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130. évfolyam

Különszám II.

Hungarian Veterinary Journal

Alapította ­ Established by

Dr. NÁDASKAY Béla

1878

Vol. 130. Supplement II. (Oral and Poster Abstracts) FÔSZERKESZTÔ ­ EDITOR-IN-CHIEF Dr. VISNYEI László SZERKESZTÔBIZOTTSÁG ­ EDITORIAL BOARD Dr. Beregi Attila, Dr. Búza László, Dr. Fodor László, Dr. Gál János, Dr. Huszenicza Gyula, Dr. Karsai Ferenc, Dr. Kégl Tamás, Dr. Kovács Katalin, Dr. Laczay Péter, Dr. Nagy Béla, Dr. Ózsvári László, Dr. Sályi Gábor, Dr. Solti László, Dr. Tanyi János, Dr. Tekes Lajos, Dr. Tóth József, Dr. Varga János, Dr. Vetési Ferenc, Dr. Visnyei László (elnök), Dr. Vörös Károly Editors: Dr. Fábián Tiborné, Szabó Judit, Verseci Miklós SZERKESZTÔSÉG ­ EDITORIAL OFFICE H-1078 Budapest, István u. 2. Hungary Levélcím: 1400 Budapest 7. Pf. 2. Telefon: (36-1) 34-13-023 (36-1) 47-84-100/8961,8960, 8962 Telefax: (36-1) 34-13-023 Internet: http://www.univet.hu/mal E-mail: [email protected] KIADÓ ­ PUBLISHER Magyar Mezôgazdaság Kft. H-1141 Budapest, Mirtusz u. 2. Telefon: (36-1)-470-0411, 273-2301, 273-2309 Telefax: (36-1)-470-0410 Felelôs kiadó: Hájos László ügyvezetô igazgató Ügyvezetô igazgató: Sári Enikô Marketingvezetô: Papp János All right reserved. Any information from the journal shall be used only with reference to Magyar Állatorvosok Lapja (Hungarian Veterinary Journal). Minden jog fenntartva. A lapból értesüléseket átvenni csak a Magyar Állatorvosok Lapjára való hivatkozással lehet. A hirdetések és egyéb reklámkiadványok tartalmáért a kiadó felelôsséget nem vállal. Nyomdai elôkészítés Lucavi Studió Bt. H-1131 Budapest, Reitter Ferenc u. 177. Telefon/fax: (36-1)-237-0582 Nyomás OLÁH NYOMDAIPARI KFT. H-1211 Budapest, Központi út 69­71. (Csepel Mûvek) Felelôs vezetô: a nyomda ügyvezetô igazgatója INDEX: 25531 HU ISSN 0025-004X

Contents

Nutrition and Metabolic Disorders Poster Abstracts Mastitis / Udder Health and Milk Quality Poster Abstracts Infectious and Zoonotic Diseases (Public Health) Poster Abstracts Sheep, Goat and Buffaloes Sheep Poster Abstracts Goat Poster Abstracts Buffaloes Poster Abstracts Camelids Poster Abstracts Herd Health Management and Epidemiology Poster Abstracts Reproduction and Biotechnology Poster Abstracts Lameness Poster Abstracts Calf Physiology, Managemenet and Diseases Poster Abstracts Animal Behaviour and Welfare Poster Abstracts Genetics and Breeding (Crossbreeding) Poster Abstracts Good Veterinary Practice (Antibiotic Resistance) Poster Abstracts Surgery, Anaesthesia, Pain Management and Diagnostic Imaging Poster Abstracts Food Safety Poster Abstracts Parasitology Poster Abstracts Global Warming Poster Abstracts Animal Hygiene, Housing and Biosecurity ­ Poster Abstracts Miscellaneous Poster Abstracts WORKSHOP Reproduction Workshop Perinatal Mortality Workshop Mycoplasma Workshop Abortion Workshop IBR Workshop Poster Abstracts Education Workshop Poster Abstracts Genetics Diseases Workshop 3 15 44 53 74 83 111 117 135 141 144 147 149 161 181 189 217 221 223 229 246 248 249 250 252 254 255 259 268 270 273 278 289 290 291 293 300 314 314 319 321 322 323 326 327 329 330 Contents 1

PREFACE

The World Buiatrics Congress (WBC) has a long tradition. Following the initiative meeting held in Hannover (Germany) in 1960, the World Association for Buiatrics was founded at the 2nd International Congress on Cattle Diseases in Vienna in May, 1962. From this time onwards the Biennial World Buiatrics Congresses were held in different countries of the world, involving all continents. It is a great honour for Hungary to organise the 25th Jubilee Congress of the World Association for Buiatrics particularly because this is the first time the WBC is held in a Middle European country. Since the foundation of the world association there have been tremendous developments in the practice of bovine medicine and greater emphasis was laid on issues like global control of infectious diseases, metabolic disorders caused by malnutrition, herd health management, animal welfare, food safety, environmental protection ­ mentioning only the most important ones. It's originating from our human nature that we often tend to overemphasize the importance of one of the above mentioned areas at the expense of the others, forgetting to maintain the healthy balance between them. This attitude appears in the relation of individual bovine medicine and bovine herd health management. Our profession requires to treat animals individually and to protect their life also on a herd level. We have to find the reasonable balance between individual treatments and herd-level preventive measures. We do hope that the 25th Jubilee Congress will help us to find that in our practice. The goal of the World Association for Buiatrics is to disseminate scientific and practical knowledge on ruminant medicine, management, genetics, production and disease control. The congress will serve as a platform for scientists from universities and research institutes, for practitioners working with dairy and beef cattle and other ruminants, consultants, residents, post-docs, PhD and graduate students of veterinary medicine and animal science in order to provide continuing education for cattle veterinarians worldwide every two years. We are very proud to mention that this year an outstanding number of 1377 abstracts have been submitted for the XXV Jubilee World Buiatrics Congress. Until now, the highest number of abstracts (a bit over 1,000) was submitted for the Nice congress in 2006. At this year's Budapest congress 408 of 1,377 abstracts have been selected for oral presentations by our international scientific committee. At the same time the majority of the poster presentations (n=860) have been accepted by our national organising committee. In cases of poster abstracts there was a very important selection criteria namely to give possibilities to the author(s) from all over the world to take part in the congress, to experience and practice the mode and level of presentations and to get more familiar with the requirements. The oral and poster presentations will be published in the Supplement II of the Hungarian Veterinary Journal (Magyar Állatorvosok Lapja) founded by Prof. B. Nádaskay in 1878.

Prof. Ottó Szenci

President of the Organising Committee

Prof. Endre Brydl

President of the Scientific Committee

Notice to Readers

Oral and poster abstracts published in the Hungarian Veterinary Journal are not peer-reviewed or referred. All statements, opinions and conclusions contained in abstracts in the Hungarian Veterinary Journal are there of the author(s), and are not necessarily those of the National and International Scientific Committees.

Nutrition and Metabolic Disorders

2 Polyunsatured Fatty Acid in Dairy Cows: a Case of Functional Feed (NAT3) G. Gabaldo , A. De Palma , A. Fusari , M. Pizzicara , 3 2 S. Tinelli , A. Ubaldi 1 Original Process, Cholet, France 2 University of Parma, Department of Animal Health, Parma, Italy 3 Veterinary Practitioner, Verona, Italy Omega 3 fatty acids can cross the rumen wall not being saturated by saturates. In this mechanism, DHA plays a fundamental rule conditioning the rumen flora to convert C18:2 fatty acids in C18:1 (trans vaccenic acid). This switch in the fatty acids production produces an increase in CLA and DHA milk fat fractions other than other types of unsaturated fatty acids specific for dairy milk, that ameliorates the nutritional characteristics of the milk for humans. Experimental study was performed utilising commercial product (FOG3) containing an association of olyunsatured Fatty Acid (PUFAs) from vegetable-origin like Linolenic Acid and from animal-origin like E.P.A. and D.H.A. (NAT), associated with antioxidant molecules (Vitamin E and Selenium) and stimulating factors of the rumen fauna (FAP). The trial involved totally 44 heads of dairy cows by 5 farms, one in the province of Verona (10 treatments and 10 controls), where only the zoometric and milk quality indices were tested and 4 farms in the province of Bari, where metabolic, endocrinological, fertility, zoometric and milk quality indices were tested in 6 cows each farm (3 cases and 3 controls each). Biochemical analysis performed stressed that treatments involved in this study influenced particularly parameters correlated to lipid status. The study confirms that omega 3 fatty acids influenced lipoproteic fractions (LDL and HDL) and total cholesterol levels that can be use to point out the necessity to administer omega 3 fatty acids. In treated and control subjects, plasma progesterone levels were measured. Reproduction performance tests showed a good correlation between analytical data and the levels of progesterone and the proof of pregnancy. In the milk collected by treated cows we noted a significant increase in CLA (C18:2) +109,34%, DHA (C22:6) + 223,80%, and palmitoleic acid (C16:1) +18.57%, while a decrease in ratios of EPA (C20 :5) -57.14%, ALA (C18:3) -20.79% and stearic acid (C18:0) 26.09% was observed. Key words: PUFAs, omega 3, DHA, CLA, EPA, transition cow, fertility 3 Three Methods of Supplementing Selenium to Intensively Grazed Beef Cattle J. Maas University of California, Veterinary Medicine Extension, Davis, United States Objectives: To compare the efficacy of supplementing selenium (Se) to grazing beef cattle by injection, intra-ruminal bolus, and Se fertilization of pastures. Materials and Methods: Weaned beef heifers (16 per group) intensively grazing irrigated pastures were studied using 4 treatments in 4 replicates for 2 years and the trials lasted 120 days. Group 1: one intra-ruminal osmotic pump bolus supplying 3 mg Se for 120 days; group 2: 2.5 mg Se/45.4 kg subcutaneously; group 3: pastures fertilized with 10g Se per hectare (sodium selenate); and, group 4: Se deficient heifers with no Se supplements. The Se fertilized pastures were treated prior to the grazing season and all other treatments were administered on day 0. Blood samples for Se analysis were collected at days 0, 30, 60, 90, and 120 of each trial period. Selenium was analyzed by ICP spectroscopy and data was analyzed by ANOVA methods. Results: Blood Se concentration (ppb; µg/l) data is listed as values for day 0, 30, 60, 90, and 120, respectively. Year 1. Group 1: 13 ± 4, 94 ± 28, 130 ± 19, 184 ± 15, and 234 ± 27. Group 2: 13 ± 5, 30 ± 6, 20 ± 8, 15 ± 3, and 8 ± 6. Group 3: 13 ± 5, 113±28, 135 ± 23, 159 ± 20, and 148 ± 16. Group 4: 12 ± 4, 14 ± 5, 10 ± 6, 5 ± 1, and 4 ± 4. Year 2. Group 1: 16 ± 3, 88 ± 7, 126 ± 13, 172 ± 19, and 200 ± 19. Group 2: 20 ± 4, 31 ± 4, 27 ± 3, 18 ± 2, and 10 ± 3. Group 3: 17 ± 3, 46 ± 11, 56 ±9, 46 ± 8, and 40 ± 8. Group 4: 22 ± 8, 13 ± 6, 9 ± 2, 9 ± 4, and 7 ± 4. The blood Se concentrations were significantly (P < 0.0001) increased above controls (group 4) for groups 1 and 3 in both years and at all times after day 0. Group 2 blood Se

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concentrations were increased (P < 0.05) above control cattle only at day 30 (year 1 and 2) or day 60 (year 2 only). Conclusions: The use of intra-ruminal Se boluses or Se fertilization of pastures resulted in superior increases in blood Se concentrations in cattle intensively grazing irrigated pastures vs. cattle receiving a single Se injection. The bolus achieved more consistent increases in blood Se concentrations. 4 Clinicopathological Evaluation of Downer Dairy Cows with Fatty Liver P. Kalaitzakis , N. Panousis , N. Roubies , N. Giadinis , 3 1 E. Kaldrymidou ,H. Karatzias 1 School of Veterinary Medicine, Aristotle University, Clinic of Farm Animals, Thessaloniki, Greece 2 School of Veterinary Medicine, Aristotle University, Laboratory of Clinical Diagnosis and Clinical Pathology, Thessaloniki, Greece 3 School of Veterinary Medicine, Aristotle University, Laboratory of Pathology, Thessaloniki, Greece Objective: To evaluate the severity of fatty liver and macro mineral status in downer dairy cows and asses the usefulness of determination of serum ornithine carbamoyl transferase (OCT) activity, bile acids concentration, and other variables for prognosis. Materials and Methods: 36 Holstein downer cows were used. Blood and liver biopsy specimens were obtained as soon as possible after cows became recumbent and before any treatments were administered. Liver tissue was examined histologically and classified according to the severity of fatty liver. Serum activities of liver-derived enzymes (OCT, GDH, SDH and also AST, ALT, GGT, ALP) and concentrations of total lipids, triglycerides, bile acids, glucose, ,-hydroxybutyrate, acetoacetic acid, total bilirubin, nonesterified fatty acids, cholesterol and macro minerals (Ca, Mg, K, Na, P) were determined. Results: The majority of the downer cows had fatty liver infiltration; 44% were classified as moderate and 44% were classified as severe fatty liver infiltration. Survival of the downer cows with moderate and severe fatty liver was poor. Serum activities of OCT and GDH, and NEFA/cholesterol ratio were indicators of fatty liver. Serum bile acids concentration was not an accurate indicator of fatty liver. Conclusions: Fatty liver is quite common in downer cows. Serum activities of OCT and GDH are useful indicators of fatty liver. NEFA/cholesterol ratio is also recommended for diagnosis of fatty liver in downer dairy cows, whereas determination of bile acids concentration is not. Key words: fatty liver, OCT, GDH, downer cows 5 Vitamin E and Oxidative Damage in Heifers R. Bouwstra , R. Goselink , P. Dobbelaar , M. Nielen , 2 1 J. Newbold , T. Werven 1 Faculty of Veterinary Medicine, Utrecht University, Animal Health Department, Utrecht, Netherlands 2 Provimi Research and Innovation Centre, Brussels, Belgium The question is whether measurements of oxidative and vitamin E status in blood of a periparturient cow are representative for the total body, given that blood concentrations of vitamin E and oxidative stress products change around this period. This study investigated the relationship between oxidative damage and the effect of vitamin E supplementation in blood, milk and liver tissue in sixteen periparturient heifers. The daily vitamin E intake of the vitamin E supplemented Holstein Friesian heifers (n=8) was 3000 international units and started two months before calving; the control heifers (n=8) were not supplemented. Oxidative damage was determined on the basis of malondialdehyde (MDA) concentrations. Blood was sampled nine times before calving, on calving day and twice after calving. Liver biopsies were taken at week -5, -1, and 2 relative to calving day. Milk was obtained from all heifers immediately after calving, the first two milkings and on days 3, 7 and 14 at 6 am. Serum and liver tissue were analyzed for vitamin E, cholesterol and MDA and milk samples were analyzed for vitamin E, MDA, fat, protein and somatic cell count. The results show that vitamin E supplements increased both absolute vitamin E concentrations and the ratio of vitamin E to cholesterol in

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Nutrition and Metabolic Disorders 3

blood and liver tissue. Absolute vitamin E concentration in milk tended to be higher in supplemented cows. Based on the increased MDA blood concentrations at calving, it seems that dairy heifers experience oxidative stress. The effect of vitamin E on MDA differs between the blood, liver and mammary gland. Vitamin E supplementation could not prevent the rise in blood MDA at calving, but the significantly lower MDA blood concentrations of supplemented cows in the two weeks after calving suggest vitamin E has a role in recovery from parturitionrelated oxidative stress. Vitamin E supplementation reduced oxidative damage in liver, while no obvious effect was found on milk MDA concentrations. A strong relationship was found between blood and liver tissue vitamin E and the ratio of vitamin E to cholesterol. MDA concentrations in blood and milk were also strongly related. The results show that the relationship between oxidative damage and vitamin E differs within blood, liver tissue and milk. This implies that oxidative and vitamin E status calculated on the basis of blood values alone should be interpreted with caution and cannot be extrapolated to the whole animal. Key words: vitamin E, oxidative damage, dairy heifer 34 Serum Non-Esterified Fatty Acid and Beta-Hydroxybutyrate in the Transition Period and their Associations with Disease in Dairy Cows M. Carson , S. LeBlanc , S. Godden , M. Capel , 4 5 1 1 M. Overton , J. Santos , K. Leslie , T. Duffield 1 Ontario Veterinary College, Population Medicine, Guelph, Canada 2 University of Minnesota, Population Medicine, St. Paul, USA 3 Perry Veterinary Clinic, Perry, USA 4 University of Georgia, College of Veterinary Medicine, Athens, USA 5 University of California, College of Veterinary Medicine, Davis, USA Introduction: Serum non-esterified fatty acid (NEFA) and betahydroxy-butyrate (BHB) concentrations provide insight into the metabolic health of transition dairy cows. The objective of this study was to further characterize the relationship of serum NEFA and BHB concentrations in the transition period with clinical disease in dairy cows across different regions of North America. Methods: A field study was conducted using 2403 Holstein cows in 56 dairy herds in 4 regions of Canada and the United States. Once weekly, after the morning feeding, blood was collected from cows in the week before their expected calving date, and again from the same cows in weeks 1, 2, and 3 postpartum. Serum was stored at -20°C within 8 hours of collection. NEFA and BHB were measured using a Hitachi 911 auto-analyzer. Results: Results were available from 2149 (confirm) cows. Considered alone and assuming equal weight on sensitivity and specificity, the optimal cut-points for prediction of LDA were, prepartum: NEFA < 0.5 mEq/L; week 1 postpartum: NEFA 1.0; or week 1 or 2 postpartum: BHB 1400 mol/L (i.e. subclinical ketosis (SCK)). The 23% of cows with NEFA 0.5 in week -1 were 2.8 times more likely to subsequently have LDA than cows below this cut-point. The 20.5% of cows with NEFA > 1.0 in week +1 were 4.6 times more likely to develop LDA. The prevalence of SCK and relative risk for affected cows to develop LDA were 14% and 4.4 in week +1, and 16% and 3.6 in week +2. Cows that did not have elevated NEFA pre- or postpartum or SCK had the lowest risk of LDA (0.8%). Conversely, cows that had both high postpartum NEFA and SCK, with or without high NEFA prepartum, had a 15 to 16% incidence of LDA. The magnitude, timing, and duration of elevations in NEFA and BHB combine in a complex manner to predict the risk of LDA. Among the 53% of cows with NEFA > 0.3 mEq/L in week -1, there was a significant (P = 0.005) but modest increase in risk of RP (9% vs. 6%; RR = 1.6). Conclusion: These data confirm the associations of NEFA and BHB with health and support their use as tools for monitoring or investigation of transition dairy cows. However, used alone, the positive predictive value of these associations is low, which is expected given the multifactorial nature of both RP and LDA. 35 The Effect of Organic and Inorganic Dietary Selenium Sources on Selenium Levels in Blood, Colostrum and Milk and Metabolic Profile in Dairy Cows D. Kumprechtova , J. Illek , N. Ballet 1 Veterinary and Pharmaceutical University Brno, Clinic of Ruminant Diseases, Brno, Czech Republic 2 Lesaffre Feed Additives, R&D, Lille, France

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Objective: The aim was to compare effects of two levels of organic selenium (Se) sources and an inorganic selenium source on blood, colostrum and milk Se levels, blood chemistry parameters in dairy cows under field conditions. Material and methods: 36 Holstein cows from 5 weeks (wk) antepartum (a.p.) to 12 weeks postpartum (p.p.). Treatments (n=9): 1. NC no Se added, 2. PC - 0.3 ppm Se (sodium selenite), S1 0.3 ppm Se (Se yeast), 3. S2 - 5 ppm Se (Se yeast). At 5 and 1 wk a.p., at the calving, and 4, 8, 12 wk p.p., blood samples were analysed for Se, GSH-Px, TP, albumin, urea, billirubin, AST, GMT, CK, vit. E. Colostrum and milk was collected at the calving and 4, 8 and 12 wk p.p. and analysed for Se. Results: Initial Se status in all the cows was marginal. NC showed suboptimal Se levels for whole period. S1 and S2 showed significantly (P < 0.01) higher values than NC and PC. Table 1. Se levels in whole blood Treatment PC (0,3ppm Se) Se (µg/l) Se (µg/l) Mean S.D. Mean S.D. 69,16 68,2a 67,0a 65,7a 68,9a 73,52a 5,0 70,3 5,7 6,2 90,8a 7,0 5,6 93.6b 7,9 5,7 86,1b 33,7 4,3 105,1b 7,0 5,03 110,0b 15,9 NC S1 (0,3ppm Se) Se (µg/l) Mean S.D. 77,7 97,7b 109,1c 129,1c 128,6c 129,22c 7,3 7,7 7,9 7,7 11,2 9,58 S2 (5 ppm Se) Se (µg/l) Mean S.D. 70,6 6,7 155,0c 22,1 152,2d 16,5 170,8d 19,8 184,4d 9,9 193,49d 15,32 P

5 wk a.p. 1 wk a.p. Calving 4 wk p.p. 8 wk p.p 12 wk p.p.

< 0,01 < 0,01 < 0,01 < 0,01 < 0,01

Colostrum Se levels were higher (P < 0.01) in S1 (128.17 µg/l) and S2 (155.58 µg/l) than in NC (94.68 µg/l) and PC (99.3 µg/l).Overall mean milk Se levels in S1 (20.4 µg/l) and S2 (50.0 µg/l) were higher (P < 0.01) than in NC (10.9 µg/l) and PC (16.6 µg/l). GSH-Px activity in whole blood in S1 and S2 was higher than in NC and PC (P < 0.01). CK activity decreased with higher org. Se doses. TP, albumin, urea, billirubin, AST, GMT, and vit. E were within the physiological ranges, non-significant differences. Conclusions: Se levels in whole blood, colostrum and milk, and GSHPx activity were significantly increased by organic Se supplementation. The response was dose dependent and consistent. Selenium from organic source was more bio available than selenium from sodium selenite. Dietary Se at a tolerance dose (5 ppm) had no detrimental effect on the blood chemistry parameters analysed. Project no. 1G46086 (NAZV). Key words: dairy cows, organic selenium, blood, milk, metabolic profile 36 The Influence of Subclinical Hypocalcemia on Production and Reproduction Parameters in Israeli Dairy Herds Ch. Gild, Ni. Alpert, Mi. Van-Straten Hachaklait, Ceasaria, Israel Introduction: Nearly all mature cows experience some degree of Hypocalcemia (HypoC) during the 1st hours post calving. Most cows cope with these low Ca concentrations without apparent clinical signs however, in some cases Ca concentrations decline to levels that disrupt neuromuscular function, resulting in the clinical syndrome known as milk fever. It has been established that cows suffering from clinical milk fever are susceptible to a variety of secondary problems. To the best of our knowledge, no one has evaluated the impact of subclinical HypoC on (re)production parameters. Most trials compare the affect of different diets on the occurrence of HypoC (clinical and subclinical). Big emphasis is given to the subclinical condition although its adverse effects have not been proven. Our objective was to investigate the association between subclinical HypoC and (re)production parameters in Israeli dairy herds. Materials and methods: Blood samples for corrected calcium were collected from 634 mature cows from 5 farms. Disease and reproduction data were recorded for 180 days. Milk and milk solids data originated from monthly milk recordings. Subclinical HypoC was defined as serum corrected Ca below 7.5 mg/dl 8-20 hours postpartum in the absence of milk fever. Associations between subclinical HypoC and retained placenta, metritis, ketosis, stillbirth and uterus prolapse were investigated using chi-square tests. Time to conception data was analyzed using the Kaplan-Meier method. The relationship between subclinical HypoC and milk production was investigated with a marginal model for repeated measurements.

4 XXV. Jubilee World Buiatrics Congress 2008

Results: Occurrence of clinical and subclinical HypoC was 2.5%, 18.8% respectively. Compared with normocalcemic cows, the subclinical hypocalcemic cows produced 3.2, 2.7 and 1.9 kg more milk in the first three milk recordings, respectively. Sub clinical hypocalcmic cows did not have an increased risk for post parturient diseases nor compromised reproduction parameters in comparison to normocalcemic cows. Conclusions: We could not demonstrate a negative impact of the subclinical condition on (re)production parameters. We hypothesize that although Ca blood levels decline to sub normal values, the short time period and physiological adaptations enable cows to undergo this disturbance without apparent (re)production loss. Our results suggest that anionic diets to prevent subclinical HypoC are not warranted in Israel as the incidence of milk fever is relatively low. 37 Fatty Liver and Serum Concentrations of Insulin and Glucose in Dairy Cows D. Kirovsci , H. Samanc , M. Jovanovic , N. Fratric , 4 2 2 D. Gvozdis , I. Vujanac , B. Dimitrijevic 1 Faculty of Veterinary Medicine Belgrade, Department of Physiology, Belgrade, Serbia 2 Faculty of Veterinary Medicine Belgrade, Department of Internal Medicine, Belgrade, Serbia 3 Faculty of Veterinary Medicine Belgrade, Department of Pathology, Belgrade, Serbia 4 Faculty of Veterinary Medicine Belgrade, Department of Pathophysiology, Belgrade, Serbia The aim of this study was to determine the relationship between fatty liver and serum concentrations of insulin and glucose in dairy cows during peripartal period. Twenty-one Holstein dairy cows were used in this study. The cows were classified into three groups (n=7 per group) according to the histopathologic findings in hepatocytes: Group 1: cows with mild fatty liver, Group 2: cows with moderate fatty liver and Group 3: cows with severe fatty liver. Liver samples for histopathologic analysis were collected from cows by percutaneous needle biopsy on day 10 postpartum. Liver samples were fixed in paraformaldehyde solution, cut and stained with hematoxylin and eosin. Blood serum insulin and glucose concentrations were determinated in blood samples obtained from v. jugularis at the middle of dry period (30 to 35 days prior parturition), 5 to 10 days prior parturition and 10 days after parturition. Insulin concentration was measured by radioimmunoassay. Glucose concentration was measured spectrophotometrically. Insulin concentration was significantly lower at day 10 postpartum than 5 to 10 days prior to partus (p < 0.05) and 30 to 35 days prior to partus (p < 0.01) in all three groups of cows. No significant differences in serum insulin concentrations were present among groups of cows in all examined time periods. In Group 3 serum glucose concentration was significantly lower 10 days postpartum than 5 to 10 days prior to parturition (p < 0.001) and 30 to 35 days prior to parturition (p < 0.01). In Group 1 significant decrease of glucose concentration was determinated from 5 to 10 days prior to parturition to 10 days postpartum (p < 0.05). There was no significant decrease in serum glucose concentration between examinated time periods in Group 2. The serum glucose concentration in Group 3 was significantly lower then in Groups 1 and 2 both 30 to 35 days prior to parturition (p < 0.05) and 10 days postpartum. Our results indicate that cows with severe fatty liver have low glucose levels, probably because of impaired gluconeogenesis. The level of glycemia was not correlated with serum insulin concentrations which indicate that glucose concentration is under complex endocrine control during peripartal period. Key words: dairy cows, fatty liver, insulin, glucose 38 Metabolic Syndrome in Cows: TNF-1 and Fat Metabolism in Heifers with Different Body Condition (BCS) at Parturition M. Fürll, N. Jatzke, K. Bauer, I. Steinhöfel, J. Gottschalk, U. Sack, A. Einspanier University of Leipzig, Internal Medicine, Leipzig, Germany Objectives: Recent studies in humans and other animals have shown that fat tissue is not a simple energy storage organ, but releases a lot of molecules which exerts important endocrine and immune functions. Adipocytes secrete numerous bioactive substances, including hormones, growth factors and cytokines. Abdominal fat accumulation has been shown to play crucial roles in the development of the human metabolic syndrome. This study describes the effect of body condition on metabolic profiles,

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hormones and cytokines in Holstein cows during the peripartal period. Maybe there are some similarities with the components of the human metabolic syndrome. Material and methods: 15 cattle were fed with a high energetic diet (hED) until they became pregnant, 15 heifers were chosen because of their high back fat thickness (hBFTh), and 15 cattle were fed in optimum conditions (oED). Blood samples were taken 1 week before parturition (w a.p.), 3 days after parturition (d p.p.) and 1 month p.p. to determine metabolic, hormonal as well as immunological parameters. Results: The FFA, insulin and TNF1 concentration (medians) are shown in the table. FFA (mmol/l) 1 w 3 d 3­5w a.p. p.p. p.p. hED oED hBFTh 160 610 200 190 450 275 270 612 230 insulin (nmol/l) 1 w 3 d 3­5 w a.p. p.p. p.p. 0,16 0,07 0,12 0,13 0,09 0,09 0,16 0,16 0,13 TNF1 (ng/ml) 1 w 3 d 3­5 w a.p. p.p. p.p. 27 10 10 33 22 10 29 10 34

In high back fat thickness, the cows insulin concentrations were significant higher at the third d p.p. than in the other groups; the high back thick fatness cows showed the highest glucose concentrations during the whole sample period. We found a positive correlation between TNF-1 and FFA concentrations in cattle with hBFTh (r=0.63). In hBFTh heifers and in cattle the concentrations of IGF-I were significant lower at day 3 p.p. Not only a positive correlation existed among the IGF-I and the FFA concentrations (r=-0.52) but also a negative correlation among IGF-I and haptoglobin in all groups (r=-0.61). Conclusion: In conclusion the study shows that changes of energy fat metabolism - including TNF-1 from adipose tissue is comparable with the metabolic syndrome in humans and may be directly involved in the pathogenesis of many postpartum disturbances. Key words: metabolism, body condition, metabolic syndrome, heifers 39 Influence of Gluconeogenesis on Metabolic Status and Appereance of Oxydative Stress in Peripartal Cows B. Dimitrijevic , S. Borozan 1 Faculty of Veterinary Medicine Belgrade, Clinic for Ruminants and Pigs, Belgrade, Serbia 2 Faculty of Veterinary Medicine Belgrade, Chemistry, Belgrade, Serbia In ketotic condition of cows, due to presence of ketone bodies, appearance of reactive oxygen species (ROS) and reactive nitrogen species (RNS) might occur. Durable presence of ketone bodies may cause cells susceptible to irreversible damage, what is reflected in increased lipid peroxidation through damaging of cell membranes, as well as increased level of lactate dehydrogenase (LDH) enzyme in extra cellular space. The objective of this study was to examine impact of keton bodies on ROS and RNS production, levels of lipid and protein damage in different level concentration of keton bodies in sample of cows blood. Research was conducted on sample of Holstein Friesian cows (n=15), during few first days after calving, and presence of keton bodies was proven. Control group was made up of cows with negative test on ketone bodies. Production of ROS was examined through activity of superoxide dismutase (SOD) and catalase (CAT) enzymes in erythrocytes, whereas production of RNS was monitored through the contents of nitrites in blood plasma of the examined animals with ELISA. The level of lipid peroxydation was monitored through malondialdehyde (MDA). The effect of free radicals provoked by keton bodies on hepatocytes was monitored through determination of izoenzymes distribution of LDH (LDH1-LDH5) by electroforesis. Research results showed significantly increase in SOD activity (P<0,001). Activity of CAT enzyme was significantly increased and was 95,59±17,46 U/g Hb, in control group 63,82±5,44 U/g Hb. The contents of nitrites in ketotic cows was 7,8±1,2 µM/L, in control group 4,8±0,50 µM/L. Distribution of LDH izoenzymes was LDH1 37,96±2,59%, LDH2 22,99±2,82%, LDH3 18,93±1,85%, LDH4 3,19±0,20%, LDH5 15,78 ±6,28% and in control group: LDH1 45±3,01%, LDH2 35±2,62%, LDH3 22±1,56%, LDH4 4,07±0,5% and LDH5 1,31±0,7%. The level of (MDA) was 44,96±6,21 nM/gHb, and 10,27±2,14 nM/gHb in control group. The effects of keton bodies results in increased production of SOD, hydrogen peroxide and RNS, depending on keton bodies concentration. Joint impact of ROS and

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RNS causes increased lipid peroxidation and accelerated proteolysis what is reflected in increased presence of LDH5, as hepatocytes destruction. Key words: cow, keton bodies, oxydative stress, ROS, RNS 70 Effect of BCS Change during the Dry Period on the Performance of Dairy Cows T. Van Werven, D. Speksnijder, J. Papen, M. Nielen Faculty of Veterinary Medicine, Utrecht University, Farm Animal Health, Utrecht, Netherlands This abstract shows the preliminary results of an on going blind trial on the effect of orally supplemented vitamin E during the dry period on the performance of dairy cows. Most studies are interested in either calving BCS or amount of BCS loss postpartum. This study evaluates the effect of BCS change (loss or gain) during the dry period on the postpartum performance of the cow. Five dairy farms were selected to participate in a blind field trial to investigate the effect of orally supplemented vit E. From each farm approximately 60 dry cows are enrolled into the study 6-8 weeks prior to calving date. Dry cows are divided in two groups (A and B), one group is fed 135 IU and the other group 3000 IU of vit E during the dry period. Body condition score is measured at a 5-point scale once a week during the dry period, at calving and at the first milk test-day. Milk test-day records are collected for fat, protein and SCC. Data on retained placenta are collected and defined as failure to expulse the placenta within 12 hours pp. Change of body condition score during the dry period is calculated by regression analysis which provides cows with an increasing or decreasing BCS during the dry period. Cows with an unchanged BCS during the dry period are included in the group of cows with increased BCS. Until so far milk test-day records of 94 cows are presented. At the beginning of next March all cows will be included. From these preliminary results some interesting trends are demonstrated. There seems to be a difference between the two vitamin E groups (blind). Cows in group A show more often an significant increase of BCS during the dry period than cows in groups B, corrected for herd effects. Cows with average BCS<3 during the dry period showed an higher risk on retained placenta than cows with BSC>3, corrected for increasing or decreasing slope during the dry period. Preliminary trends indicate that cows with a BCS loss during the dry period are less at risk for a fat/protein ratio > 1.4 than cows that gained BCS, corrected for days in milk. These results indicate that cows with BCS loss in the dry period might have a more efficient energy metabolism after parturition and therefore be less at risk for metabolic diseases like ketosis after parturition. Those cows might experience fewer metabolic changes during the transition from late gestation to early lactation. The possible effect of vit E on BCS needs further research. Key words: body condition score, dry period, dairy cows, vitamin E 71 Metabolic Syndrome in Cows: TNF-1 Concentrations before and after Parturition in Healthy and Ill Cows B. Fürll , G. Hädriich , M. Fuerll 1 University of Leipzig, Department of Veterinary Physiology, Leipzig, Germany 2 University of Leipzig, Department of Internal Medicine, Leipzig, Germany Objectives: Today, cattle health is especially disturbed because of a strained metabolism. Cytokines of the visceral fat depot, TNF-1 for example, can affect the metabolism negatively. Therefore we investigated cows with the aims: a) Are there any differences between the TNF-1 concentrations in healthy and ill cows around parturition? b) Can those differences be noted before parturition already? c) Are there any TNF-1 changes specific for certain diseases? Material and methods: 20 healthy cows as well 103 cows with various diseases (table) were examined 10 days before parturition (d a.p.) and 3 d after parturition (p.p.). Blood samples were analysed byHitachi 912, TNF-1 by an ELISA. Results: The TNF-1 concentrations a.p. and p.p. of healthy (medians 36:37 ng/ml) and ill (140:55 ng/ml) cows differed significantly. Table: TNF-1 (ng/ml), FFA (mmol/l) and haptoglobin (Hp, g/l) concentrations (medians) in healthy cows and cows with different diseases p.p. (*p=0.05)

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d p.p heal- ret. endo- still- ova- twin mas- Dislothy pla- met- birth rian birth titis catio centa ritis cysts abomasi (DA) TNF-1 -10 3 FFA -10 3 Hp -10 3 35,6 37,3 80 570 0.20 0.74 90,7* 117,5* 115 870* 0.09 1.51* 50,4 47,8 180* 872* 0.33* 1.52* 26,8 19,9 150* 790* 0.09 1.45* 23,9 33,4 10,0 19,8 115 310 875* 875* 0.11 0.40* 1.22* 1.86* 76,4 50,5 210* 620 0,08 1,01 55,9* 10 85 1270* 0,28 1,04

The TNF-1 concentrations correlate a.p. with the back fat thickness (p>0.05); they are at their highest (p<0.05) in cows with ret. placenta, followed by cows with mastitis and DA (p<0.05). The FFA concentrations are also especially increased (p<0.05) in cows with mastitis and DA. Increases in Hp concentrations have started to rise at approx. 10 d a.p. and vanished again 4 weeks p.p. Hp concentrations also differ in their behaviour from TNF-1 concentration; they do not correlate with each other. However, Hp correlates with FFA r = 0.84, i.e. FFA and Hp have the same dynamics. Conclusion: TNFa concentrations are increased a.p. and p.p. in sick cows; they correlate positively with the BFT. In cows with ret. placenta, mastitis and abomasal displacement, the TNF-1 correlates positively a.p. with the FFA. Hp concentrations are risen in calving and puerperal disorders. The study supports the hypotheses that during early lactation, adipositas influences the diseases via cytokines. Key words: cattle, metabolic syndrome, cytokines. TNF-1, pp diseases 72 Prevalence of Subclinical Metabolic Disease in Transition Dairy Cows on Large Dairies in Peru J. Bryk-Lucy , D. Nydam , A. Delgado Castro , 3 2 A. Gonzales Zariquiey , M. Montenegro Vega 1 Cornell University, Population Medicine, Ithaca, United States 2 Universidad Nacional Mayor de San Marcos, Facultad De Medicina Veterinaria, Lima, Peru 3 Universidad Nacional Mayor de San Marcos, Dean of Veterinary College, Lima, Peru Blood concentrations of non-esterified fatty acids (NEFA) and betahydroxybutyrate (BHB) and pH of rumen fluid can be monitored to indicate cows or herds at risk for excessive metabolic disease and related economic losses. Very little information is available about the risk of metabolic disease in the transition cows near Lima, Peru. The objective of this study was to determine the prevalence of excessive negative energy balance (NEB), subclinical ketosis (SCK), and subacute ruminal acidosis (SARA) in transition cows in dairy herds located within a 160 km radius of this rapidly growing dairy center. Blood from 128 Holstein cows from 16 herds 2-16 days prepartum and 166 Holstein cows from 19 herds 3-21 days postpartum was tested for NEFA and BHB concentrations, respectively. NEFA and BHB concentrations were analyzed using the DVM NEFA Meter and Abbott Precision Xtra Ketone Meter, respectively. The average herd size was 350 milking cows. Rumen pH was tested in 47 Holstein cows from 9 herds using rumenocentesis. A herd-level prevalence of >15% elevated metabolic indicators was considered an alarm level for significant risk of clinical metabolic disease and economic loss. Herd-level incidence of displaced abomasum, retained placenta, and low milk production over the past 30 days was also recorded. Of the sampled herds, 87.5% had a > 15% prevalence of NEFA levels greater than the cut-off point of > 0.4 mmol/L, and the prevalence of herds with >15% subclinical ketosis (cut-off of 1.4 mmol/L) was 52.6%. The within herd prevalence of NEB and SCK ranged from 14% to 80% and 8% to 44%, respectively. The prevalence of SARA (rumen pH <5.5) was 0% in all herds sampled. Only two herds of all herds sampled reported an incident of displaced abomasum. At least one cow was considered low producing in 64% of herds with >15% elevated NEFA and in 75% of herds with >15% elevated BHB. At least one retained placenta was reported in 85% of herds with >15% elevated NEFA and in 100% of herds with >15% elevated BHB. Further modelling of the data will be discussed at the meeting. The results suggest that subclinical metabolic disease in transition cows is present in dairy herds surrounding Lima, Peru at levels that may negatively impact milk production, reproductive performance, and profitability. There is opportunity to

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improve management practices to minimize subclinical disease in transition cows and maximize profitability. Key words: metabolic disease, transition cows, subclinical ketosis, negative energy balance, Lima Peru 73 Evaluation of three Laboratory Methods Measuring NEFA in Cow's Blood T. Gaal, P. Ribiczey Szent István University Faculty of Veterinary Science, Department of Internal Medicine, Budapest, Hungary Non esterified fatty acids (NEFA) in the blood are good markers of negative energy balance (NEB) in dairy cows. There are several laboratory methods to assess NEFA concentration. One method is based on a colorimetric, non-enzymatic technique at room temperature (Duncombe, 1964), the other one applies colorimetric, enzymatic determination at 37 °C (Matsubara et al., 1983). Recently a portable ® photometer appeared on the market (DVM-NEFA ) using a colorimetric technique for NEFA measurement at room temperature. As laboratory methods need a well-equipped laboratory with skilled personnel, introduction of a cow-side test is more than welcome. However, widely used NEFA reference values were established by the laboratory methods, therefore critical evaluation of DVM-NEFA is recommended. The goal of this study was to compare NEFA results determined with 3 methods. On a herd of 900 HF dairy cows blood samples were collected from dry cows (n=17) and fresh cows (n= 14). Blood serum NEFA was measured immediately by 3 methods. For the non-enzymatic method commercial chemicals were used while for the enzymatic method a Randox NEFA test kit was applied on an automatic chemistry analyzer. Results were analyzed by t-test and regression analysis. Pre-partum NEFA values determined by DVMNEFA, non-enzymatic and enzymatic methods were 0.258+0.200, 0.105+0.090 and 0.093+0.088 mmol/L, respectively. Post-partum these were 0.706+0.327, 0.300+0.146 and 0.293+0.179 mmol/L. Prepartum NEFA was lower than post-partum with all methods (P<0.01). Both pre- and post-partum significant differences were found between the results of DVM-NEFA and the two laboratory methods (P<0.05): DVM-NEFA results were more than 2 times higher than those completed by 2 laboratory tests. Good correlation was found both preand post-partum between DVM-NEFA and laboratory results (correlation coefficients: 0.86-0.93). As substantial differences were found between the results, establishing reference values for DVM-NEFA is recommended. The widely accepted 0.400 mmol/L NEFA as upper reference value in dairy cows was determined several years ago by laboratory tests and most probably could not be used on DVM-NEFA. Applying the 0.400 mmol/L NEFA as upper reference limit, all post-partum cows should have been qualified as a severe case of NEB based on the 0.706 +0.327 mmol/L NEFA value achieved on the DVM-NEFA instrument. Key words: NEFA, NEB, DVM-NEFA, enzymatic method 74 Non Invasive Detection of Hepatic Steatosis in Dairy Cows with Calibrated Ultrasonographic Image Analysis A. Starke , J. Thijssen , G. Weijers , A. Haudum , 1 3 2 1 K. Herzog , P. Wohlsein , C. De Korte , J. Rehage 1 University of Veterinary Medicine Hannover, Foundation, Clinic for Cattle, Hannover, Germany 2 Radboud University Nijmegen Medical Center, Clinic for Cattle, Nijmegen, Netherlands 3 University of Veterinary Medicine Hannover, Foundation, Department of Pathology, Hannover, Germany In dairy cows, rising liver fat content increases the risk of other peripartal disorders. Until now, liver biopsies have to be obtained invasively for a precise quantification of the hepatic fat content. The objective of this study was to investigate the diagnostic value of ultrasonographic image analysis for a non invasive detection of hepatic steatosis in cattle. 151 German HF dairy cows with left sided abomasal displacement were included in the study. Abomasal displacement was surgically corrected during laparotomy. Ultrasonographic images (fixed equipment settings) were obtained transcutaneously and intraoperatively. During surgery, a liver biopsy was taken (liver triglyceride content (gold standard); patho-histological examination). A custom designed software package for computer-aided ultrasound diagnosis (CAUS) was developed. The quantitative comparison of

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transcutaneous and intra-operative images enabled the correction for the attenuation effect of skin and subcutaneous fat/muscle layer on the mean echo level in the liver. The effects of the beam formation and attenuation of liver tissue on the echo level vs. depth were taken into account by estimating the mean echo level vs. depth curve of healthy livers. The residual attenuation coefficient (dB/cm) in fatty liver vs. normal liver was estimated and compensated for. In the control group (normal liver (n=10)) skin plus fat/muscle layer attenuation was 4.8 dB/cm. In transcutaneous images (n = 151), the mean liver tissue echo level correlated with the triglyceride content: r = 0.56, correlation of the residual attenuation coefficient with triglyceride content was r = 0.80. Intra-operatively, the correlation between mean liver tissue echo level and triglyceride content was r = 0.72, correlation of residual attenuation coefficient and triglyceride content: r = 0.82. The obtained results show clearly that accumulating fat leads to enhanced ultrasound attenuation in bovine liver. The developed calibrated method offers an accurate alternative for the invasive assessment of liver fat content. The authors show in this study the feasibility of computer aided ultrasound for noninvasive diagnosis of hepatic steatosis of the liver. Key words: fatty liver, ultrasonography, CAUS, liver biopsy 75 The Effect of Body Condition Score at Calving and Supplementation with Yeast on Feed intake, Blood Metabolites and Days to First Ovulation in Peri-parturient Dairy Cows R. Alibrahim, P. Duffy, L. O'Grady, M. Beltman, A. Kelly, V. Gath, M. Doherty, F. Mulligan University College Dublin, School of Agriculture, Food Science and Veterinary Medicine, Dublin, Ireland The aim of this study was to determine the effect of body condition score (BCS) at calving and supplementation with a live yeast culture 1026 (Yea-Sacc , Saccharomyces cerevisiae, CBS 493.94) beginning two weeks pre-calving on feed intake, indicators of negative energy balance and days to first ovulation in early lactation dairy cows. Forty Holstein/Friesian cows of mixed parity were randomly allocated to two groups depending on BCS approximately 60 days prior to parturition. Both groups were fed grass silage ad-libitum for the entire 60 days precalving and half of each group were supplemented with a dairy concentrate containing 2.5g of thermo-stable yeast supplement or a control concentrate for the last 14 days pre-calving and during the first 70 days of lactation. Feed intake was monitored individually in early lactation. Blood samples were collected via jugular venipuncture on the day of calving and at days 5, 15, 25 and 35 post-calving and analysed for glucose (GLS) non-esterified fatty acids (NEFA) and beta-hydroxybutyrate (BHB). Dairy cows in the high BCS group had a lower (P<0.06) feed intake than cows in the low BCS group in weeks 1 and 2 of lactation. However, there was no effect of BCS group on the average feed intake over the first 70 days of lactation. High BCS group cows tended to have higher NEFA concentrations (P=0.08) on the day of calving and higher BHB concentrations (P=0.09) at 5 days post-calving when compared with the low BCS group. Low BCS group cows had higher (P<0.01) GLS concentrations at day 25 post-calving and at day 35 post-calving (P=0.06). High BCS group cows had higher BHB concentrations at day 25 (P=0.07) and 35 (P<0.01) post-calving and had higher NEFA concentrations at day 25 (P<0.01). Cows supplemented with S. cerevisiae tended to have lower NEFA concentrations at day 25 post-calving (P<0.10). There was no effect of BCS group on days to first ovulation (31.5 vs 29.0 days), size of the first dominant follicle (17.9mm vs. 17.5mm) or the size of the first ovulatory follicle (18.6 vs. 18.1mm). Cows fed S. cerevisiae ovulated 7.5 days earlier (34.0 vs. 26.5 days) than control cows. However this difference was not significant. Low BCS group cows had higher feed intake and improved indices of energy balance in early lactation without affecting days to first ovulation. Further investigation with greater animal numbers is appropriate for the effect of yeast supplements on days to first ovulation post-partum in dairy cows. 122 Effects of Various Dietary Fat Supplementations on Liver and Blood Parameters in Energy Deficient Transition Cows R. Karcagi , T. Gaal , P. Ribiczey-Szabo , R. Jolankai , 1 F. Husveth 1 Georgikon Faculty of Agriculture, University of Pannonia, Department of Animal Science, Keszthely, Hungary 2 Faculty of Veterinary Science, Szent István University, Department of Internal Medicine, Budapest, Hungary

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An experiment was conducted to study the influence of calcium soaps (CAS) or triglyceride (HTG) of hydrogenated palm oil on liver and blood parameters indicating the energy status and lipid metabolism of cows in the peripartum period. Holstein-Friesian dairy cows were fed on a corn-silage based diet supplemented with 11.75 MJ NEL per day of CAS or HTG or without fat supplementation (control) in a period from d 21+3 prior to the expected calving to d 100+5 postpartum. At d 21+2 (basal sample) and d 15+2 prepartum and d 5+2, 25+3 postpartum liver biopsy and blood samples and at d 100+5 postpartum an additional blood sample were collected from the cows. Total lipid (TL), glycogen (GL) content, fatty acid composition of liver lipids and glucose (GLU), insulin (INS), free fatty acid (NEFA), beta hydroxy butirate (BHBA), triglyceride (TG), cholesterol (Ch), aspartat-aminotransferase (AST) contents from the blood were determined. No differences were detected in the liver fat among the groups at d 14 prepartum or d 25 postpartum. At d 5 postpartum, however, both control and CAS-cows had higher TL and lower GL concentrations in the liver than HTG-cows (P<0.05). The variations in the liver lipid concentrations were accompanied by significant changes in the proportion of C16:0, C16:1n-7, C18:0, C18:1n-9, C18:2n-6 and C20:4n-6 fatty acids in the liver lipids. Considering blood constituents HTG-cows had higher GLU and INS concentrations than cows in the control group. HTG resulted in a higher level of GLU than CAS all duration of the experiment. HTG-cows had lower NEFA concentrations than cows in the control group, however, compared to the CAS-group, NEFA in the HTG group showed significantly lower values only at d 15 prepartum and d 100 postpartum. During the whole experiment HTG-cows expressed the lowest BHBA concentration comparing to the other two groups. No significant differences were detected in TG and CHOL in the blood among the groups at either sampling times. HTG-group had significantly lower AST values than control at all sampling times, while compared to the control group CAS-cows showed significantly lower AST only at d 100. It is concluded from our results that fat supplementations can decrease deficiencies of lipid metabolism resulted from energy deficiency in lactating cows around calving. However, HTG had somewhat higher beneficial effect on the metabolism than CAS. 123 Peripartum Heifer Response to Source of Starch in Prepartum diet. I. Milk Production and Ruminal Parameters H. Mirzaei Alamouti , K. Yazdi , H. Amanlo , A. Tohidi 1 University of Tehran, Department of Animal Science, Karaj, Iran 2 University of Zanjan, Department of Animal Science, Zanjan, Iran To determine if source of starch in close-up diets improves lactation performance and ruminal parameters , Holstein heifers (n=40) were fed two different source of ruminally fermentable starch 1) 18.6% ground corn, or 2) 18.6% rolled wheat from d -21 relative to expected parturition until parturition. After parturition, all animals received the same diet. The heifers were group-fed from d -21 to -7 relative to expected parturition and fed individually from d -7 until 7 d in milk and group-fed after 7 d until d 28. Heifers were required to have at least 5 d of prepartum dry matter intake (DMI) data to remain in the study. The prepartum diets were 1.56 Mcal of NEl/Kg and 15% CP. These diets were fed ad libitum. Close-up diet affected DMI, NEl intake, energy balance and urea pH in prepartum. The DMI changes, body condition score (BCS) and milk yield were similar between treatments, but wheat diet improved milk yield and components. There was no carry over effect of close-up diet on DMI, NEl intake and BCS in postpartum. Wheat diet has higher milk protein and lower milk urea nitrogen than corn diet. Prepartum diets did not affect ruminal fermentation characteristics. In summary, wheat grain as a rapidly fermentable source of starch did not compromise periparturient health and performance of Holstein heifers. 124 Crimping of Barley Grain with High Moisture - Scandinavian Studies H. Miettinen KEMIRA OYJ, R&D, Helsinki, Finland Crimping was developed in Finland in the end of 1960s. In 1918 British researchers had established that grain attains its peak nutritional value when the moisture content of the grain is between 35% and 45%.

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However, it took about 50 years before this knowledge was successfully turned into a method to process and preserves the grain when still moist. Traditionally, grain is not harvested until it is dry enough to be ground by a hammer mill. In crimping, the grain is combined moist and run through the crimper machine, which will break and flatten the grains. Crimped grain is stored in storage silos or plastic bags as silage. After this the question should brought up in everybody despite the early recognition why received significance of preservation of barley (and other grains) bigger attention just in a past decade. The based and the first reason should be, rises of energy prices, and as a result of this, the high costs of drying and the traditional feed preparation. Further reasons why this technologies getting more popular in worldwide, that the earlier harvesting, and over the reduce in-field losses, the less weather depended. Practical experiments by farming and livestock research institutions in Finland, Sweden, UK and Hungary and elsewhere have confirmed, that crimped feed has higher nutritional values, it increases the animals' growth and milk production, improves milk quality and the animals' health, and in addition, helps cut costs. For the good fermentation, in order to ensure the protection of nutrients and for the aerobe stability of the feed it is advantageous to use organic acid additives, as preservatives. Finnish examinations (Jaakkola, et al., 2003 MTT Agrifood) showed that in case of organic acid treated barley, the pH decrease in shorter time for value is necessary to the lactic fermentation and will standing on this level longer time than the untreated barley. Furthermore observable, that in the case of the treated barley the Ammonia- N content smaller. In studies of Crimped pea-oats we, can see, the aerobic stability of the organic acid treated grain are better, as the stability of the untreated grain or in case of treating with biological. Key words: crimping, barley, preservation, fermentation 125 Crimping of Maize with High Moisture ­ Feed Hygienic and Economic Aspects J. Karnóth , H. Miettinen , G. Kovács , I. Palkó , 5 6 7 L. Wágner , J. Péter Szûcs , E. Brydl 1 NOACK Hungary Ltd, R&D, Budapest, Hungary 2 KEMIRA OYJ, R&D, Helsinki, Finland 3 Pannon University Faculty of Georgikon Agricultural Science, Department of animal nutrition, Keszthely, Hungary 4 Taurina-Kanizsa Ltd, R&D, Kaposvár, Hungary 5 Pannon University Faculty of Agricultural Science, Department of Animal Breeding and Nutrition, Mosonmagyaróvár, Hungary 6 University of Szeged Faculty of Agriculture, Institute for Animal Nutrition Science &Technology, Szeged, Hungary 7 Szent István University Faculty of Veterinary Science, Department of Animal Hygiene, Herd Health and Veterinary Ethology, Budapest, Hungary It is essential these days to use silage technologies, which on one hand meet the strict regulations about quality assurance, animal hygiene and environment protection, and on the other hand provide high quality forage at lower costs. The aim of the study was to adopt and develop forward a system of processing and storage of grain crops, which is not depend on the low moisture content of the crops, so it makes a more flexible harvest possible and it can contribute to animal farming more successful competitiveness. This preservation method has been used in Finland and other Scandinavian countries successfully for many years. This technology includes the processing of the grain crops with crimping and the direct addition of a fermentative-preservative organic acids that depends on the ripeness of the grains (they could have even 30% of moisture content) and on the storing technology. It can be seen from our studies that this technology ensures fermentation in the right direction. According to our microbiological analyses, the number of moulds in after opening was between 100 and 1000. We made comparative analyses on the mycotoxins F-2, T-2 and DON with the hypothesis that the moulds did not start producing toxins in the maize harvested early, while toxin production may have started in the maize harvested traditionally in October. The DON contamination of the samples at the time of the treatment (end of August) and the time of feeding (October) was below trace level, while 7 out of ten samples of the maize harvested traditionally in October had a higher level of DON. The results of mould and mycotoxin tests show that the treatment with organic acids kills the fungi and bacteria present at harvest and prevent their multiplication. It can be seen from our aerobic stability tests, that the untreated control silages were considerably less stable than the treated silages. We analysed two varieties, there lasted their stability

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8 XXV. Jubilee World Buiatrics Congress 2008

only for less than 3 days. All treated wet corn samples (both varieties) protected their stability for 7 days. The fermentation dynamic, aerobic stability, mould and mycotoxin examinations have proven that from feed hygienic point of view this technology is more advantageous comparing to the traditional technologies. Taking into consideration the technology and the feeding requirements this technology can be fully inserted to the cattle feeding system. Key words: corn, crimping, preservation, fermentation, mycotoxin 160 Necessity of the New Biotechnological Methods for Avoiding Subacute Ruminal Acidosis S. Latos Dr. BATA Biotechnological Co, Reserch & Development, Ócsa, Hungary There are several strategies for feeding dairy cows, but all of them based on the roughages, rich in fiber. The ingredients added to the ration (energy-, and protein carriers, vitamins, minerals) can be used for filling the gap between the value of the basic forages and the requirements of the cow. To fulfill the energy demand of the early lactation, we have to increase the energy and decrease the fiber content of the ration. The most used energy sources are the treated cereals. The treatments are increasing the digestibility or the rumen metabolisation rate of the non-structural carbohydrates in the cereals.The high yielding status can only be maintained for short time periods but the larger problem is, that these rations are harmful for the health status of the cow. In these cases evolves the subacute ruminal acidosis (SARA). It can be defined as a temporary rumen state, with moderate pH decrease (pH 5,5-5,0) in the rumen for several hours. Cows suffering in SARA are showing the symptoms such as decrease in dry matter intake and in yield, lower milk fat, diarrhea, body condition loss. Not only a ration with high level of NSC can cause SARA, management faults (feeding techniques, sludge feeding, selection) can support the development of the problem. Even the environment (heat stress) can play a role to SARA.The fermented cereals (wet corn) taking more place in the last years can also higher the risk of SARA. They contain high amount of lactic acid and living Lactobacilli. In the acidotic rumen (anaerobic environment, pH>5,5) they can activate and start to produce even more lactic acid which can lead to an acute ruminal acidosis.In practical farm circumstances, with high NSC and critical NDF level rations it is almost impossible to avoid the SARA. This shows the subservience of using biotechnological products and methods which can help to avoid or minimize these damaging processes due to their biological activity. A typical example the world-wide used live yeast (Saccharomyces cerevisiae), which can directly affect the fermentation, unlike the so called yeast-metabolites. It is able to consume the oxygen in the rumen, and reacts with the lactic acid utilizing microorganisms. This helps to stabilize the pH, consequently minimizes the risk of SARA, resulting not only a higher production but furthermore a better health status and fertility rate. Key words: rumen acidosis, biotechnology, prevention 161 Comparison of the Effect of Feeding of Crimped versus Dry Corn Grain in High Producing Dairy Cows E. Brydl , A. Tirián , L. Könyves , V. Jurkovich , L. Tegzes , 1 2 2 3 4 R. Farkas , L. Horváth , M. Török , J. Karnóth , H. Miettinen 1 Szent István University Faculty of Veterinary Science, Department of Animal Hygiene, Herd Health and Veterinary Ethology, Budapest, Hungary 2 Gorzsa Agricultural Co, Dairy Farm, Hódmez_vásárhely, Hungary 3 NOACK Hungary Ltd, R&D, Budapest, Hungary 4 KEMIRA OYJ, R&D, Helsinki, Finland The aim of the study was to measure the effect of feeding of crimped corn grain with high moisture (DM 70%) vs. dry one on the rumen fermentation, energy, protein and acid-base metabolism, health status, reproductive performance, daily milk production and milk composition of high producing dairy cows. Kemisile 2S was used as preservative at crimping with 4.2 kg/t. The experiment was carried out at a dairy farm with 1000 head of Holstein Friesian cows. Two hundred in calf second and third parity Holstein Friesian cows were selected randomly to control and experimental groups, 3 weeks prior to expected parturition. Rumen fluid, blood and urine samples were taken for laboratory examinations once a week in the first month and once a month onwards the experiment for four times. The corn silage based diet was identical for both groups except for the control animals received dry and the

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experimental cows crimped corn. The total concentration of volatile fatty acids in rumen fluid of the experimental cows was superior to the controls by 12.1% on average. The acetate ratio of the experimental cows was superior to the controls by 13.4% on average (p<0.001). The glucose concentration in the blood samples was higher in the experimental cows than in the controls by 5.7% on average. The acetoacetic acid concentration in blood samples of the experimental cows was inferior to the controls by 46.8% on average (p<0.05). The NEFA concentration of blood samples in the experimental cows was inferior to the controls by 22.4% on average. The occurrence rate of hyperketonaemia and subclinical fat mobilization syndrome was less in the experimental cows by 16% and 5%, respectively. The BCS loss was less in the experimental cows than in the controls by 0.4 score. The time for calving to first heat, to first AI and to conception was shorter in the experimental cows by 1.6, 6.6 and 4.4 days, respectively. The daily milk yield of the experimental cows exceeded the control's milk production by 4.33 kg on average (p<0.001). Conclusions can be drawn as following: more favourable rumen fermentation, more balanced energy and protein metabolism, better health status, higher daily milk yield, higher milk protein concentration, better reproductive performance can be achieved by feeding of crimped corn instead of dry one. Key words: crimped corn, dairy cow, rumen fermentation, health, milk production 162 Effect of Fresh Alfalfa Feeding on Blood Ammonia in Dairy Cows T. Gaal1, P. Ribiczey1, R. Karcagi2, F. Husveth2 1 Szent István University Faculty of Veterinary Science, Department of Internal Medicine, Budapest, Hungary 2 Georgikon Faculty of Agriculture, University of Pannonia, Department of Animal Science, Budapest, Hungary Blood ammonia concentration in ruminants is <60 µmol/L, which increases following high nitrogen intake. To prevent ammonia toxicosis supplementation of the ration with fresh legumes or urea is not used in dairy cows. The minimum toxic concentration of ammonia causing clinical signs is not well defined. The aim of this study was to evaluate blood ammonia level in dairy cows having been fed a daily ration of 2-5 kg fresh-cut alfalfa. On a herd of 250 Holstein-Friesian dairy cows the animals were fed on a corn silage based diet. Close-up cows were supplemented with grass hay and corn grain while cows after calving received corn grain, alfalfa silage and alfalfa hay. To increase the protein intake the farmer decided to add 5-7 kg/day/cow freshly-cut alfalfa to the diet of cows from the 8th week after calving. Blood metabolic profile was made before introducing this bad practice (pre-alfalfa results) at the herd and it was repeated after 10 days of alfalfa supplementation in 19 highproducing cows (post-alfalfa results). Blood ammonia was measured by Blood Ammonia Checker (BAC) instrument while other blood parameters were analyzed by commercial test kits on an automatic chemistry analyzer. Adding fresh-cut alfalfa had no clinical consequences and did not influence the milk production. Pre-alfalfa blood ammonia level was 40 +13 µmol/L. Post-alfalfa ammonia was immeasurable on BAC in 5 of 19 cows (BAC has an upper cut-off point of 236 µmol (400 µg) /dL ammonia. Mean post-alfalfa ammonia (including these 5 cows' result as 236 µmol/L) showed a 5-fold increase (201 ±19 µmol/L, p<0.001). As we supposed an analytical error, 5-5 blood samples of dry and fresh cows not fed with alfalfa were evaluated by BAC. Ammonia concentrations in these samples were lower than 50 µmol/L.Pre-alfalfa plasma urea concentration (BUN) was 5.4±2.3 mmol/L, while post-alfalfa BUN nearly doubled (10.4±3.0 µmol/L, p<0.001). Other metabolic parameters did not show significant changes. The post-alfalfa blood ammonia concentration was extremely high compared to the 60 µmol/L upper reference limit and was reflected in elevated BUN. Previously we found such a high ammonia level in ammonia toxicosis. In this case, however, neither the behaviour nor the milk production of cows had any significant changes. Probably the upper reference limit of blood ammonia in healthy cows ought to be reevaluated. Key words: blood ammonia, urea, alfalfa-feeding, reference value 163 A New Bale-forming Technology: Higher Density and Fermentation Quality in Alfalfa Silage for Ruminants Sz. Orosz1, Z. Bellus2, S. Kapás1 1 Szent István University Faculty of Agricultural and Environmental Sciences, Department of Animal Nutrition, Gödöllô, Hungary

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Hungarian Institute of Agricultural Engineering, R&D, Gödöllô, Hungary

of study: The authors investigated a new bale-forming system: dry matter limitation of the new technology and effect of short (29% DM: 4 hour) and long term wilting (52% DM: 12 hour) on bale parameters and fermentation profile in new-type alfalfa bales. Materials and methods: Alfalfa was mowed with theoretical chop size of 20-30 mm. Baling was carried out by a Göweil LT Master variable chamber baler-wrapper machine. Pressurization: 150 bar. -1 Output: 18-20 bales hour for alfalfa. Film for wrapping was applied with thickness of 25 µm in 7 layers, respectively. The pH, lactic and th th th volatile fatty acid composition were analysed on the 13 , 30 and 70 day of fermentation according to the Hungarian National Standards (Hungarian Feed Codex, 2004). Results: Extremely high density was carried out with the new -3 technology: 213-278 DM kg m due to high pressurization (150 bar) and small particle size (20-30 mm) compared to conventional bales -3 (90-200 DM kg m ) (Table 1). Better homogeneity of the new-type bales compared to the conventional bales, owing to chopped and mixed forage, was confirmed by the low variation in wet bale weight (var. coeff. 1.4%). High density results extremely good anaerobe circumstances for fermentation (Table 2). It was found higher pH, lactic acid content and lower acetic acid content in bales with 52% DM-content. Table 1. Bale characteristics in alfalfa baled with the new baler system (n=15) 29% DM Bale wet weight (kg) Bale dry weight (kg DM) Wet density (kg m-3) Dry density (kg m-3) 904±25,1a 262±7,3a 734±10,3a 213±3,0a 52 % DM 657±13,0b 342±6,6b 534±7,9b 278±4,1b

Materials and methods: 105 clinically healthy Holstein Frisean cows were assigned randomly at 21-14 days before expected calving. Blood and urine samples were taken for metabolic profile test on <-14 prepartum, + 4, 10-14, 28-35, 56-63 and 84-91st day pp. Blood acetoacetic acid, blood plasma non esterified fatty acid (NEFA) and glucose concentration, aspartate-amino-transferase enzyme activity, urine pH and net acid base excretion (NABE) concentration and urine ketone concentration were determined. Parallel with metabolic tests from the 4th day postpartum the metritis was monitored by clinical examinations. From the 28-35th days the ovarian cycle was examined by ultrasound. The indices of calving, clinical examinations, sex and vitality of calves, data of milk production and reproduction were recorded. Results: In the <-14 days prepartum the plasma NEFA concentration shown a positive (OR 102.1; P<0.05), and the urine NABE concentration shown a negative (OR 0.99; P<0.05) correlation with PR. There was a negative correlation between NEFA and NABE in the same period (Person` corr: -0.24; P<0.05). The level of 2+ prepartum ketonuria increased the risk of PR (OR: Inf.; P<0.05). There was not relationship between the parity, sex and vitality of calves, manual assistance for calving and PR. The PR increased the risk of pueperal metritis (OR: 27.3; P<0,0001). The subsequent metabolic status, milk production and reproductive performance were not affected significantly by PR. Conclusions: The plasma NEFA and urine NABE concentrations and the >2+ urinary ketone positivity measured in <-14 days prepartum indicates the higher risk of placenta retention in dairy cows. The placenta retention indicates the relevant risk of the puerperal metritis. Key words: risk factors, retained placenta, energy balance, acid-base metabolism, dairy cow, reproduction, milk production 165 The Effect of Micro-capsulated Yeast Supplementation on Rumen Fermentation in Sheep Fed with High-energy Diet V. Jurkovich , J. Kutasi , H. Fébel , L. Könyves , 1 1 A. Tirián , E. Brydl 1 Szent István University Faculty of Veterinary Science, Budapest, Hungary 2 Dr. Bata Ltd, Ócsa, Hungary 3 Research Institution of Animal Breeding and Nutrition, Herceghalom, Hungary The aim of our study was to investigate the effects of trehalose producing yeast supplementation on the rumen fermentation of sheep fed with a diet rich in wheat and barley. Trehalose producing yeast strains are supposedly more viable in the rumen than trehalose nonproducing strains. A self controlled study was carried out using 3, rumen canullated merino wethers. In the first experimental phase (A) the sheep received trehalose producing yeast (Live-Sacc Dairy), in the second experimental phase (B) the animals received trehalose nonproducing yeast supplementation (Live-Sacc) mixed in the ration. The third experimental phase (C), without supplementation, served as control. Rumen fluid samples were taken before and after the morning feeding. Rumen pH, ammonia and VFA concentrations were measured. DM, NDF and starch degradation rates were also determined. Statistical analysis has not been carried out at the time of the abstract, thus significance of the following findings can not be stated. Rumen pH decreased in all groups after feeding (as expected) and no considerable difference was found between them. TVFA, acetate, propionate and butyrate concentrations increased after feeding, and considerable difference was found between groups. All VFA concentrations were the highest in Phase A. Ammonia concentrations remained unchanged in all phases. Degradability of dry matter showed no significant difference between Phase A and C, but it was considerably lower in Phase B. Degradability of NDF and starch was highest in the animals treated with Live-Sacc Dairy. The preliminary results demonstrated that trehalose producing yeast strain supplementation considerably modifies rumen fermentation parameters. This study was financially supported by National Office for Research and Technology (grant no.: OMFB-1213/2004) Key words: sheep, yeast, rumen fermentation 232 The Effect of Feeding of Micro Capsulated ViableYeast on Rumen Fermentation, Health and Milk Production of Dairy Cows E. Brydl , A. Tirián , L. Könyves , V. Jurkovich , 1 2 2 2 3 L. Tegzes , L. Horváth , M. Török , Zs. Szabó , J. Kutasi

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Table 2. Fermentation characteristics in alfalfa baled with the new baler system (n=5) Day of fermen13th day 30th day 70th day Treatment pH Total acid -1 g kg 28,9a 30,0a 34,8a 37,6b 32,9a 35,1a Lactic Acetic Butyric acid acid acid g kg-1 g kg-1 g kg-1 21,3a 24,4a 26,9a 30,8b 25,2a 28,2v 7,6a 5,5b 7,8a 6,7a 7,6a 6,8a 0,0a 0,0a 0,0a 0,0a 0,2b 0,1a

29% DM 52 % DM 29% DM 52 % DM 29% DM 52 % DM

4,84a 4,87a 4,60a 4,79b 4,49a 4,74b

Conclusions: It was confirmed that the new baling system is able to form bales in a wide range of dry matter content (290-520 g kg-1 in alfalfa) with high density and excellent fermentation profile. Key words: bale-forming technology, high density, fermentation quality, alfalfa silage 164 Periparturient Risk Assessments for Placenta Retention in Dairy Cows L. Könyves , O. Szenci , V. Jurkovich , L. Mrs. Tegzes , 1 3 4 1 A. Tirián , N. Solymosi , Gy. Gyulay , E. Brydl 1 Szent István University Faculty of Veterinary Science, Department of Animal Hygiene, Herd Health and Veterinary Ethology, Budapest, Hungary 2 Szent István University Faculty of Veterinary Science, Clinic for Large Animals, Budapest, Hungary 3 Szent István University Faculty of Veterinary Science, Department of Biomathematics, Budapest, Hungary 4 Private practitioner, Martonvásár, Hungary The aim of the study: was to measure how the metabolic parameters of energy and acid-base metabolism indicate the risk of placenta retention (PR) in dairy cows. The relationship between animal- and environment related factors and PR, the consequences of PR for subsequent puerperal endometritis (PM), metabolic status, milk production and reproduction were analysed as well.

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10 XXV. Jubilee World Buiatrics Congress 2008

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Szent István University Faculty of Veterinary Science, Department of Animal Hygene, Herd-health and Veterinary Ethology, Budapest, Hungary 2 Gorzsa Agricultural Co, Dairy Farm, Hódmezôvásárhely, Hungary 3 Dr. BATA Biotechnological Co, Research & Development, Ócsa, Hungary The aim of the study was to measure the effect of feeding of micro capsulated viable yeast on the rumen fermentation, health and milk production of diary cows. The experiment was carried out at a large scale dairy farm in Hungary between February and August 2007. The milk production of the cows was nearly 10 000 l yearly. Two hundred in calf second and third parity Holstein Friesian cows were selected randomly to control (No Yeast Received, NYR) and experimental (Micro capsulated Viable Yeast) groups, 3 weeks prior to expected parturition. Rumen fluid, blood and urine samples were taken for laboratory examinations once a week in the first month and once month onwards the experiment six times. The total concentration of volatile fatty acids (VFAs) was normal in both groups however in the experimental cows it was higher by 4.4% and the rumen fluid pH was more stabile. Lower concentration of NEFA, BHB and urea concentration was found in the experimental cows one month after parturition indicating more balanced energy metabolism and better health. The average daily milk production was higher by 0.6 l and the days spent for different diseases e.g. mastitis, lameness etc. was less by 33 d in the experimental group. The reproductive indices were more favourable in the experimental animals than in the controls. This work was supported by the project of GAK ALAP1-00003/204. Key words: micro capsulated, viable yeast, dairy cow, rumen fermentation, health, milk production 233 Subacute Rumen Acidosis in Intensive Italian Dairy Herds: Occurrence and Clinical Approach M. Gianesella , C. Stelletta , C. Cannizzo , 2 1 P. De Battisti , M. Morgante 1 Universita' Degli Studi di Padova, Dipartimento di Scienze Cliniche Veterinarie, Legnaro (Pd), Italy 2 Cortal Extrasoy S.P.A., Cittadella (Pd), Italy Subacute rumen acidosis (SARA) represents one of the most important metabolic disorders in intensive dairy farms that affects rumen fermentations, animal welfare, productivity and profitability. The aim of the present study was to study the occurrence of SARA in intensive Italian dairy herds and to determine the correct clinical approach to evaluat this disease. During the last four years forty commercial dairy herds were investigated. In each farm a general investigation was performed, with particular attention to herd management and presence of typical clinical signs of SARA. Twelve cows in good body condition, between 5 and 60 day in milk and without clinical signs of disease were selected randomly from each herd, to perform rumenocentesis and obtain rumen fluid. Ruminal pH was determined immediately after sampling and concentration of SCFA in ruminal fluid was determined on samples after storage. Comparisons between TMR after distribution and residual food were be evaluated by sieving and by chemical composition. Milk yield quality was determined in the cows were we performed the rumenocentesis. Results were subject to ANOVA and correlation analysis using SIGMA STAT 2.03. We had no sampling problems with the 480 cows of our study series, and no animal developed any health problem during or after the procedure. The results indicated the presence of SARA in 40% of herds (more than 33% cows with rumen pH < 5.5), a critical situation (more than 33% cows with rumen pH < 5.8) in 25.7% of farms and a normal rumen pH condition in 34.3% of herds. In particular dairy herds show an average SCFA concentration of 162, 147 and 121 mmol/L for low pH, critical pH and normal pH herds, respectively. The differences among diet composition were not significant even if herds with SARA showed a light discordance between initial composition and residual feed, a result which suggests more attention in TMR preparation and management, especially in large herds where the attitude of the personnel responsible for feeding the cows is of paramount importance. Milk yield quality were not statistical different in three groups: in particular dairy herds show an average of 3.8, 3.6 and 3.5 of milk-fat % for low pH, critical pH and normal pH herds, respectively, and the same values (3.3) of milk protein %. Although ruminal pH is reported to drop in experimental situations of inducing low milk-fat, this alone does not justify those situations to be characterized as being SARA.

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234 Measuring Rumen pH and Temperature by an Indwelling and Wireless Data Transmitting Unit ­ Application under Different Feeding Conditions J. Gasteiner , S. Rosenkranz , M. Fallast , T. Guggenberger 1 Federal Research Institute Raumberg-Gumpenstein, Animal Care and Animal Health, Irdning, A-8952, Austria 2 University of Technology Graz, Sciencepark, Graz, Austria Subacute rumen acidosis is a common and economically important herd health problem of dairy cattle and there is a crucial need for monitoring systems. An indwelling wireless data transfer system for monitoring rumen ph and temperature was assembled. Readings (user selectable) were taken every 30 minutes. Stored data are transmitted using a radio transceiver communicating with the ISM-band frequency 433 MHz. The system is controlled by a microprocessor. Data (pH, temperature) are sampled with an A/D converter and stored in an external memory chip. The latest class of the indwelling system can be administered orally, but to service the measuring units, experiments were conducted using 5 ruminally cannulated steers. After calibrating and check for proper operation rumen pH und temperature measures were carried out under following feeding conditions: 1. 100 % roughage for 1 week, measuring last 3 days 2. Daily pasture and roughage ad lib. for 3 weeks, measuring last 7 days 3. 50:50 roughage: concentrate for 7 days, measuring 7 days In experiment 3, 2 measuring units were placed simultaneous in one animal to compare and to verify data. At the same time, rumen samples were taken at intervals (1hour) and pH was detected with a pH meter. Statistical analysis was conducted using GLM (Statgraphic Plus 5.1) and Bonferroni-Holm-Test. Experiment 1: rumen temperature (mean 38.40±0.70 °C) was influenced significantly by drinking water but it is not connected with feeding time. Mean pH was 6.49±0.39 and nadir was pH 6,14. Experiment 2: mean rumen temperature was 38.12±0.80 °C and mean pH was 6.36±0.22. Nadir during pasture (day) was pH 5.34, nadir during feeding roughage (night) was 6.16. Pasture had a significant influence on rumen pH. Experiment 3: mean rumen temperature was 38.55±0.83 °C and mean pH was 6.37±0.24. Nadir was pH 5.29. Decline of rumen pH was significantly related to the feeding of concentrate. When comparing the results of 2 simultaneous measuring units, the absolute statistical error was 0.6±0.65 °C for temperature and 0.15±0.19 for pH, differences could be seen due to rumen dynamics and non-homogenous mixture of ingesta. Results indicate that the present method is a useful tool for scientific applications. As the measuring system can also be administered to uninjured cattle, an adapted rumen pH measuring system will also be assembled for practical purposes in future. Examinations were carried out in compliance with the National Animal Experiment Law (Tierversuchsgesetz 1988) 235 Blood and Ruminal Biochemical Modifications and Clinical Signs after 1 and 2 % Body Weight Intoxication with Soybean Meal in Dairy Steers D. Raboisson, A. Ferrieres, F. Enjalbert, F. Schelcher National Veterinary School of Toulouse, Department for Farm Animals and Herd Health, Toulouse, France Effects of massive ingestion of rapidly fermentable carbohydrate diets on cattle are well known, but no data are available on massive ingestion of soybean meal. The aim of the trial is to describe the clinical signs and the ruminal and blood biochemical modifications of bulls receiving an large quantity of soybean meal. In two successive trials (8 weeks apart), soybean meal (46% crude protein) was introduced via the canula in the rumen of four Holstein steers, respectively at 1% (3,5 to 3,7 kg) and 2% body weight (7,8 to 8,3 kg). Clinical examination, ruminal and blood biochemical analysis were performed every two hours for 4 days. Measured parameters were blood pH, PCO2 and HCO3- (Opti-CCA, Idex), blood NH3 (Vitros 250, Ortho Clinical Disgnostic), ruminal pH (pH-meter), ruminal volatile fatty acids (VFA) (gas chromatography) and ruminal N-NH3 (spectrophotometry). In the 2% study, ruminal pH firstly decreased to 5,67 ± 0,15 at H4 (4 hours after soybean meal ingestion), in parallel to an increase of VFA concentrations and a moderate and stable concentration of N-NH3 between H4 and H12. Then ruminal pH increased slowly to basal level (pH= 6,47 ± 0,18) up to H20 (VFA decreased and N-NH3 increased from H12 to H26). Ruminal pH finally increased slightly up to H32 and returned to basal level. Blood pH

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Nutrition and Metabolic Disorders 11

and plasma HCO3- increased from H4 to H40 and then decrease to basal level until H80. Blood NH3 concentration peak (H20 : 378,2 ± 103,3 µmol/L) was detected a few hours after blood pH peak (H12). Uremia peaked at H26. Hyperammoniemia was associated with an hyperglycemia (H24). Clinical signs (moderate muscle tremors, head shaking, stamping, weackness and moderate prostration) occurred in the 2% trial between 12 and 28 hours after soybean meal ingestion. Clinical signs were detected during hyperamoniemia, suggesting causual relationship between symptoms and blood NH3 concentration. Similar but less demonstrative biochemical modifications occured with 1% soybean meal ingestion, but no clinical sign was detected. Clinical signs and major biochemical modifications occured only with ingestion of large quantities of soybean meal, but spontaneous ingestion of such a quantity by cows is unlikely. 236 The Effect of Rumen Mucosal Lesions on Growth in South African Feedlot Cattle P. Thompson , W. Schultheiss , A. Hentzen 1 University of Pretoria, Production Animal Studies, Onderstepoort, South Africa 2 Schering-Plough Animal Health, Johannesburg, South Africa 3 Supreme Livestock Services, Heidelberg, South Africa Rumen health is well recognized as a primary necessity for bovine health and if compromised it will negatively affect performance. It is at risk when abrupt transition to a more energy dense ration takes place early on in the beef feedlot feeding period. This is also the time when most cases of undifferentiated bovine respiratory disease (UBRD) occur. Excessive pH fluctuations may cause subclinical or clinical acidosis and result in permanent damage to the rumen mucosa. Although such lesions are commonly seen in apparently healthy cattle at slaughter, there are no reports of their effect on the performance of feedlot cattle. The rumen mucosa of 1935 cattle was examined at slaughter, representing 15 groups from five feedlots. The presence and size of focal mucosal scarring ("stars") and diffuse mucosal lesions were recorded. Lung lesion scoring was also done on most animals. Live mass on arrival at the feedlot and average daily gain (ADG) during the feeding period were obtained for each animal. The effects of rumen mucosal lesions on ADG were then estimated using mixed-effects multiple regression models. Stars were recorded in 50.9% (varied from 3.3% to 72.3% between groups) and diffuse lesions in 56.1% (5.5% to 93.6%) of all animals. Pulmonary lesions were seen in 52.3% (32.3% to 77.1%) and severe pneumonia (lung parenchymal or pleural adhesion score >2) in 13,6% (4.8% to 33.9%) of all animals. Mean ADG was 1.406 kg (1.176 to 2.070 kg). When adjusted for processing mass, sex, feedlot and group, calves with rumen stars showed an average reduction in ADG of 46 g (P < 0.001). The presence of a diffuse mucosal lesion was associated with a 60 g reduction in ADG (P < 0.001). Calves with both lesion types showed a 79 g reduction in ADG (P < 0.001). However, the effect of each of the rumen lesions on ADG varied between groups. In one group of 185 animals, the presence of rumen stars was positively associated with the presence of severe pneumonia (odds ratio = 4.9; P = 0.03). This information is important for feedlot managers, supporting the fact that transition from adaptation to grower or finisher rations should be made with care. Efforts should be made to limit pH fluctuations which may cause permanent damage to the rumen mucosa that will increase days on feed to attain the desired end mass. Routine inspection of rumen mucosa at the abattoir may be also considered as a tool to monitor nutritional management in the feedlot. Key words: rumen mucosal lesions, pneumonia, feedlot cattle, growth 237 Case Report of Iodine Deficiency in an Organic Dairy Farm I. Schlemmer , M. Metzner , S. Reese , U. Ebert , W. Klee 1 University Munich, Clinic for Ruminants, Munich, Germany 2 University of Munich, Institute of Veterinary Anatomy, Munich, Germany 3 Bavarian Health and Food Safety Authority, veterinary pathology, Munich, Germany In an organic dairy farm with 40 Holstein-Friesian cows and offspring in Bavaria the farmer described tumours in the region near the larynx in five of 15 calves (up to the age of six months). Additionally the growth fell behind and the calves showed increased incidence of pulmonary diseases. In September 2006, three calves were sent to the Clinic for Ruminants for examination. Later the farm was visited. The general condition of several animals of the age of 1 day to 6 months

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was poor, the animals were emaciated, their development was reduced and their respiratory rate severely increased. Blood samples were taken from animals of different age groups for examination of glutathione-peroxidase activity (GSH-Px), thyroxin (T4) and selenium (Se) concentrations. 3 animals admitted to the clinic showed poor general condition, emaciation and severe pathological lung findings. Two of the calves had visible and palpable masses at the ventral aspect of the larynx. A main finding in blood examination was very low GSH-Px (12 ­ 42 U/g Hb). These animals were euthanized and submitted to post-mortem examination. All three animals had masses in the area of the larynx in size of 7-15 cm in diameter. Additionally, the animals had severe bronchopneumonia, and one animal also had fibrinous pleuritis. Histological findings included hypertrophy and hyperplasia with hyperaemia of the thyroid gland. The bacteriological findings were Pasteurella multocida in the lungs of two of the animals and BRSV-antigen in one animal. On the farm, nine calves had palpable masses in the area of the larynx of up to 10 cm in size. T4 levels were very low in three out of nine animals (< 0.5 µg/l) and the GPX was below 8 U/gHb in all animals examined. Analysis of the grass silage, fed ad libitum, yielded an iodine concentration of 0.29 mg/kg. Mineral feed was offered to the animals; however, it did not contain any Se or iodine. Another one day old calf with goitre, Sedeficiency and bronchopneumonia was treated in the clinic with potassium iodide, selenium, vitamin E and antibiotics. After 4.5 months the calf had recovered and was discharged. It was concluded that the farm had an iodine deficiency problem, with clinical signs of goitre, Se- deficiency and bronchopneumonia. There are no restrictions for mineral supplements in commandments for organic farms, especially for selenium or iodine. In regions with iodine and Se- deficiency in the soils there should be a special attention to mineral supplements. 238 Dynamics of Bone Metabolism in High Pregnant Heifers and First Lactation Cows J. Staric, M. Nemec, J. Jezek, M. Klinkon, T. Zadnik UL, Veterinary faculty, Clinic for Ruminants, Ljubljana, Slovenia Bone metabolism can be assessed by using biochemical markers of bone metabolism. Biochemical markers of bone metabolism are degradation products, by products of formation and enzymes involved in bone degradation or formation. They indicate whether bone is forming, degrading or remodelling more intensively. 88 late pregnant HF heifers and first lactation cows in different physiological periods, 1 month before calving, up to 10 days before calving, within 48 hours after calving, 10 to 20 days in milk and at the peak of lactation were involved in the study. The study was performed during winter, when all animals were housed. Venous blood samples for measurement of bone specific alkaline phosphatase (BALP), C-terminal telopeptide crosslinks of collagen I (CTx) and total calcium (tCa) were taken according to the protocol. Mean serum tCa has the lowest value in a group within 48 hours after calving, 2,12 ± 0,09 mmol/L and highest during the peak lactation, 2,56 ± 0,13 mmol/L. Interestingly the mean BALP has the highest value in a group within 48 hours after calving, 37,8 ± 7,9 U/L and the lowest value during the peak lactation, 22,3 ± 4,53 U/L. The highest mean CTx value was measured in group of cows 10 to 20 days after calving, 1,152 ± 0,292 ng/L and the lowest in heifers up to 10 days before calving, 0,513 ± 0,197 ng/L. All mentioned differences were also statistically significant at p<0,05. Despite Ca shortage in post parturient time, bone formation marker BALP has the highest values which indicate that there could be a strong involvement of hormones associated with calving and that low blood calcium does not lower bone formation at that time, which we expected. Bone degradation is most intensive between 10th and 20th day of lactation in this study, what could be associated with increasing milk production and demand for calcium which is supplied by bone tissue. Bone tissue is predominantly forming during last 10 days of pregnancy. Most probable reason for this is building up of Ca reserve for lactation. Key words: Bone turnover, BALP, CTx, cattle, blood 281 Change in Hepatocellular Phosphorus Content during the Transition Period in Dairy Cows W. Gruenberg , R. Staufenbiel , P. Constable , H. Dann , 4 J. Drackley 1 Purdue University, Veterinary Clinical Sciences, West Lafayette, United States

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Freie Universität Berlin, Veterinary Clinical Sciences, Berlin, Germany 3 Miner Agricultural Research Institute, Chazy, United States 4 University of Illinois, animal sciences, Urbana - Champaign, United States Hypophosphatemia and disturbed liver function are common findings in dairy cows during early lactation. Low serum phosphorus (P) concentrations have been associated with altered liver function and decreased regenerative capacity of the liver. To our knowledge, neither the liver P content nor the relationship between serum P concentration ([P]) and liver P content have been studied in periparturient dairy cows. We therefore investigated the change in hepatocellular P content in dairy cows from late gestation to early lactation. Liver biopsy samples of 6 dairy cows were obtained at -65, -30, -14, +1, +14, +28 and +49 days relative to calving. Amounts of P, potassium (K) and magnesium (Mg) in liver were determined using inductively coupled plasma-mass spectrometry and expressed as mg/g or µg/g liver dry weight. The amount of DNA in the biopsy sample was determined in order to explore the potential confounding effect of changes in cell volume on liver P content. Serum [P] was measured at -50, -36, -14, +1, +14, +28 and +49 d relative to calving. The amount of P in wet weight liver (PW), non fat wet weight (PNFW) liver, and per cell (indexed to DNA; PDNA) was calculated; similar values were calculated for K and Mg. Repeated measures ANOVA was used to identify changes in measured and calculated values over time. Liver P decreased significantly from 12.4±1.1 mg/g at -14d to 8.2±1.4 mg/g at +14d, increasing thereafter to 11.7±1.5 mg/g at +49 d. Similar changes in dry weight liver were observed for Mg and K. PW and PDNA showed the same pattern of change over time with similar trends for the Mg and K content in wet liver tissue as well as indexed to DNA. In contrast, the amounts of K and Mg in non fat wet tissue remained constant over time whereas a significant decline of PNFW at +14d was determined. Serum [P] tended to increase from calving to +14d and did not reflect changes in liver P content over time. Our findings indicate that time-dependent changes of the electrolyte contents in dry liver weight are largely due to changes in hepatic water and fat content. As indicated by the P, Mg and K content in non-fat wet liver weight, the cytosolic concentrations of Mg and K were stable throughout the study period whereas the cytosolic P concentration significantly declined at +14 d. Our findings further indicate that serum [P] cannot be used to estimate liver P content. 282 Effect of Vitamin D on the Bone Metabolism of Dairy Cattle with Hypophosphatemia JT. Wang, JL. Li, SW. Xu, FQ. Shi Northeast Agricultural University, College of Veterinary Medicine, Harbin, China The morbidity of mineral metabolic obstacle in cows was very high in Daqing and Anda regions in China, especially in some areas of Anda in which most areas belonged to alkaline soil which was composed of high content of NaHCO3. There was 5mg/kg available phosphorus in soil roughly. Dairy cattle in these areas were found to have hypophosphatemia with detecting the content of phosphorus in serum, and they showed dyskinesia and low productive ability for long time. The aim of this study was to find out the method which prevented and cured dairy cattle with hypophosphatemia, and illuminated the effect of vitamin D on the bone metabolism of cows with hypophosphstemia. 30 cows with obvious clinical symptoms which were hypophosphatemia, dyskinesia was assigned randomly. 10 cows as control feed low phosphorus grass (p<0.15%), 10 cows as group 1: feed high phosphorus grass (p>0.2%), 10 cows as group 2: feed high phosphorus grass (p>0.2%), 6 and got vitamin D3 (2.5x10 IU, weekly) by intramuscular injection and one capsule of vitamin D3 (10mg/500kg, daily) by oral. Blood samples (serum) were withdrawn on 1d, 30d, 60d, 90d and 120d. The content of vitamin D (including vitamin D, 25-OH-D, 1,25-(OH)2-D), calciumphosphorus- CT and BGP in the serum of group 2 were higher, and APK activity was lower than control and group 1. However, these items are near between control and group 1: the content of serum protein, GOP and GPT activity in serum had no difference in three groups. After 120 days the clinical symptom of 10 cows in group 2 was obviously slight, 10 cows in group 1 were better than control. These results clearly demonstrated that the content of calcium-phosphorusCT, BGP, vitamin D3 and its metabolism activity in serum of the cows with hypophosphstemia were changed obviously with adding vitamin

D and high phosphorus grass. With enhanced the content of calcium, phosphorus, CT and BGP in serum, vitamin D could effect the bone metabolism of dairy cattle with hypophosphstemia, and prevented dairy cattle hypophosphatemia Key words: cows, hypophosphstemia, vitaman D, bone metabolism 283 Changes T3 and T4 Plasma Concentrations in Dairy Cows during Lactation K. Sinka, J. Illek, D. Kumprechtova, P. Novak Veterinary and Pharmaceutical University Brno, Clinic of Ruminant Diseases, Brno, Czech Republic, Czech Republic Objective: The objective of this study was to determine plasma concentrations of T3 and T4 hormones in dairy cows in preparturient period (20 to 10 days prepartum), in early lactation (15 to 20 days in milk -DIM), peak lactation (60 to 80 DIM), and late lactation (220 to 250 DIM). Material and methods: The study was performed in a Holstein herd with average milk yield of 10,650 kg per 305 day lactation. Cows were housed loose in pens and fed total mixed ration consisting of maize silage, clover-grass silage, meadow hey, straw and concentrate. Proportions of TMR components and nutrient contents reflected the lactation phases and were in accordance with standard recommendations. Selected cows from different phases of calving to calving interval were withdrawn blood from the coccygeal vein, using the HEMOS tubes with heparin. Group A: late pregnancy cows (10-20 days prepartum), Group B: early lactation (15- 20 days prepartum), Group C: peak lactation (60-80 DIM), Group D: late lactation (220-250 DIM). In each group 10 cows without clinical disease signs and in optimum body condition were monitored. Hormones T3 and T4 were measured by chemiluminiscence, using the automatic analyzer IMMULITE 1000. Results were statistically evaluated. Results: The late pregnancy cows (A) showed the lowest T3 concentrations and the highest T4 concentrations. In early lactation there was a significant decrease in T4 and non-significant increase in T3. In peak lactation, T4 levels were still low but non-significantly higher than in the early lactation. In late lactation there was a significant increase in T4 and significant decrease in T3. The late lactation values were similar to those measured in late pregnancy cows. Table 1. T3 and T4 plasma concentrations (nmol/l) Group T3 X S.D. C.V. T-test 1.71 0,179 0,105 A-B ** (P<= 0.01) A T4 74.64 8,562 0,115 A-B *** (P<= 0.001) T3 1.93 0,183 0,095 B-C N.S. B T4 36.9 3,157 0,086 B-C ** (P<= 0.01) T3 1.95 0,135 0,069 C-D *** (P<= 0.001) C T4 T3 D T4

43.96 1.77 63.96 7,096 0,095 7,592 0,161 0,054 0,119 C-D *** (P<= 0.001

Conclusion: Plasma concentrations of thyroid hormones were changing throughout the calving to calving interval. In the early lactation there was a significant drop in T4 concentrations, remained low in peak lactation, and increased in late lactation only. T3 concentrations were significantly higher in early and peak lactation than the initial values. In late lactation, T3 concentrations were gradually decreasing and approached the values measured in late pregnancy cows. The study was carried out within the project NAZV no. 1B44035 284 Predictive Value of Serum Biochemical Analysis on Treatment Outcome in 72 Recumbent Cows J. Berchtold , H. Maier , P. Constable 1 Tierärztliche Praxis Haiming, Obing, Germany 2 Synlab.Vet., Augsburg, Germany 3 Purdue University, School of Veterinary Medicine, Department of Veterinary Clinical Sciences, West Lafayette, United States Introduction: The objective of this study was to determine the predictive value of serum biochemical analysis on treatment outcome in recumbent cows.

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Materials and Methods: Serum samples from recumbent cattle were selected from samples submitted to a laboratory in Germany for analysis. A questionnaire was completed in order to obtain data on the duration of recumbency, time of sampling, breed, and treatment outcome. Serum concentrations of Ca, Phosphorus, Mg, Na, K, total bilirubin, urea, and total protein were measured, and serum activities of AST, CK, and GLDH were determined. Analysis of variance was used to compare serum values of cows that did not recover and were sampled before (group NR1) and after (group NR2) their first treatment for recumbency, and recovered cows that were sampled before (group R1) and after (group R2) their first treatment, respectively. Nonnormally distributed variables were log transformed before analysis and P <0.05 was considered significant. Results: Serum was analyzed from 72 cows aged 3 to 12 years, with 51 (70%) of cows recovering. Breeds included 45 Fleckvieh, 6 Holstein, 19 Braunvieh, 2 Red-Pied, and one mixed. Serum was obtained before treatment in 38 cows (groups R1, NR1) and after one or several treatments in 34 cases (groups R2, NR2). Duration of recumbency when serum was obtained was <1 day in 41 cows. Group NR2 cows had significantly higher geometric mean serum activities for AST (718 U/L, range 162 to 2,616) and CK (10,025 U/L, range 184 to 62,952), compared to cows in group NR1 (AST, 293 U/L; CK, 2,443 U/L), group R1 (AST, 184 U/L; CK, 1,138 U/L), and group R2 (AST, 228 U/L; CK, 1,754 U/L). Serum calcium concentration was lower in group R1 cows than group R2 and NR2 cows, but was similar to cows in group NR1. Overall mean serum potassium concentrations were lower in Holstein (3.4+0.5 mmol/L) than in Fleckvieh (4.6+0.8 mmol/L) or Braunvieh (4.2+0.2 mmol/L) cows. Other serum parameters did not differ between the 4 groups and between breeds. Conclusions: Higher serum activities for AST and CK in cows not recovering from recumbency indicate the development of muscle damage secondary to recumbency, with high serum AST and CK activities being associated with a poor prognosis. Our results suggest that potassium supplementation should be routinely performed in lactating recumbent Holstein cows because they have a lower potassium concentration than recumbent Fleckvieh and Braunvieh cows. Key words: recumbency, serum biochemistry, muscle damage, hypokalemia 285 Abnormal Dynamic Changes of Plasma Leptin are Associated with the Occurrence of Sub-clinical Ketosis in Dairy Cows around Parturitions B. He, J. Huang, T. Hu, Y. Liao, Y. Du, J. Ma, G. Li GuangXi University, College of Animal Science and Technology, Nanning, Guangxi, China, China In order to understand the link between leptin and ketosis in Holstein cows around calving, The 16 Holstein cows around parturitions were chosen randomly from a dairy farm in Nanning area, their blood samples were collected within Days 10 prepartum and at Days 0,1,2,7,14,21,28,35,42,49,56 postpartum. The concentrations of leptin, ketone bodies, glucose and lipids in the blood plasma were analyzed. Then the examined cows, according to diagnostic criteria of ketosis, were divided into two groups, i.e. the cows with subclinical ketosis (SK) and the cows without subclinical ketosis (nSK). The concentration and dynamic change characteristics of each above parameter were analyzed between two groups, the linear correlations between parameters were also analyzed in SK and nSK groups. The concentrations of plasma glucose and lipids were determined by routine methods, the level of leptin by RIA and ketone bodies by salicylaldehyde spectrometry. The results shows that: The SK group had significantly lower average leptin concentration (P=0.000024) and higher average ketone bodies concentration (P=0.00045) than nSK group. Although the SK group had lower average glucose concentration than nSK group, the difference between them was not significant (P=0.0575). There was no significant differences in average lipids levels between two groups (P=0.05). For correlations, both SK and nSK groups had highly significant correlations between leptin and ketone bodies, also between glucose and ketone bodies concentrations, so did between leptin and lipids, between ketone bodies and lipids in nSK group. On the dynamic curve, the changes of leptin, ketone bodies and glucose in SK group displayed lower frequency wave with various amplitudes within test period, the leptin exhibited a long-span wave with the peak less than 1.1ng/ml. In nSK

group all parameters except lipids displayed high frequencies fluctuations. In conclusion, the lower frequency changes of leptin, ketone bodies and glucose concentrations are the results of energy metabolism dysregulation in the cows with SK. The occurrence of a single long-span wave in leptin dynamic change within 8 weeks postpartum may be associated with the underfeeding SK. Key words: dairy cow, leptin, ketosis, dynamic correlation, lipids 286 Flying Scapulas as a Symptom of Muscular Dystrophy in Dutch Dairy Heifers K. Jobse , R. Bouwstra , M. Holzhauer 1 DAP Nijkerk-Wellensiek, Nijkerk, Netherlands 2 Faculty of Veterinary Medicine, Utrecht University, Animal Health Department, Utrecht, Netherlands 3 Animal Health Service Deventer, Deventer, Netherlands Introduction case report: Within one week after turning out heifers on pasture, a dairy farmer consulted his local veterinary practitioner. Three heifers out of a group of 30 displayed a symptom in literature described as "Flying Scapulas". This abnormality is characterised by a separation of both scapulas from the thorax due to bilateral rupture of the serratus ventralis muscles. After this first consult all heifers were housed and provided extra vitamin E and selenium (Se). Materials and Methods: During this first consult blood was sampled from all three heifers. Three weeks later blood was sampled from the two remained heifers. Blood was analyzed for vitamin E, glutathione peroxidase and serum creatine phosphokinase activity. One heifer was euthanized three days after the first consult and one heifer three weeks later. On this heifer post mortem research was carried out. Results: The first blood samples revealed a vitamin E as well as a Se deficiency in all three heifers while CPK levels were too high. After three weeks of supplementation vitamin E and Se levels were normal. One heifer did not clinical recover and still had a high CPK blood level and was therefore euthanized. Post mortem research showed a bilateral rupture of the serratus ventralis muscles. No new clinical cases of muscular dystrophy have been observed after housing and providing extra vitamin E and selenium. Conclusion: The most likely reason for developing the specific symptoms of "Flying Scapulas" is muscular dystrophy caused by an inadequate vitamin E and Se supply during winter, followed by excessive exercise after turn-out. The authors' opinion is that this is the first report in which both a vitamin E and selenium deficiency have been found in animals with the clinical symptoms of Flying Scapulas. This also is the first case-description of cattle with this specific symptom in The Netherlands. Key words: case-description, flying scapulas, muscular dystrophy 287 Total Lipid and Triacylglycerol Determination in One Analytical Approach in Small Amounts of Bovine Hepatic Tissue R. Busche, A. Haudum, A. Starke, J. Rehage University of Veterinary Medicine Hannover, Foundation, Clinic of Ruminants, Hannover, Germany Fatty liver is the most frequent liver disease in dairy cows in early lactation and an important risk factor for other periparturient diseases. The increase of the total lipid (TL) content is mainly due to accumulation of triacylglycerol (TAG) but also to a rise in other hepatic fat fractions. Since fine needle biopsies commonly yield no more than 100mg of hepatic tissue the aim was to develop an analytical combined method for TL and TAG determination in small amounts of liver tissue. Liver bioptates (10g) taken from 10 different HF cows during surgical correction of abomasal displacement were splitted into equal portions and stored at -85°C. TL were extracted from homogenised tissue samples (ca. 50mg) with 2ml hexane:isopropanol (3:2, v/v; at 20°C, 24h, constant agitation). Then 1ml Na2SO4 (0.455 mmol/l) was added. Samples were vortexed, centrifuged (400g, 10min) and the hexane phases with extracted lipids were transferred to clean tubes. For complete lipid extraction 1ml hexane was added to the bottom phase, then vortexed, centrifuged, and the hexane supernatant was used again. After hexane evaporation the dry extract was used for gravimetric TL determination. For TAG analysis the dry lipid extract was dissolved in 1ml hexane and aliquoted according to TL content. Commercial test kits on an automated analyser without modifications revealed incomplete enzymatic TAG hydrolysis. Thus, an extra incubation step for complete hydrolysis was included, and various

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14 XXV. Jubilee World Buiatrics Congress 2008

lipases (porcine pancreatic lipase type II and type VI-S, microbial lipase), detergents (SDS 10%; Lubrol), and incubation periods (0.5-16 h) were tested. Triolein (Sigma, Steinheim, Germany) was used as internal standard. The coefficient of variance (CV) of TL analysis in the ten liver bioptates (range: 40-314mg/g FW) was in average 2.2% (max. 4.5%; three repetitions). For the extra incubation step prior to the final enzymatic TAG analysis (MTI diagnostics, Idstein, Germany) microbial lipase, Lubrol, and 16 hours of incubation provided best results. Mean recovery of Triolein was 101% (97%-107%) with mean intra and inter day (10 samples, 5 repetitions) CV% of 0.75% and 2.7%, resp. The liver TAG (range: 4-260 mg/g FW) analysis showed mean intra and inter day CV% of 2.5% (0.35-5.6%) and 3.4% (2.3-4.9%), resp. The presented combined method for TL and TAG determination in small amounts of bovine liver tissue was simple, accurate and reproducible. This work was supported by WILHELM SCHAUMANN STIFTUNG, Hamburg, Germany. 288 Study on the Mechanism in the Damage of Erythrocyte Membrane in Low-phosphorus Cows SW. Xu, FQ. Shi, DW. Xuan Northeast Agricultural University, College of Veterinary Medicine, Harbin, China Objectives: Investigate the mechanism in the damage of erythrocyte membrane (EM) in low-phosphorus cows. Materials and Methods: These cows with field cases were divided into three groups, including hemoglobinuria group (HG), low-phosphorus group (LPG) and control group (CG). The phospholipid composition, skeletin, antioxidant function and shape of EM were determined. Results: The phospholipid composition, skeletin, antioxidative function and shape of EM obviously changed in HG and LPG. (1) Phosphatidylethanolamine (PE) content in HG was significantly lower than that in LPG and CG, but sphingomyeline (SM) and phosphatidycholine (PC) + phosphatidylserine (PS) content in HG was significantly higher than that in LPG and CG. In comparison between LPG and CG, PC + PS content were lower and SM content was higher in LPG. Significant positive correlation and negative correlation were observed between serum phosphorus and PE content, serum phosphorus and SM content, respectively. (2) There were no difference was found in EM skeletin between LPG and CG. Spectrin I, spectrin II, and band IV-2 content was lower in HG than that in LPG and CG, but band III was higher in HG than that in LPG and CG. (3) SOD activity and GSH-Px activity in HG and LPG was significantly lower than that in CG. MDA content in HG and LPG was significantly higher than that in CG. There were a significant positive correlation between serum phosphorus and erythrocytic SOD activity and GSH-Px activity, and a negaitive correlation between serum phosphorus and erythrocytic MDA content. (4) Through the observation of scanning electron microscope, the course of erthrocytolysis attributable to low phosphorus intake was proved: the erythrocytes changed from biconcaval shape to acanthocytes, spherocytes and to be destroyed eventually with the decresed serum phosphorus content. Treatment with phosphorus preparation could signficantly alleviate the change in erythrocyte shape and make it to return to normal shape. Conclusions: The phospholipid composition, skeletin, and shape of EM changed, and the antioxidant function of EM decreased in lowphosphorus cows. These further caused the occurrence of the haemolysis. Key words: cow; low-phosphorus; erythrocyte membrane 289 Prevalence of Digital Dermatitis in First Lactation Cows Presented at Auction M. Hulek , I. Sommerfeld-Stur , J. Kofler 1 University of Veterinary Medicine Vienna, Department of Horses and Small Animals, Clinic of Orthopaedics in Large Animals, Vienna, Austria 2 Veterinary Practice Michael Hulek,Oberneukirchen, Austria 3 University of Veterinary Medicine Vienna, Department of Animal Breeding and Reproduction, Institute of Animal Breeding and Genetics, Vienna, Austria Objective: The risk of introducing digital dermatitis (= DD) to dairy herds by the introduction of infected, newly purchased cattle is assumed to be considerable. The aim of the study was to assess the prevalence of DD in first lactation cows (FLCs) presented as breeding cattle at the monthly auction in one auction centre in Austria over a period of 10 months.

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Material and Methods: The FLCs to be examined were selected randomly for claw examination for each auction from the monthly auction catalogue. At each auction a minimum of 25% of FLCs were selected, with an average of 36% of all FLCs of all 10 auctions. After obtaining owner's consent, the hindclaws were examined in a walk-in crush for DD. DD lesions were evaluated by the parameters localisation (plantar, interdigital, dorsal aspect), diameter in cm and lesion type (M1-M4). Other claw lesions and trimming status of the claws were also noted. Results: From a total of 1110 FLCs registered for the ten auctions on the catalogues, 399 were chosen for examination, and of these 199 FLCs could be examined. In 63 cows from the random sample the owners did not consent to the examination. A total of 24 FLCs were found to have DD lesions on one or both hindlimbs, with at least one cow detected at nine of the ten auctions. The size of lesions ranged from 0.5 to 3 cm in diameter. The prevalence of DD determined at 10 auction dates within 10 months was 12.06%. Conclusions: The results of this study show that about 12% of FLCs presented as breeding cattle at auction were affected with DD. This result suggests that in the worst case, if each of these FLCs affected with DD is introduced into a DD free herd, 24 new herds could be infected by DD during a 10 months period from one single auction centre in Austria. An additional risk is the infection of other animals at the auction from the use of shared walkways and pens, with a potentially much larger number of herds affected at a later stage. In order to reduce both these risks, we recommend that dairy farmers purchase only cows free from digital dermatitis. An examination of the claws for the presence of DD lesions should be carried out in all cows one or two weeks before they are presented at auction. This examination should be performed by trained experts and the findings should be documented in a special protocol, which should be presented with the cow at the auction, and be available for inspection by all potential buyers.

POSTER ABSTRACTS

443 Influence of Age, Season and Physiological State on some Biochemical Parameters in South Eastren Algerian Desert Goats H. Nadia , M. Toufik , M. Bakir , B. Mabrouk 1 Batna University, Veterinary, Batna, Algeria 2 Batna University, Virology, Batna, Algeria

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Blood plasma Ca, P, Mg, Na, K, and Fe were analysed in order to establish biochemical references, and to study the influence of age, season and physiological state on the variation of these parameters. The results demonstrate: ­ Ca, Mg, and Na levels were high at birth and decrease with age. ­ The season had a signficant effect on the levels of these ions, for example Ca, Mg, K decreased and inversely Na increased during dry season. ­ The values obtained for the plasma Ca, Mg, Na, K, Ca:P and Fe levels in pregnant goats were 80.02±4.84 mg/l, 22.14±1.61 mg/l, 142±1.73 mEq/l, 6.43±0.40 mEq/l, 1.29±0.37 and 91.11±18.84 µg/100ml respectively. They were significantly higher than in non pregnant and lactating females. ­ Ca (77.67±3.20 mg/l), Mg (20.44±1.66 mg/l), K (5.65±0.57 mEq/l) and Ca:P (1.35±0.35) were lower in lactating goat compared to pregnant and non pregnant goats. These values of plasma minerals can be used as reference to detect metabolic and/or nutritional disorders in goat. Key words: mineral metabolism, goat, lactating, desert 44 The Evaluation of Vitamin A and ß-carotene Levels during Postpartum Period in Semi Industrial Dairy Farms in Iran M. Tehrani Sharif , R. Mozaffary , J. Gholami Seyed kolaee , 2 2 M. Rezaee ghale , A. Cheraghzadeh 1 Islamic Azad University , Garmsar branch, Department of Pathobiology , Faculty of Veterinary Medicine, Garmsar, Iran 2 Islamic Azad University , Garmsar branch, Student, Garmsar, Iran 3 Shahid Chamran University, Student, Ahwaz, Iran Objective of study: Vitamin A, retinol, plays a vital role in vision sense. Due to the presence of large amounts of beta-carotene in cattle's foods &

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vitamin A stores in liver, the hypovitaminosis is not provable. Increased secretion of vitamin A in milk during postpartum period, increases the risk of hypovitaminosis occurrence in cattle's feed on a poor beta- carotene nutrition. The aim of this study was to evaluate the value of vitamin A & beta-carotene during postpartum period in some semi industrial farms with poor beta-carotene based nutrition in Ghaemshahr Iran. Material & methods: Blood samples were obtained from 88 dairy cows during postpartum period. The samples were centrifuged and froze pa at -20 °C. To prevent direct contact of samples with air and light, serum were preserved at dark pockets. Cows were classified at 9 groups based on postpartum period (1,2,3,4,5,6,7,8,9 month after parturition). Vitamin A & beta-carotene were determined by spectrophotometric method. Results: The mean value of vitamin A & beta-carotene were 38.06±9.16 µg/dl & 33.8±9.74 µg/dl respectively. The mean value of vitamin A in 1,2,3,4,5,6,7,8,9 month postpartum were 33.98±7.26, 29.51±3.5, 35.23±1.44, 37.20±7.62, 38.74±9.06, 38.09±8.20, 39.91±6.96, 42.89±4.87, 46.05±5.01 respectively. The mean value of beta-carotene in 1,2,3,4,5,6,7,8,9 month postpartum were 37.02±3.45, 40.17±1.56, 26.79±3.40, 40.29±6.26, 18.07±4.02, 36.15±3.6, 59.61±8.36, 36.31±7.31, 31.11±8.19 respectively. Conclusion: The mean values of vitamin A were normal in spite of consumption of poor beta- carotene based nutrition. It might be due to high level of liver stores. There were not significant differences in mean vitamin A during postpartum period. The mean beta-carotene were lower than reference ranges during post partum period. The decreased level of beta-carotene indicates the nutritional deficiency of beta-carotene. Key words: Vit A, beta carotene, postpartum period 445 Comparison of Clinical and Laboratuar Responses for Laparotomic Omentopexy and Laparoscopic Abomasopexy in Cows with Left Abomasal Displacement K. Yigitarslan , N. Yavru , S. Avki 1 University of Mehmet Akif Ersoy, Faculty of Vet.Med, DEPARTMENT OF SURGERY, Burdur, Turkey 2 University of Selcuk, Faculty of Vet. Med., DEPARTMENT OF SURGERY, Konya, Turkey The objective of this study was to compare the metabolic responses and recovery period in cows with left abomasal displacement following by surgical treatment either with laparotomic omentopexy or laparoscopic abomasopexy. Forty cows brought to the Faculty of Veterinary Science (The Universities of Selcuk and Mehmet Akif Ersoy) and diagnosed with left abomasal displacement were used. The cows were divided for two groups. Twenty cows (group 1) were treated with laparotomic omentopexy and the other 20 were treated with laparoscopic abomasopexy. There were no statistically differences between both groups concern about sex, age, number of calving, time after parturition, diseases period, level of displacement, feces quantity, body temperature, heart frequency and serum lactate levels. Serum K and Cl levels were nd reached to normal levels in laparoscopic abomasopexy group at 72 th hours postoperatively. The serum cortizol levels were maximum at 0 hours both in laparoscopic abomasopexy and laparotomic omentopexy groups, and it was decreased under the preoperative levels after 24 hours. In cows treated with laparoscopic abomasopexy, the serum cortizol levels th nd were statistically decreased at 24 and 72 hours compared to the laparotomic omentopexy group (p<0.05). Serum ß-hidroxybutirate levels th were statistically decreased in laparoscopic abomasopexy group at 24 th and 72 hours compared to the laparotomic omentopexy group (p<0.05). As a conclusion, laparoscopic abomasopexy technique may be superior than laparotomic omentopexy due to the reduced risk of abdominal contamination, less intestinal movement loss in postoperative term, rumen contraction number reached normally earlier, small incision for enter to abdominal cavity, definite of diagnose and less operative stress. 446 The Levels of Serum Amiloid A and Haptoglobin Concentration in Cows with Abomasal Displacement H. Guzelbektes, A. Coskun, M. Ok, I. Sen University of Selcuk, Faculty of Vet. Med., Department of Clinical Sciences, Faculty of Veterinary Medicine, Konya, Turkey The objectives of study: The purpose of this study (1) was to determine concentrations of selected acute phase proteins (serum amyloid A and haptoglobin), haematological and biochemical changes in cows with abomasal displacement.

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Materials and methods: Forty-two Swiss® Holstein dairy cows with left displacement abomasums (LDA) and sixteen Swiss® Holstein dairy cows with right displacement abomasums (RDA) were used in this study. Clinically healthy post parturient cows (n = 9) from a local dairy farm were used as control group. Blood samples were collected from the jugular vein from all the cows. Surgery was performed in all cows with displacement abomasum (DA). The abomasum was repositioned followed by an omentopexy. Results: The mean concentration of haptoglobin and serum amyloid A in cow with DA without concurrent diseases were slightly increased compared to control groups. The concentration of haptoglobin and serum amyloid A in cows with DA with infectious diseases were greater high compare to cows with DA with non-infectious disease. Serum glucose concentration in cows with DA had increased in general. But serum glucose concentrations in cows with RDA were more increased compared with cows with LDA. The serum albumin levels in all cows with DA were slightly decreased compare to control group cows. Conclusion: Acute phase proteins level in cows with DA without concurrent disease were slightly increased. Although concentration of Hp and SAA in cows suffered from ketosis and fatty liver was mildly increased, concentration of Hp and SAA in infectious diseased cows with DA increased more. But there was no relationship between elevated acute phase proteins level and prognosis of DA. We can say that elevated acute phase proteins level in cows with DA are primarily because of the relationship of concurrent disease and stress. Acknowledgment: This study was supported by University of Selcuk, Scientific Research Project Coordinating. Key words: serum amyloid A and haptoglobin, abomasal displacement, cows. 447 Ameliorative Effects of Boron on Serum Profile in Buffalo (Bubalus Bubalis) Fed High Fluoride Ration V. Bharti , M. Gupta 1 CCS Haryana Agricultural University, Hisar, Department of Veterinary Physiology, Hisar, India 2 Indian Veterinary Research Institute, Division of Physiology & Climatology, Bareilly, India An experiment was undertaken to evaluate the protective role of boron on the serum profile of buffalo calves fed a high fluoride ration. Twelve male Murrah buffalo (Bubalus bubalis) calves of 6-8 months age were divided into three groups of four calves in each group. They were fed basal diets and supplemented with sodium fluoride (NaF, 60ppm) alone or in combination with borax (Na2B4O7.10H2O, 140ppm) for 90 days. However, boron (B) was added in the ration as borax to make @140ppm boron (elemental B) on DM basis in treatment II. Dietary F caused a significant (p<0.05) depressing effect on serum Ca and Zn on day 90, which was improved with B supplementation. However serum Fe and Cu did not show any significant change on F as well as F+B supplementation. The serum ALP and phosphorus level were increased significantly (p<0.05) upon F feeding but declined significantly (p<0.05) when B was fed. These findings suggest that boron has beneficial effect on serum minerals and ALP in buffalo calves fed high fluoride ration. Key words: boron, buffalo, enzyme, fluoride, mineral 448 Clinical Field Evaluation of a Butaphosfan + Vitamin B12 Compound (Phosphorum B12/Catosal® - Bayer) in the Treatment of Subclinical Ketosis in Dairy Cows V. Cuteri , L. Nisoli , A. Attili , A. Romero Tejeda , A. Fruganti 1 University of Camerino, Veterinary Science, Matelica (MC), Italy 1 Bayer Health Care, Veterinary Science, Milan, Italy Sub-clinical Ketosis (SK) is a metabolic disorder in high-producing dairy cattle characterized by abnormal levels of ketone bodies in blood, urine and milk and absence of clinical signs. The inability of the cow to regenerate the energy lost to sustain the lactation, leads in economic losses through decreased milk production and reproductive performance. The aim of this study was to evaluate the effect of Butaphosfan and Cianocobalamine (Phosphorum B12® 10% inj. solution) on dairy cattle with sub-clinical ketosis. During February to September 2007, a negativecontrolled, blinded, multi-centre and randomised clinical study was carried out on 79 primiparous and multiparous animals reared in 4 Italian farms. One group (n=42) was treated with Phosporum B12® (KP) at a dose of 25 ml/animal i.m., once a day for 5 consecutive days, while the Sub-Ketosis Control (KC) group (n=37) was not treated. Milk was

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16 XXV. Jubilee World Buiatrics Congress 2008

collected from each cow and tested for the presence of Betahydroxybutyrate (BHBA) - an indicator of abnormal fat metabolism using Ketolac® and following manufacturer's instruction. Appetite and bovine activity (ALPRO DELAVAL System) were evaluated for 10 days post enrolment (PE). Rectal temperature and symptoms of clinical ketosis, Left Displaced Abomasum or other diseases, were also recorded. Milk BHBA 200 Ïmol/L was used as the cut-off value for diagnosis of SK while values <100 µmol/L were considered surely negative. After 10 days PE, the incidence of SK based on the milk test was statistically different between th groups (P=0.0213), with a significant decrease from 5th (48.65%) to 10 day (23.81%) only in the treated KP group (P=0.0129). Inoculation of Phosphorum B12® increased the prevalence of cows cured (milk BHBA <100 µmol/L) at the 5th day of treatment (16.67%), showing a statistical difference (P=0.0401) vs KC group (2.70%). Also, the treatment improved the average milk production to 27.53L for KC group vs 30.94L for KP group (P=0.0106). Even if the appetite reduced in both groups, only untreated cows showing a serious diminishment and anorexia at 5th day recorded a significant decrease (P<104) in the average of milk yield (19.27L) in comparison to the animals with normal appetite (28.93L). No differences were observed about temperature, activity and other symptoms. The administration of Phosphorum B12® resulted in a sooner reduction of BHBA levels in milk and in a better recovery of milk production. 449 Calcium Levels and Some Biochemical Parameters in Lambs with White Muscle Disease S. Kozat Yuzuncu Yil University, Ozalp Vocational High School, Özalp-Van, Turkey White Muscle Disease (WMD) or Muscular dystrophy is a nutritional and enzootic disease of lambs that is characterized by muscle degenerations. In muscle dystrophy, the total calcium values are elevated. This study tested the hypothesis whether serum calcium values change in WMD of lambs or not. 10 healthy and 15 WMD lambs of Akkaraman breed were included in the study. Prior to treatment (at day 0), serum calcium values of lambs with WMD were higher than those of healthy control group (P <0.01). Similarly, prior to the treatment the CK, LDH, AST, ALT, and ALP values in lambs with WMD were significantly higher compared to those of control (P <0.001, P <0.001, P <0.001, P <0.01 and P <0.05, respectively). After treatment (at day 30), none of the parameters were significantly different than healthy controls (P >0.05). In addition to biochemical parameters mentioned above, simultanous serum calcium values were elevated in WMD lambs. Key words: White muscle disease, lamb, calcium, biochemical parameters 450 Evaluation of Whole Cottonseed as Effective Fiber Source in Early Lactating Holstein Cows M. Salehpour , H. Amnalou 1 Islamic Azad University ,Roudehen Branch, Agriculture College, Tehran, Iran 2 University of Zanjan, Animal Science Department, Zanjan, Iran Twelve Holstein cows in early lactation were used imbalanced changeover design to evaluated the effect of replacing alfalfa Neutral Detergent Fiber (NDF) with NDF from whole cottonseed at deferent levels (0,7,14,20) on dry matter bases that replaced to alfalfa hay. The diets were similar as crude protein and neutral detergent fiber on dry base. Cows fed with total mixed ration individually. Dry matter intake and milk yields were higher for cows fed high whole cottonseed (14,20%) diets than for that alfalfa control diet. High whole cottonseed diet had best efficiency. Milk fat percentage and milk fat yield were highest for high whole cottonseed diets, but no significant difference were found among the another milk composition compound (protein, lactose and SNF). Milk component yields were significant difference (p<0.05). Rumen pH and fecal pH were affected not significantly by experimental diets. Chewing activity increased linearly when increased whole cottonseed percentage. Increased whole cottonseed percentage associated increased physically effective fiber. Digestibility results were indicated that control diet and 7 percentage whole cottonseed diets had highest dry matter digestibility (p<0.05), But organic matter, crude protein and NDF digestibility were not affected significant among diets. Daily weight gain and body condition score change were

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affected by diets that14 and 20 percentage whole cottonseed diets were indicated highest daily weight gain and body condition score (p<0.05). Plasma urea nitrogen was higher for 14 and 20 percentage whole cottonseed diets. Key words: early lactation, whole cottonseed, alfalfa hay, effective NDF, chewing activity 451 Effect of Molybdenum and Sulphur on Copper Status and Mohair Quality in Merghoze Goat M. Moeini , E. Nooriyan , M. Souri , E. Mikaeeli 1 Razi University, Animal Science, Kermanshah, Iran 2 Razi University, Animal Nutrition, Kermanshah, Iran This study was made on the effects of a normal diet containing 12.8 mg Cu/kg DM, which supplemented with molybdenum and sulphur on the copper status and fibre quality in eight 1- year Iranian Merghoze goat. One group (n=4 mean weight 31±2.0 kg) was treated with Mo and S supplements for 20 weeks, the second group (n=4 mean weight 32± 2.1 kg) served as controls. In addition of blood sampling for measuring copper status in plasma, the copper content and quality of fleeces were measured every 6 weeks. Mohair measurements were carried out by 2 taking patch samples (10 x 10 cm ) from the mid-side area of the goat. The analytical set consists of plasma copper concentrations (Pl Cu), Trichloroacetic acid soluble copper concentrations (TCA-Cu), and fleece copper content. The results indicated that the addition of 30 mg Mo and 2.5g S /kg DM to the normal diet did produce sub clinical copper deficiency. One such visual symptom was the loss of fleeces pigmentation and poorer crimp being observed. The results showed that there was a significant decrease in Pl Cu (P<0.5) along with a significant increase in thiomolybdate (MoS) production after 4 months. The Pl Cu minus TCA-Cu plasma became more than 2 µM in the blood of treated goat, indicating that there was a significant thiomolybdate formation in the body. The signs of copper deficiency are likely to be from thiomolybdate formation in the body, which reduced mohair quality in Merghoze goat. Key words: copper deficiency, molybdenum, sulphur, fiber, Goat 452 An Investigation of Zinc Deficiency Effects in the Nutrition of Holstein Heifer and Determine the Best Form Zinc Supplements for its Composition M. Salehpour, M. Hajipour Islamic Azad University, Roudehen Branch, Agriculture College, Roudehen, Iran A 100-d study was conducted to determine zinc concentration of nutrition and it interaction effects on serum zinc concentration "immune system" health and daily weight gain and also for determining the best mineral supplements of zinc absorption(zinc sulphate and oxide)on 24 Holstein heifers with average age if 6 month and weight163 kg in completely randomizes design(CRD) and a 2*2 factorial experiment form including 4 treatments consisting six heifers in the vertical integration of Babol Gavedasht in Mazandaran province. The amount of zinc fed animals in all treatments including two levels and two factors(one equals zinc sulfate and the other zinc oxide) and ration of fed have formulated in a way that energy , protein and all nutrients except zinc were similar. The results of experiments indicate that about 70% of feeds have relative deficiency of zinc but no sings of deficiency including allopecia, parakeratosis, decrease of appetite and hock inflammation. Also, the results show there was no significance between zinc supplements have been seen during the whole period particularly and twenty eight day of experiment but fed zinc supplements has been significant especially zinc sulphate levels in the amounts of zinc in blood serum. 453 The Effect of Selenium and Vitamin E Administration to Heifers during the Late Stage of Pregnancy on Copper and Zinc Status M. Moeini , E. Mikaeili , A. Kyanei 1 Razi University, Animal Science, Kermanshah, Iran 2 Faculty of Veterinary Medicine, Animal Medicin, Kermanshah, Iran This study was conducted to determine the effect of injection of selenium and vitamin E to pregnant heifers at the last stage of gestation on serum Se, Cu, Zn and Fe status. Fifty Holstein heifers were

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Nutrition and Metabolic Disorders 17

randomly assigned to one of five treatments. Four and two weeks before expected calving, the heifers were injected intramuscularly 0 ml (T0), 10 ml (T1), 20 ml (T2), 30 ml (T3), 40 ml (T4) selenium and vitamin E respectively. The control (C) group received no supplement. Blood samples were collected from heifers four weeks before expected calving and at calving day. Blood samples of calves were drowning from the jugular vein at birth and 7 days of age. Serum concentrations of Se were determined using hydride generation Atomic absorption Spectrophotometer. The result indicated that the serum concentration of Se, Cu and Fe increased in T3 and T4 treatment heifers at calving day but no change seen in serum concentration of Zn. Serum Cu concentrations in calves of treated groups increased at birth and first week of age but was not significantly different. The serum concentration of Cu and Fe increased in calves of heifers given Se and vitamin E compound esp. in T3 and T4 treatments but the serum concentration of Zn decreased. It seems that the Zn: Cu and Zn: Fe ratios be disturbed which lead to Zn reduction. It can be concluded that Zn should be supplemented when more than 40 mg/Kg Dm administered to pregnant heifers. Key words: Zn, Cu, Se, interaction, heifers 454 Effect of Mixed Enzymes on Alfalfa Hay Nutritive Value Using in vitro Gas Production and in situ Techniques B. Baghbanzadeh Nobari, A. Taghizadeh, H. Janmohamadi, S. Alijani, H. Paya, G. Moghadam Dept. of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran Fibrolytic enzymes are typically cellulase that degrade cellulose or xylanase that act on hemicellulose (Beauchemin et al., 1995). Enzymes applied to forages immediately before incubation enhance the digestion of dry matter (DM) and neutral detergent fiber (NDF) suggesting that fibrolytic enzymes applied at feeding can improve digestion in ruminants. 455 Changes of Serum Parameters in Monensin-contained Diet of Gezel Sheep B. Baghbanzadeh Nobari , A. Rezapour , P. Namavar , 2 G. Moghadam 1 Dept. of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran 2 Dept. of Animal Science, Faculty of Agriculture, Islamic Azad University of Tabriz, Tabriz, Iran Five Gezel male lamb treated with 4 levels of monensin (0, 10, 30, and 60 mg/kg of diet) in a change-over design. Blood samples were taken one hour before and 2 hrs after feeding. Serum was separated and freezed up to analysis. Serum levels of glucose, beta-hydroxy-butyrate, triacylglycerol, total protein, albumin (albumin/ globulin), and blood urea nitrogen (BUN) was measured by colorimetric methods. Considering liver health, serum activity of AST, ALP, and GGT was measured. Ruminal fluid sample was taken for pH analysis (ruminocynthesis). Total and differential white blood cell count and phagocytosis test undertaken. Results of this study showed that monensin in a level of at least 30 mg/kg of diet could effectively inhibit acidosis occurrence. Acidosis has a negative effect on appetite; acidotic animals have lower levels of glucose in serum. But, monensin does not have any independent positive effect on serum glucose. Even two-folds of this dosage had no negative effect on liver. Monensin does not any stimulatory effect on natural immunity either independently or dosedependently. Generally staying, monensin may be inhibiting acidosis elated complications such as natural immune depression. Key words: gezel, immunity, monensin, sheep 456 D-vitamin Status of Cows in Preparturient and Postparturient Periods under Injection of Cholecalciferol L. Yuskiv , B. Kurtiak , V. Vlizlo 1 Institute of Animal Biology UAAS, Cattle Nutrition, Lviv, Ukraine 2 Institute of Animal Biology UAAS, Scientific Center for Prion Infections Study, Lviv, Ukraine The aim of our work was the investigation of influence of different doses (210 and 420 IU D3/kg body weight) of vitamin D3 on vitamins status of cows in preparturient and postparturient periods under the parenteral

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injection of vitamin D3 once before calving and three times after calving with interval of seven days. The experiment was carried out on the three groups of cows of Black-and-White breed, 5-6-years-old age, in a winterspring period. The cows of first group, to which vitamin D did not injection governed as control. To the cows of 2-nd and 3-rd groups the vitamin D3 was injected intramuscularly in abovementioned doses. In blood the level of 25-hydroxyvitamin D, the concentration of calcium, magnesium, inorganic phosphorus, the activity of total alkaline phosphatase and its isoenzymes at 3-5 days before calving, 5-7 days and 55-60 days after calving were determined. It was established, that the content of 25-OH D3 in the blood serum of cows of 1-st, 2-nd and 3-rd groups at 3-5 days before calving was correspondently: 18,75; 25,95 and 32,85 nmol/l, and at 5-7 days after calving 15,83; 20,83 and 31,88 nmol/l. At 55-60 day after calving the concentration of 25-OH D3 in blood serum of cows was the greatest and varied from 20,48 till 35,0 nmol/l. Thus in the cows blood of 2-nd and 3-rd groups before and after calving dose dependence increase of concentration of calcium of total, protein-linked and ultra-filtrates was observed. The concentration of magnesium in the blood of cows of 3-rd group in the 5-7 day after calving was statistically significantly higher about 19%, and in the 55-60 day about 6 %, comparatively with the control. At the same time, in the blood of cows of 2-nd and 3-rd groups, comparatively with the control group, dose dependence increase of concentration of inorganic phosphorus and decrease of the activity of alkaline phosphatase and its isoenzymes (bone and intestinal) was observed. Parenteral injection of cholecalciferol to cows before and after calving maintained the level of 25-OH Vit D3 and effected on the exchange of mineral components of blood, that prevents the decrease of their level in postparturient period. The character of these changes depend on the physiological state of cows and on the doses of vitamin D3. Key words: cows, prepartum, postpartum, calcium, 25-hydroxycholecalciferol 457 Clinical and Subclinical Cases of Phosphorus Deficiency and Hypophosphatemia in Egyptian Buffaloes H. Morsy , A. Sheikh , M. El-gamal 1 Faculty of Veterinary Medicine, Suez Canal University, Department of Animal Nutrition, Ismailia, Egypt 2 Faculty of Veterinary Medicine, Department of Animal Nutrition, Ismailia, Egypt 3 Faculty of Veterinary Medicine, Department of Animal Nutrition, Cairo, Egypt The correlationship between food intake, hypophosphatemia, blood contents, pregnancy and health condition were done in 35 bufalloes 24 months before parturation at 7-9 years old (9 healthy as a control,12 with clinical and 14 with subclincal hypophosphatemia according to blood , serum and feed analysis and clinical examination). Analysis of feed stuff revealed a lower level of phosphorous, a higher level of calcium and molybdenum with a normal level of copper. Clinical cases of hypophosphatemia showed normal temperature, elevated pulse and respiration, sluggish rumen movement, depraved appetite, pica, dullness, emaciation, red haemoglobin urine and pale or sometimes icteric mucosus membrane. The subclinical cases showed a normal temperature, pulse, respiration, depraved appetite, dullness and sometimes locomotors disturbances. The blood level of hemoglobin content and PCV were significantly decreased in clinical cases only. Serum levels of inorganic phosphorous was highly significant decreased with a decrease of serum copper level in clinical cases and significantly decreased of phosphorous in subclinical cases. Serum calcium and molybdenum levels were significantly increased in clinical cases and slightly increased in subclinical cases. Serum magnesium, sodium, potassium and chloride showed a non significant changes in clinical and subclinical cases. Treatment with Sodium dibasic phosphate (60 gm diluted in 300 ml distilled water was given 4 times within 12 hours intervals, The first injection was intravenous, the others was subcutaneous) as well as daily 20 ml catozal intravenous and 250 gm bran and 10 gm copper sulphate per os in food for one week was satisfy for treatment the clinical cases. Subclinical cases were treated with oral daily of 30 gm sodium dibasic phosphate, 125 gm of bone meal and 2-4 kilogram of brain for a period of one month. Correction of feed supply was helpful and advised in both cases. It is concluded that diet, blood and serum analysis and the general health condition were helpful for diagnosis of clinical and subclinical hypophosphatemia in pregnant buffaloes,

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18 XXV. Jubilee World Buiatrics Congress 2008

correction of diet and phosphorus supplement was useful for treatment and prevention of hypophophatemia. 458 Ketosis in Dairy Cows under Obligatory Reduction of Feed Intake A. Elsheikh , T. Allam , H. Morsy 1 Faculty of Veterinary Medicine, Animal Medicine, Zagazig, Egypt, Egypt 2 Animal Health Research Institute,Zagazig, Animal Medicine, Zagazig,Egypt, Egypt 3 Faculty of Veterinary Medicine, Suez Canal University, Nutrition and Clinical Nutrition, Ismailia, Egypt Clinical findings, post-mortem, laboratory examination of blood serum and urine as well as milk production and body weight were done in lactating Friesian cows subjected to a reduction of food intake at the first and second month of parturition. Clinical symptoms included loss of body weight, dehydration, sunk eyes, reduction of milk production, crossing of fore limb, lameness, constipation, ketoneuria and acetone odour were appeared within 3-7 weeks after restriction of food intake. Post-mortem of dead or emergency slaughtered cows showed a yellow gelatinous foamy fluid inside and around visceral organs, oedema of pericardium, and intestine. Enlargement greasy and fatty degeneration of liver which was easily perforated on pressure. Serum levels of ketone bodies and total lipids were significantly increases meanwhile serum levels of glucose, total protein, calcium, phosphorus and magnesium were significantly decreased. It is concluded that prolonged feed restriction causes chronic wasting form of ketosis, serous complication and severe economic loss among dairy cows due to sharp drop of milk production and loss of body weight. 459 RBC and WBC Counts in Lactating Dairy Cows Affected by Subacute Ruminal Acidosis M. Morgante , M. Gianesella , C. Cannizzo , P. Dalvit , 1 1 2 M. Gatto , LM. Coppola , L. Moro 1 Universita' degli Studi di Padova, Dipartimento di Scienze Cliniche Veterinarie, Legnaro (PD), Italy 2 Istituto Zooprofilattico Sperimentale delle Venezie, Legnaro (PD), Italy The influence of subacute ruminal acidosis (SARA) on ruminal microbiology has received considerable attention, but less is known about systemic manifestations that arise from SARA probably because it is well known that variables such as breed, stage of growth, reproductive status, and stage of lactation have an influence on many blood parameters. The aim of this study was to investigate possible changes in some haematological parameters in dairy cows affected by SARA. During autumn 2007 the study was carried out on 12 highly productive farms, stationed in different zones throughout northern Italy. In all farms, 12 cows in early lactation, in good body condition and without clinical signs of disease were selected randomly from each herd, to perform rumenocentesis and obtain rumen fluid. Blood samples for hematological profiles were obtained from the same cows that had rumenocentesis performed by jugular venipuncture. The herds were divided into 3 groups depending on the mean rumen pH: group A farms with pH>5.8 (normal), group B pH between 5.6 and 5.8 (critical) and group C, pH<5.6 (acidosis). Data were statistically analyzed by analysis of variance to verify the effect of the group. The 3 groups were homogeneous for average dry matter intake and all the animals were on absence of external clinical signs of disease. Day in milk (DIM) and body condition score (BCS) were not statistical different (p<0.05) in 3 groups: in particular dairy herds show an average of 35±19, 28±16 and 32±18 of DIM and an average of 3.02±0.29, 3.07 ±0.22 and 3.06 ±0.18 of BCS for group A, B and C, respectively. RBC counts, HGB, HCT, RDW, PLT and MPV were not significantly different in three groups, whereas the concentration of total WBC increased statistically (6.62 3 ±1.73, 6.84±1.91 and 8.73 ±1.98 10 /µl) in group A, B and C, respectively. In particular dairy herds show an average differential 3 WBC counts of 2.8±0.94, 3.4 ±1.01 and 3.8 ±0.96 10 /µl of 3 neutrophils, 2.4±0.64, 2.6 ±0.71 and 2.9 ±0.69 10 /µl of lymphocytes 3 and 0.34 ±0.02, 0.23 ±0.01 and 0.22 ±0.01 10 /µl of basophils for group A, B and C, respectively. These data suggest that modifications of some haematological parameters can be observed and in particular

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the total and differential WBC count were significantly higher in the dairy cows belonging to the farms affected by SARA. 460 Effect of Dietary Buffer Addition on the Concentrations of Trans-18:1 Fatty Acids and Conjugated Linoleic Acid (CLA) in the Milk of Dairy Cows I. Vudmaska, O. Golubets, V. Vlizlo Institute of Animal Biology UAAS, Cattle Nutrition, Lviv, Ukraine CLA and some trans-18:1 fatty acids (trans-10 and trans-11 18:1) have been shown to promote health-related effects in human and animals, including anticarcinogenic and antiatherogenic effects and effects on body composition and fat metabolism. The objective of this study was to determine the effects of varying amounts of dietary concentrate and buffer addition on incorporation of trans-18:1 fatty acids into milk fat and on synthesis of CLA by the mammary cells. The effect of concentrate to forage ratio and buffer (sodium bicarbonate) supplementation on milk fat composition in Holstein cows was objective of study. The 3 diets were examined: 80:20 concentrate to forage ratio, and 50:50 forage to concentrate ratio with (1% of DM) and without sodium bicarbonate. The fatty acid profile was determined by GLC, using a 100-m SP-2560 capillary column (Supelco, Inc.). Cows fed high-concentrate diet have lower ruminal pH (6.14). The addition of buffer to this diet has prevented this effect (pH 6.72). Low ruminal pH appears to be a factor that results in the inhibition of the some steps of FA bio-hydrogenation that convert of unsaturated fatty acids to stearic acid. Altered ruminal function resulting from low ruminal pH for cows fed the high-concentrate diet may play a role in the increased production of trans-18:1 fatty acids in the rumen. Milk trans-18:1 fatty acids were higher for cows fed the high-concentrate diet without buffer than for cows fed the low-concentrate diet without buffer and high-concentrate diet with buffer (5.52 % vs. 4.66 % and 4.71 %). The milk trans-10 18:1; trans-11 18:1; trans-10, cis-12 18:2 and cis-9, trans-11 18:2 concentrations in cows fed the low-concentrate diet, high-concentrate diet without buffer and high-concentrate diet with buffer were 0.23, 1.76, 0.34 %; 3.36, 2.12, 3.13 %; 0.05, 0.16, 0.08 % and 1.21, 0.87, 1.28 %. At the same time, concentration of the 18:0, total 18:1, total 18:2 and total 18:3 fatty acids in the milk of cows fed these diets were 10.02, 11.28, 10.71 %; 23.26, 28.90, 26.91 %; 3.19, 3.68, 3.57 % and 1.38, 0.92, 0.67 %.The milk fat concentration was lower for cows fed the high-concentrate diet without buffer (3.04 % vs. 3.74 % for cows fed low-concentrate diet). Supplementation of high-concentrate diet with sodium bicarbonate has elevated the milk fat to 3.78 %. Key words: cows, milk, trans-18:1 fatty acids, CLA, sodium bicarbonate, pH 461 Cytokine Synthesis in Calves Fed with the Feedingstuffs Supplemented by Tylosine or Prebiotics M. Szymanska-Czerwinska, D. Bednarek National Veterinary Research Institute, Department of Cattle and Sheep Diseases, Pulawy, Poland, Poland The aim of the study was to examine the effect of tylosine and prebiotics (mannan-oligosacharides and ,-glucans) on the cytokine synthesis in calves. The calves were fed with feedingdstuffs supplemented either by tylosine (group I) or prebiotics (group II). The calves in group III served as the controls. The additives, i. e. tylosine and prebiotics were used in a form of commercial pharmaceutical preparations: Tylan (Elanco) and Alphamune (Alpharma), respectively. The experimental calves were fed with traditional feedingstuffs supplemented either by tylosine in a recommended dose of 700 mg of the active substance per animal (group I) or prebiotics (, glucans and mannan-oligosaccharides) in dose 14 g per animal (group II). The animals in group III served as the controls and received the same feedingstuffs without the additives. The both kinds of feedingstuffs were given to the calves for seven weeks. Blood samples from calves were taken twice a week by the jugular vein puncture before the calf morning feeding. Duration of the blood collection was 7 weeks too. The following cytokines were assayed: interleukin 1 (IL-1), 2 (IL-2), interferon (IFN) and tumor necrosis factor (TNF). The cytokines were investigated in the calves serum using: ELISA tests and biological method by Daniluk and Kandafer-Szerszen. The obtained results showed, that in blood sera of calves the concentration of cytokines significantly increased in both experimental group (I and II). However, the significant higher increase of the concentration was

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observed in group II of the animals treated with prebiotics. According to the data presenting in the literature ,-glucans and tylosine are considered as powerful immunostimulators of macrophages and other immune effector cells (e.g. monocytes, neutrophiles, NK etc.) mainly by binding with their specific receptors (CR3, TLR, CD16/CD18 etc.) situated on a surface of the cells and activated them. Key words: tylosine, prebiotics, cytokines, calves 462 Effects of Omentopexy on Tbars and Plasma Frap in Dairy Cows with Left/Right Abomasal Displacement P. Mudron , J. Rehage 1 University of Veterinary Medicine, Clinic for Rminants, Kosice, Slovakia 2 University of Veterinary Medicine Hanover, Foundation, Clinic for ruminants, Hanover, Germany Objective of study: Higher levels of free radicals and reactive oxygen species can result in a damage of biological macromolecules and tissues. Therefore, the present investigation was aimed to study the effects of omtopexy stress on thiobarbituric acid reactive substances (TBARS) and ferric reducing ability of plasma (FRAP) in dairy cows suffering from the left and right abomasal displacement (LAD and RAD, respectively). Material and methods: Ten H-F dairy cows, (5.26 years), admitted for treatment of left abomasal displacement (omentopexy), were used in our study. Blood samples were drawn from the jugular vein prior to surgery, immediately and then 15, 30, 60, 90 minutes, and 2, 5, 10, and 24 hours after reposition of abomasum. In the study on the right abomasal displacement ten H-F dairy cows, (4.74 years), admitted for treatment of right abomasal displacement, were used. Abdominal surgery (omentopexy) was performed in a standing position 1-2 hours after admission. The average surgery duration was 26.1 minutes (without anaesthesia). Blood samples were drawn from the jugular vein in the same intervals as in the cows with the left abomasal displacement. Results: LAD Surgical stress resulted in a significant increase in plasma cortisol concentrations (p<0.001) with the highest mean levels 15 minutes after surgery (32.4±18.1 µg/l) as well as in serum levels of glucose (p<001) and NEFA (p<0.001). Similarly, surgery transiently enhanced the levels of TBARS (p<0.001). The highest TBARS plasma levels (0.91±0.18 µmol/l) were recognized 60 minutes after reposition of abomasums. There were no significant differences in FRAP in dairy cows before and after reposition of abomasum. RAD ANOVA showed significant changes in plasma cortisol concentrations within the study (p<0.05). In contrast, both TBARS and FRAP were not significantly affected. Conclusion: The data of the trial with the left abomasal displacement indicate that stress reaction caused by surgical correction of left displacement of abomasum can be associated with higher production of TBARS. Although the data of cows with the right abomasal displacement indicate that stress reaction caused by surgical correction (omentopexy) of the right abomasal displacement is not strong enough to overwhelm the stress response to pre-surgical events (transport, sickness, new environment), in generally, stressful events seem to be able to lift lipid peroxidation at the higher levels. 463 Blood Gas Parameters in Dairy Cow Affected by Subacute Ruminal Acidosis M. Morgante , C. Cannizzo , M. Gianesella , P. Dalvit , 1 1 2 F. Bevilacqua , L. Coppola , A. Stefani 1 University of Padua, Department of Veterinary Clinical Science, Legnaro (Pd), Italy 2 Istituto Zooprofilattico Sperimentale delle Venezie, Legnaro (Pd), Italy Subacute ruminal acidosis (SARA) may be a common and economically important problem for dairy cattle but clinical signs of SARA in a dairy herd can be subtle and easily overlooked. The diagnosis of SARA in a herd or a group, is difficult and the technique greatly employed is the measurement of ruminal pH in a subsample of cows. According several researchers rumenocentesis may be useful for the collection of rumen fluid for pH determination but this technique is often too difficult in field conditions. The aim of this trial was to study an alternative method of diagnosis of SARA, particularly by the evaluation of some hematological parameters, especially blood gas. During autumn 2007 twelve farms have been monitored: in all farms, twelve cows in early lactation, in good body condition and without clinical signs of disease were selected randomly

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from each herd, to perform rumenocentesis and obtain rumen fluid. At the same time blood samples for haemogasanalysis were obtained by jugular venipuncture from the same cows that had rumenocentesis. The herds were divided into three groups depending on the mean rumen pH: group A counted farms with average ruminal pH > 5.8 (normal), group B included farms with average ruminal pH between 5.6 and 5.8 (critical), and in group C, dairy farms presented an average ruminal pH < 5.6 (acidosis). Data were statistically analyzed by analysis of variance to verify the effect of the group. The three groups were homogeneous for average dry matter intake and all the animals were on absence of external clinical signs of disease. Blood bicarbonates, pH and partial pressure of CO2 didn't show significant differences but oxyhemoglobin (%O2 Hb), partial pressure of O2 (pO2), calculated and measured oxygen saturation (sO2c and sO2m) were significantly different. In particular %O2 Hb was 70.73±8.52, 66.25±8.95, 64.92±8.63 - pO2 was 41.18±5.03, 33.43±4.95, 34.67±5.01 mmHg - %sO2c was 76.70±7.97, 64.31±7.88, 66.57±7.84 %sO2m was 70.80±8.64, 66.19±8.05, 64.96±8.95 in group A, B and C respectively. These data suggest that cow attended by SARA probably don't have any problem in acid-base equilibrium but this condition could be related with some problems in blood oxygenation status. 464 Effect of Yea-Sacc 1026 Supplementation on Productive Response of Dairy Cows Fed Corn Silage-based Diets during Summer V. Novais R. Cabrita , C. Gomes , J. Fonseca , S. Andrieu 1 Universidade do Porto, Centro de Estudos de Ciencia Animal, Porto, Portugal 2 Cooperative Agricola de Vila do Conde CRL, Vila do Conde, Portugal 3 Universidade do Porto, ICBAS, Instituto de Ciencias Biomedicas de Abel Salazar, Porto, Portugal 4 Alltech, Biotechnology Centre, Dunboyne, Ireland Objectives of study: The objectives of the study was to evaluate the effects of Yea-Sacc®1026 (Saccharomyces cerevisiae CBS 493.94, Alltech) supplementation on feed intake and performance of dairy cows fed corn silage-based diets during summer in Portugal. Materials and methods: Effects of Yea-Sacc®1026 on feed intake and productive response of dairy cows fed corn silage-based diets were evaluated in an randomized complete block design experiment, using 12 Holstein cows. Animals were fed a diet comprising (dry matter basis) 42% corn silage, 8% coarsely chopped ryegrass hay, and 50% concentrate (mixture of raw-materials) without (Control) or with 1 g Yea-Sacc®1026 _per kg (Yea-Sacc). Experiment lasted for 35 days. During the first seven days, cows were fed the control diet (covariate period). At the end of the covariate period, cows were blocked by 4%fat corrected milk, parity number, and days in milk in six blocks of two animals each, and randomly assigned to one of the two treatments (Control and Yea-Sacc). The experiment was carried out during June and July 2006, mean maximum and minimum daily barn temperatures being 28 °C (SD = 4.6 °C) and 18 °C (SD = 3.2 °C), respectively. Results: Yea-Sacc®1026 supplementation did not affect dry matter (DM) intake (19.6 and 20.4 kg DM/day, respectively for Control and Yea-Sacc), but tended to increase milk production (P=0.072; 30.6 and 33.0 kg/day, respectively for Control and Yea-Sacc) and milk protein production (P=0.096; 0.82 and 0.88 kd/day, respectively for Control and Yea-Sacc), and decreased milk lactose content (P=0.083). Plasma urea concentration did not differ between treatments. Conclusions: Results show that the addition of Yea-Sacc1026 increase milk production and milk protein production of dairy cows fed corn silage-based diets subjected to relatively high environmental temperatures. The cost/benefit ratio of using Yea-Sacc was 1:4 (according to feed costs in Cooperativa Agrícola de Vila do Conde, Portugal, in June 2006). Key words: dairy cow, milk production, heat stress, live yeast 465 Effect of a Modified Glucomannan Fraction from Yeast Cell Wall Extract (Mycosorb) on Milk Production in Dairy Herds in South Italy M. Agovino , S. Andrieu 1 Alltech, Biotechnology Centre, Dunboyne, Ireland 2 Alltech, Alltech Italia, Bologna, Italy Objectives of the study: The objective of this study was to evaluate the effect of a modified glucomannan fraction from yeast cell wall

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20 XXV. Jubilee World Buiatrics Congress 2008

extract (Mycosorb®, Alltech Inc.) on milk quality (SCC, % fat & protein) and quantity. Materials and methods: The study was conducted on dairy cow herds where visual observations indicated the presence of mycotoxin contaminated forage. The study was conducted on 2 dairy farms. The total milk production and milk composition was recorded for the total trial period. The trial was conducted near Rome, Italy. The trial was done on a "before and after" basis with production figures being compared before and after the inclusion of Mycosorb Farm Pak. (including Mycosorb at 20% on a carrier). Mycosorb Farm Pak was distributed at 100 g/head/days for the first 10 days and 50 g/head/day after until the end of the trial. Results: FARM 1The trial was conducted with 83 lactating HolsteinFriesian cows. The basal diet consisted of corn silage, Ryegrass hay, alfalfa hay, maize meal, feed concentrate (31% crude protein), soybean meal and peas. The total milk production and milk composition was recorded for 5 months: first 2 month without Mycosorb and consecutive 3 months with Mycosorb in the diet. Mycosorb inclusion increased milk yield by 1.5 l/h/d while stage of lactation remained identical; protein percentage (3.36% vs 3.41%) and fat percentage (3.71% vs 3.97%) while somatic cell count (SCC) was reduced (529 vs 196, x1000 cells/ml). FARM 2The trial was conducted with 74 lactating Holstein-Friesian cows. The basal diet consisted of corn silage, ryegrass hay, alfalfa hay, feed concentrate (21% crude protein). The total milk production and milk composition was recorded for 12 months: 3 months without Mycosorb followed by 3 months with Mycosorb in the diet and then again 3 months without Mycosorb. With Mycosorb in the diet, SCC decreased (430 vs 288, x1000 cells/ml) and increased again (288 vs 598, x1000 cells/ml) after the removal of Mycosorb. Mycosorb inclusion increased milk yield by 2 l/h/d without change in stage of lactation at herd level. Conclusions: In conclusion, the inclusion of Mycosorb into the diet can be used to alleviate mycotoxins negative impact on productive performance and milk composition. Key words: dairy cows, milk production, mycotoxins 466 Effects of Increasing Prepartum Dietary Protein Level Using Poultry by-product Meal on Productive Performance and Health of Multiparous Holstein Dairy Cows F. Kafilzadeh , M. Yazdi , H. Amanloo 1 Razi University, Department of Animal Science, Kermanshah, Iran 2 University of Zanjan, Department of Animal Science, Zanjan, Iran The effects of two levels of crude protein (CP) using poultry by-product meal fed during late gestation on the performance, blood metabolites, some reproductive parameters and colostrum composition of Holstein cows was studied. Sixteen multiparous cows 26±6 d before calving were assigned to one of the two diets in a randomized block design to evaluate the effects of two isocaloric diets containing either 14 or 16 % crude protein. Postpartum diet was similar in both groups through wk 3 of lactation. Yields of milk, protein, lactose, fat, and SNF were not affected by prepartum crude protein level. Colostrum composition (fat%, CP% and Total solids%) blood metabolites (Ca, Glucose, Total protein, Albumin, Globulin, Urea N and Cholesterol), some of the reproductive performance indexes (Pregnancy duration, Days open, First AI, service per conception, First service Conception rate%, and Pregnancy rate%), and disease incidence were not influenced by prepartum crude protein level. Colostrum fat percent was significantly affected (P<0.04). There was no significant difference in body weight and BCS in cows received the two different diets. Prepartum blood urea N concentrations were elevated in the cows fed 16 % CP diets prepartum (P<0.02). Serum cholesterol during prepartum and postpartum periods was significantly different in the two groups of cows. In general except reduced fat in colostrum and increased blood glucose level in cows received 16% CP in diets it seems feeding 16% had no other obvious advantages over 14% protein in close up cows. Key words: poultry by-product meal, dairy cow, close-up, protein 467 Trans Fatty Acids in Milk of Dairy Cows E. Vargas-Bello-Pérez, P. Garnsworthy University of Nottingham, School of Biosciences, Loughborough, United Kingdom Lipids obtained from dairy products are an important part of the human diet in many countries; nearly 75% of the total consumption of fat from ruminant animals comes from bovine milk fat. Milk fatty acid (FA)

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composition has numerous effects on milk quality, including its physiological properties (e.g. melting point and hardness of butter, fractionation of milk fat) as well as its nutritional properties (e.g. potential positive and negative effects on human health). Some FA, such as butyric acid, oleic acid, polyunsaturated FA (PUFA) and conjugated linoleic acid (CLA) have potential anticarcinogenic and antiatherogenic effects on human health. Other FA such as some saturated FA (lauric, myristic and palmitic acids) and some trans FA, (e.g. elaidic acid) have potential negative effects on human health by increasing blood LDL-cholesterol. Trans fatty acids (tFA) are produced during biohydrogenation of PUFA in the rumen and are incorporated into the milk fat of lactating cows. There has been intensive research on trans FA, mainly on the effects of specific isomers of trans C18:1 that can be found in dairy products and margarines. The major tFA found in milk is vaccenic acid (trans-11 C18:1), which appears to have mainly beneficial effects on human health. There is also interest in the role of tFA in the milk fat depressing effects of certain dairy diets containing low fiber or fish oil. However, the exact mechanisms of this phenomenon remain unclear. Economically, milk fat depression has a great impact, causing important losses to dairy producers. Recently, however, there has been interest in milk fat depression as a possible strategy to maintain energy balance and thereby improve reproductive performance of dairy cattle. Because milk fat contains a complex mixture of FA, the research challenge is to improve the FA profile for human health without compromising the physical properties of milk or the metabolism and performance of the cow. 468 The Concentrations of Zn, Cu, Mn and Se in Milk in Relation to Daily Milk Production and Stage of Lactation of Dairy Cattle A. Pechová, L. Pavlata, R. Dvorák University of Veterinary and Pharmaceutical Sciences Brno, Clinic of Ruminants, Brno, Czech Republic The concentrations of microelements in dairy milk are studied especially in connection with nutrition of children. There is only little information about factors, which effect the concentration of various trace elements in milk. The aim of this study was to evaluate the effect of daily milk production and stage of lactation on the concentrations of zinc (Zn), copper (Cu), manganese (Mn) and selenium (Se) in cow's milk. The experiment was carried out in dairy farm of Holstein cattle with average milk production 8,562 kg. It was included 35 dairy cows, which was housed in one division. The sampling of blood and milk was realized two times in the course of four weeks. All experimental cows were fed on stable total mixed ration during whole period. The concentration of microelements in daily ration was following: Zn 105 mg/kg, Cu 17 mg/kg, Mn 66 mg/kg, Se 0.4 mg/kg of dry matter. The determination of single trace elements was carried out with atomic absorption spectrophotometry: F-AAS (Zn and Cu in serum, Zn in milk), ETA-AAS (Mn in blood, Mn and Cu in milk), HG-AAS (Se in blood and milk). The daily milk production varied from 19.6 l to 62.6 l with the average 36.6 l of milk. In term of stage of lactation there were results from 7 to 188 days of lactation (the average 105.4 days). The concentrations of single trace elements in blood/serum show that the supplementation of trace elements was adequate. The concentrations were following: Zn (serum) 1063.9±181.1 µg/l, Cu 826.8±138.1 µg/l, Mn 14.0±3.2 µg/l and Se 183.8±29.6 µg/l. The concentrations of trace elements in milk widely varied and we found the following dates: Zn 3855.2±814.7 µg/l; Cu 36.3±14.4 µg/l; Mn 20.1±8.3 µg/l; Se 28.6±7.1 µg/l. The relations between daily milk production and concentration of Zn, Mn and Se were not statistically significant. Only copper concentration in milk correlates significantly (r=-0.302, p<0.05). The relations between stage of lactation and milk concentration of trace elements were closer. We found positive correlation for Mn (r=0.419, p<0.01) and on the contrary negative correlation for Cu (r=-0.258, p<0.05) and Se (r=-0.277, p<0.05). On the basis of correlation analysis we did not find significant relations between concentrations of studied trace elements in milk and in blood/serum. The work was realized with the support of MSM Project No. 6215712402. Key words: cows, microelements, blood 469 Influence of Soybean Seeds and Hydrolyzed Palm Oil on the Metabolism of Lactating Dairy Cows V. Lubojacká, A. Pechová, R. Dvorák, L. Pavlata University of Veterinary and Pharmaceutical Sciences Brno, Clinic of Ruminants, Brno, Czech Republic

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The aim of the monitoring was to compare the influence of fat supplementation in the form of full-fat soybean seeds and hydrolyzed palm oil as energy sources for the metabolism of dairy cows in early lactation. During the first 8 lactation weeks, 17 dairy cows, divided into experimental (E; n=8) and control (C; n=9) groups, were followed. The basic diet was the same for both groups. The differences were in the composition of the concentrate mixture, which had a balanced content of nutrients, but different source of fat. The group E received heattreated soybean seeds (22% in concentrates) and the group C hydrolyzed palm oil. The two diets started to be fed on the day of st th th calving. Blood samples were taken weekly (1 -5 ) and 8 weeks after delivery. At the end of the experiment was taken ruminal fluid. During the experiment BCS in both group worsened similarly. In group E decreased from 3.8±0.2 to 3.3±0.3 and in Group C from 3.5±0.4 to 3.1±0.4. The BCS of the group E was on average by 0.3 point higher than in group C, but the difference was not significant. Despite this unfavourable factor, a better performance of hepatic metabolism was proved in the group E. In this group lower concentrations of nonrd esterified fatty acids was identified in 3 (0.96±0.38 vs. 1.76±0.63 -1 th -1 mmol.l ; p<0.01) and 8 (0.32±0.29 vs. 0.78±0.48 mmol.l ; p<0.05) week after calving. The gamaglutamyltransferase activity in serum indicates a better liver parenchyma condition in the group E where no increased activity was identified above the reference range. An increased activity was identified in the group C from 2nd week with a nd significant difference between the groups detected in 2 (0.41±0.07 vs. -1 th -1 0.66±0.25 -kat.l ; p<0.05) and 4 (0.42±0.05 vs. 0.64±0.25 -kat.l ; p<0.05) weeks after delivery. There were no significant differences found between groups in bilirubin and aspartate aminotransferase but a general tendency toward lower values is apparent in the group E. Based on these results we can conclude that feeding higher amounts of full-fat soya (22% in concentrates) does not have a negative effect on the dairy cow's metabolism. Although lipomobilization syndrome has developed in both groups, a lower level of liver affection was diagnosed in the dairy cows, which was fed heat-treated full-fat soybean seeds. The work was realized with the support of MSM Project No. 6215712402. Key words: cattle, fat 470 Relationship between Metabolic Factors and Blood Chemistry Level of Domestic Cow in Korea YJ. Do , TY. Hur , IS. Ryu , KW. Lee , SJ. Kang , CY. Choi , 1 1 3 YH. Jung , DS. Son , HR. Han 1 National Institue of Animal Science, Animal Resources Development, Cheonan, Korea, South 2 Kyung Pook National University, College of Veterinary Medicine, Daegu, Korea, South 3 Seoul National University, College of Veterinary Medicine, Seoul, Korea, South There are many factors to have effect on metabolism of cow like milk productive quantity, the number of milking days, parturition times, etc. To get the basis of the study about metabolic disease of domestic cow in Korea, we analyzed the relationship between these factors, CBC and blood chemistry level of cow using 517 blood samples from National Institute of Animal Science (NIAS) and 15 farms in Gyeonggi Province. In addition, ketone body level of 84 cows and MUN of 54 cows were detected. As a result of analyses, milk productive quantity of a year had positive relation with WBC(K/µl), MO(K/µl), EO(K/µl), Hb (g/dL), HCT(%) (p-value 0.05). MUN had high negative relation with LY(K/µl) and high positive relation with BUN, GGT, CHO.(pvalue 0.01). The number of milking days was in high positive relationship with LY(K/µl), MO(K/µl), Hb(g/dL), HCT(%) (p-value 0.01) and NE(K/µl), EO(K/µl), RBC(M/µl), PLT(K/µl) (p-value 0.05). In the milk ketone body level, 23 cows of 29 cows were in normal rage(0~0.5mg/dl) and 6 cows were in suspected positive range(1mg/dl), and then 2 suspected positive cows had clinical signs like anorexia, depression. A cow of these 2 cows was in abnormal milk ketone body level and abnormal blood sugar level and then was culled by bad prognosis. In blood sugar level, at 7days after parturition, 10 cows of 34 cows were in abnormal range and at parturition day, 6 cows of 18 cows were in abnormal range. So blood sugar level had a tendency to show high level of blood sugar at parturition day. Therefore these results are thought that can be used to compare with the aspects of metabolic diseases and for prevention and prediction of metabolic diseases we need more study from now on. Key words: metabolic disease, cow, chemistry

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471 Influence of a High Potassium Diet on Mineral Concentrations in Blood and Urine and on Indicators for Early Detection of Acid-Base Changes in Dairy Cows M. Rérat , A. Philipp , H. Hess , A. Liesegang 1 Agroscope Liebefeld-Posieux Research Station ALP, Posieux, Switzerland 2 Institute of Animal Nutrition, Vetsuisse Faculty, University Zürich, Zürich, Switzerland The aim of this study was to determine the effect of two diets, fed during the transition period and differing in their potassium content, on mineral concentrations in blood and urine around calving. Five weeks before calving, 12 cows were allocated to two groups (K35 and K15). During the prepartum period, the cows of group K35 received a diet based on hay with high potassium content (35 g/kg DM) while the cows of group K15 received a diet based on hay with low potassium content (15 g/kg DM). The two diets differed only in their potassium content. The dietary cation-anion balance (DCAB) of diets K15 and K35 was 209±18 and 492±18 meq/kg DM, respectively. Blood and urine samples were taken at the beginning of the trial; three, two, and one week before parturition; three days before parturition; at parturition, and 24h after parturition. Concentrations of minerals (Ca, P, Mg, Na, K, and Cl) were analyzed in blood and urine. Net-acid-base excretion (NABE), base-acid-quotient (BAQ), and pH in urine were determined to observe changes in acid-base homeostasis. No consistent differences in mineral concentrations in blood and urine were observed between the two groups. Only the blood concentration of Mg in group K15 increased (P < 0.05) during the last days before parturition and a significantly (P < 0.05) higher urinary excretion of potassium in group K35 from three weeks to three days prepartum was observed. Urinary pH was significantly lower in group K15 three days before parturition than in group K35. In group K15 NABE and BAQ were significantly (P < 0.05) lower two weeks, one week and three days before parturition than in group K35. These results showed that the reduction of potassium content in the diet of dairy cows before parturition induced a change in acid-base homeostasis reducing the metabolic alkalosis. The indicators NABE and BAQ showed this change earlier than urine pH and could be useful tools to detect early changes in acid-base status in the periparturient cow. 472 The Concentration of Zinc in Goat´s Milk during Short Term Supplementation of Different Forms of Zinc A. Pechová, L. Mi_urová, L. Pavlata, R. Dvorák University of Veterinary and Pharmaceutical Sciences Brno, Clinic of Ruminants, Brno, Czech Republic The aim of this experiment was to study dynamics of changes in zinc concentration in goat milk after a short-term oral supplementation of three organic and one inorganic form of zinc (Zn). Four different forms of Zn supplementation were compared on fifth groups of lactating goats (32 animals as total). The first group was used as control (C, n=7), the second (L, n=6) received zinc in the form of lactate (Zinc Chelate, Agrobac, CR), the third (A, n=6) received zinc in the form of chelate (Bioplex Zn, Alltech, USA), the fourth (T, n=6) received amino acid-polypeptidic complex with zinc (Transfer Zn 15 %) and the fifth group (O, n=6) received zinc in inorganic form as zinc oxid (ZnO). Individual forms of zinc were administered orally after the morning feeding at 500 mg Zn per head/day within the period of 30 days. Zinc concentration in milk was followed daily while its level in serum was estimated at the beginning, after 2 weeks and at the end of the experiment. At the beginning of the experiment Zn concentration in serum of goats in individual groups were as follows: C 10.6±2.5 µmol/l; L 10.7±1.4 µmol/l; A 10.4±1.2 µmol/l; T 10.7±2.1 µmol/l; O 10.5±1.5 µmol/l. In the course of the experiment Zn concentration increased significantly in all experimental groups after 2 weeks and in the control group was concentration of Zn in serum almost the same as at the start of the experiment. At the end of the experiment Zn concentration in blood serum of goats in individual groups were as follows: C 10.8±0.8 µmol/l; L 12.9±2.3 µmol/l; A 12.2±1.0 µmol/l; T 12.6±1.0 µmol/l; O 12.2±1.2 µmol/l. The concentrations of zinc in milk were at the beginning of the experiment as follows: C 37.6±9.8 µmol/l; L 31.8±9.0 µmol/l; A 38.8±3.5 µmol/l; T 35.9±5.9 µmol/l; O 37.2±7.9 µmol/l. During the experiment the concentration of zinc fluctuated in individual groups, but there were no significant differences between groups. After 30 days of Zn supplementation Zn concentration in milk of goats in individual groups were similar as at the start of the experiment. According to our results the supplementation of different form of zinc did

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not influence the concentration of Zn in milk, but significantly influenced the Zn concentration in blood serum. The efficiency of different organic and inorganic forms of zinc was almost the same. The work was realized with the support of MSM Project No. 6215712403. Key words: microelements, serum, Zn 473 The Evaluation of Coagulation Profiles in Cows with Abomasal Displacement P. Sobiech, Z. Kuleta, J. Radwinska University of Warmia and Mazury, Department of Internal Medicine, Olsztyn, Poland Abomasal displacement occurs in high-yelding cows during early lactation. The most common form of this disease is left abomasal displacement, in which the abomasum is trapped between the rumen and the left abdominal wall. The purpose of the study was to evaluate coagulation profile in cows with this disorder. Material and methods: Tests were carried out on 20 cows of H-F breed (Holstein Friesian) divided into two groups: the first group 10 cows with left abomasal displacement, the second 10 healthy cows. Diagnosis of abomasal displacement was confirmed by abdominal ausculopercussion, rectal examination and verified by operation. Laboratory tests included marking, using CoA Chrom 3003 BIO-KSEL Apparatus, such parameters as: prothrombin time (PT), fibrinogen concentration (FIB), activated partial thromboplastin time (APTT), thrombin time (TT), level of antithrombin III (AT III) and level of D-Dimer. Results: Mean values of determined parameters amounted in cattle with abomasal displacement: PT 20,43 sec., FIB 7,99 g/l, APTT 47,31 sec., TT 26,63 sec., AT III 81,33 %, D-Dim 849,33 µg/l. In healthy cows these parameters reached values as follows: PT- 25,12 sec, FIB 4,15 g/l, APTT 33,71 sec., TT 24,84 sec., AT III 111,12 %, D-Dim 286,15 µg/l. Discussion: In this study it was found that cattle with left abomasal displacement had significantly higher fibrinogen level, APTT time and extremely high D-dimer level together with significantly lower AT III level. Obtained results indicated, that ill cows showed abnormal coagulation profile which reflect the occurrence of disseminated intravascular coagulation syndrome (DIC). The laboratory diagnosis of DIC is based on prolonged APTT, PT, hypofibrinogenaemia, decreased concentration of coagulation factors, increased D-Dim level and decreased AT III activity. A diagnosis of DIC is concerning when at least three tests in coagulation profile are abnormal. According with literature data, DIC is very common complication in cattle with abomasal displacement, especially with right displacement, which is usually accompanied by a degree of volvulus. Conclusion: During left abomasal displacement in cows occurred disseminated intravascular coagulation syndrome with different level of coagulation abnormalities. 474 Bread Leftovers Utilization in Cattle Rations in Tadla Area A. Ilham, M. Mazouz, S. Alali Institut Agronomique et Vétérinaire Hassan II, Morocco, Pathologie Médicale et Chirurgicale des Ruminants, Rabat, Morocco The present study aimed to evaluate the importance of bread leftovers utilization by cattle producers in Tadla area in Morocco and to assess their nutritive value as well as their effects on dairy cow performances and diseases incidence. A follow up study was conducted during 3 months to compare the performances and the diseases occurrence in a sample consisting of 12 dairy farms among which 6 farms totalizing 51 cows used bread leftovers all the year around (F1) and the 6 others composed of 53 cows that have never used them (F2). The main results obtained are as follows: - Among all cattle producers in Tadla area, 35.8 % supplemented the rations with bread leftovers from which 16.4 % used them all the year around and 19.4 % only during periods of feed shortage. - The total quantity of bread leftovers used per year in Tadla area was estimated to about 5230 metric tons. - The incorporation rate of bread leftovers in the concentrate portion of cattle rations varied from 20 to 100 % with an average rate of 30 %. The daily quantities used varied between 2 and 6 kg per cow with an average of 2.3 kg. - The composition per kg of bread leftovers was: 88.1% DM, 0.9% CF, 11.9% CP and 3.6% Ash. The nutritive value was: 1.28 UFL (French dairy feed unit) / kg DM and 104.4g DCP / kg DM.

- There was no significant difference between F1 and F2 regarding milk production and body live weight. However, the body condition score was in favour of cows of F1 and the milk fat test was 1.27 % higher in F2. - The incidence of health problems was higher in F1 compared to F2. Thus, abortion, acidosis and bloat rates were 15.7 vs. 7.4 %, 9.8 vs. 3.7 % and 11.8% vs. 5% respectively in F1 and F2.65 Key words: bread leftovers, nutritive value, dairy cattle, performances, health problems 475 Influence of Diarrhoea on Chosen Indicators of the Blood Coagulation System at Dairy Cattle J. Radwinska, Z. Kuleta, P. Sobiech University of Warmia and Mazury, Department of Internal Medicine, Olsztyn, Poland Objectives of study: In cattle breeding important is prevention and treatment of diarrhoeas. There are many factors causing diarrhoea. The aim research was evaluation of diarrhoea influence on chosen indicators of the blood clotting system at the dairy cattle. Material and methods: Research was passed on 2 groups of dairy cows. In group I were 16 cows with diarrhoea. Group II - 20 healthy cows (control group). The blood were taken from jugular vein. Coagulation factors were marked in blood plasma. To the laboratory analysis was used photometer Coag-Chorm 3003 (Bio-Ksel). During research were marked: prothrombin time(PT), thrombin time(TT), activated partial thromboplastin time(kaolin-kephalin time, APTT), fibrinogen, antithrombinIII(ATIII) and D-dimer. During research was not identify primary reason of diarrhoea. Results: There were changes in times length between ill and healthy cows. PT at cows with diarrhoea was longer than in healthy. There was an APTT extension (mean APTT at group I - 37,675s and group II 34,24s). In group I a mean TT was 31,39s in group II 26,44s. Fibrinogen concentration at ill cattle increased (6,61g/l) to healthy cattle (5,24g/l). Mean value of D-dimer in group I and II differed about 103 µg/l. ATIII in each groups were: group I 108,7% and group II 107,7%. PT describes exogenous tract and APTT characterize endogenous track of the blood clotting. The extension of PT is observed in deficiency of II, V, VII, X factors. All mentioned factors are dependent on vitamin K. A length of APTT is connected with II, V, X, VIII, IX, X, XI, XII factors. The PT prolongation might be connected with vitamin K malabsorption. This can be induced by changed bacterial flora in intestines (production vitamin K) or handicap absorption from alimentary canal (damages at intestine epithelium during infections). An increased D-dimer concentration (product of digestion stabilized fibrin) and TT prolongation (measures activity of fibrinogen) at the group I were connected with long PT and APTT. This can be an evidence of disseminated intravascular coagulation (DIC) in cow organism. Increased concentration of fibrinogen is DIC laboratory symptom. The activity of ATIII did not differ at sick and health cattle. Conclusions: Diseases with diarrhoea have influence on whole organism also on blood coagulation system. Abnormality in this system can lead to serious problems like DIC. 476 Diagnostic Use of Pooled Metabolic Profiles in Czech Dairy Herds J. Davidek , R. Van Saun 1 Practitioner, Krasna Hora nad Vltavou, Czech Republic 2 Penn State University, Veterinary & Biomedical Sciences, University Park, United States Study objective was to compare blood analyte concentrations collected at different periods relative to calving to determine diagnostic value in herd-based assessment of transition cow problems. In the course of clinical investigations on 37 dairy herds presented for various periparturient diseases or routine monitoring, blood was sampled from 3 to 7 mature cows within defined time periods to run a pooled sample metabolic profile. Time periods were defined as (days relative to calving): Early Dry (>30 d prior), Close-up (<21 d prior) and Fresh (350 following). Analytes determined in metabolic profiles included urea nitrogen (UN), glucose (Glu), albumin (Alb), total protein (TP), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), sodium (Na), potassium (K), chloride (Cl), calcium (Ca), phosphorus (P), magnesium (Mg), total cholesterol (Chol), ,-hydroxybutyrate (BHB) and nonesterified fatty acids (NEFA). A wide range of health and feed-related problems were identified and used to assign collective

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health group (HG) categories for analysis. Blood analytes were analyzed by ANOVA with main effects of period, HG and their interaction with herd as a covariate. Herds ranged in size from 100 to 750 cows with Holstein and Simmental being the predominate breeds. A total of 40 metabolic profiles were performed. Herds were categorized into HG of no problems (NP, n=6), heat stress (HS, n=6), fresh cow diseases (FD, n=16), low production (LP, n=5) and other diseases (OD, n=7). Time period influenced (P<.0001) NEFA, Glu, Chol, BHB, TP, Na, Cl and AST concentrations. Health group influenced NEFA (P=.003), Glu (P<.0001), Chol (P=.01), Alb (P=.0003), TP (P=.04), Ca (P=.0003), K (P<.0001) and UN (P=.01) concentrations. Both AST and P tended (P=.09) to be influenced by HG. Of interest was the absence of HG effect on BHB concentration, though there was a tendency (P=.06) for a period by HG interaction. No other interactions were found. Within each period, NP herds generally had lower NEFA and TP and higher Glu, Alb, Ca, Chol and K compared to other HG herds, though mean comparison significance varied by HG within and across time periods. Study results suggest pooled samples can provide diagnostic information in detecting periparturient health problems. Variation in HG effects on blood analytes over time relative to calving suggests potential analyte patterns that may provide herd-based disease diagnostic criteria. Key words: metabolic profile 477 Characterizing Analyte Patterns in Pooled Metabolic Profiles for Disease Diagnosis J. Davidek , R. VanSaun 1 Practitioner, Krasna Hora nad Vltavou, Czech Republic 2 Penn State University, Veterinary & Biomedical Sciences, University Park, United States Study objective was to compare blood analyte concentrations collected at different periods relative to calving to determine diagnostic value in herd-based assessment of transition cow problems. Forty metabolic profiles were performed on 37 dairy herds presented for various periparturient diseases or routine monitoring. Herd size ranged from 100 to 750 cows with Holstein and Simmental being the predominate breeds. Blood was sampled from 3 to 7 mature cows within defined time periods to run a pooled sample metabolic profile. Time periods relative to calving day were defined as: Early Dry (ED, >30 d prior), Close-up (CU, <21 d prior) and Fresh (FR, 3-50 d following). Analytes determined in metabolic profiles included urea nitrogen (UN), glucose (Glu), albumin (Alb), total protein (TP), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), sodium (Na), potassium (K), chloride (Cl), calcium (Ca), phosphorus (P), magnesium (Mg), total cholesterol (Chol), ,-hydroxybutyrate (BHB) and nonesterified fatty acids (NEFA). Based on clinical diagnosis, herds were categorized into health groups (HG) of: no problems (NP, n=6), heat stress (HS, n=6), fresh cow diseases (FD, n=16), low production (LP, n=5) and other diseases (OD, n=7). Blood analytes were analyzed by ANOVA with main effects of period, HG and their interaction with herd as a covariate. Paired comparisons between NP and other HG herds were performed to characterize analyte pattern differences. Herds experiencing HS showed the most alterations in blood analyte concentrations having higher NEFA (P<.0001) and Cl (P=.005) and lower Gluc (P<.0001), Chol (P=.001), Alb (P=.0005), Ca (P<.0001), P (P=.02) and K (P<.0001) concentrations across periods. Both BHB (P=.03) and AST (P=.04) were higher in HS herds only in the FR period. Herds with FD showed higher NEFA (P=.008), BHB (P=.04) and AST (P=.05) and lower Alb (P=.0001). A period by HG interaction (P=.005) showed higher FR period BHB concentrations in FD herds. Low production herds showed only differences with lower Chol (P=.02) and Urea (P=.0007) in ED and CU periods and higher AST (P=.02) in CU period. Health effects of OD found higher CU and FR NEFA (P=.0001) and ED and CU TP (P=.02) compared to NP herds. Results of this study suggest differential patterns of pooled blood analyte changes around the time of calving are associated with specific disease conditions and have herd-based diagnostic potential. Key words: metabolic profiles, periparturient disease 478 Interpretation of Pooled Metabolic Profiles for Herd Assessment J. Davidek , R. VanSaun 1 Practitioner, Krasna Hora nad Vltavou, Czech Republic 2 Penn State University, University Park, United States

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Study objective was to evaluate diagnostic reference criteria for determining if pooled sample blood analyte concentrations can differentiate dairy herds with and without health problems. Forty metabolic profiles were completed on 37 dairy herds presented for various periparturient diseases or routine monitoring. Blood samples were collected from 3 to 7 mature cows during defined time periods relative to calving and pooled prior to analysis. Time periods relative to calving day were defined as: Early Dry (ED, >30 d prior), Close-up (CU, <21 d prior) and Fresh (FR, 3-50 d following). Analytes determined in metabolic profiles included urea nitrogen (UN), glucose (Glu), albumin (Alb), total protein (TP), aspartate aminotransferase (AST), gammaglutamyltransferase (GGT), sodium (Na), potassium (K), chloride (Cl), calcium (Ca), phosphorus (P), magnesium (Mg), total cholesterol (Chol), ,-hydroxybutyrate (BHB) and nonesterified fatty acids (NEFA). Based on clinical diagnosis, herds were categorized into health groups (HG) of: no problems (NP, n=6), heat stress (HS, n=6), fresh cow diseases (FD, n=16), low production (LP, n=5) and other diseases (OD, n=7). For each analyte measured, the pooled sample value was subtracted from a herdbased reference value and divided by the analyte's standard deviation (SD) for a given time period. T-test was used to determine if deviation was different from zero. Analyte deviations were analyzed by ANOVA with main effects of period, HG and their interaction with herd as a covariate. Within NP herds, analyte deviations different (P<.05) from zero within periods were NEFA (ED, FR), Na (CU, FR), Cl and AST (CU) and Glu, P, Mg and K (FR). Mean analyte deviation for ED, CU and FR periods for NP herds were .29, .43 and .37 SD, respectively. These deviations were analyte-specific and potentially influenced by period. Collectively, HG influenced deviation of NEFA (.85 SD, P=.002), UN (-.62 SD, P=.001), Alb (-.48 SD, P=.0003), Ca (-.29, P=.007) and AST (1.3 SD, P=.02) with a HG by period effect on BHB (FR, 1.1 SD, P=.02). However, significance, magnitude of analyte deviation and period effects were specific to and differed between HG. Results suggest evaluation of pooled samples based on number of SD the sample value deviates from herd-based population reference values have diagnostic potential. Pattern of analyte deviation and period might provide insight to specific disease present in a herd. Key words: metabolic profile, diagnostic 479 Intake, Apparent Digestibility, Ruminal and Blood Components of Holstein Cows Fed Extruded Ground Canola with or without Lignosulfonato G. Santos , C. Neves , D. Silva , W. Santos , C. Jobim , 1 3 A. Branco , F. Santos 1 Universidade Estadual de Maringá, Zootecnia, Maringá, PR, Brazil 2 Universidade Estadual de Maringá, Programa de Pós-graduaçio em Zootecnia, Maringá, PR, Brazil 3 Universidade Estadual de Maringá, Student, Maringá, PR, Brazil The objective of this research was to evaluate the Holstein cows intake, digestibility, ruminal parameters and blood components, fed with ground canola (GC), extruded ground canola (EC), ground canola with Lignosulfonato (GCL) and extruded ground canola with lignosulfonate (ECL). Eight cows, divided in a double Latin square, with four treatments and four periods with 21 days each, were utilized. Intake and total apparent digestibility of nutrients were estimated, while rumen volatile fatty acids (VFA), ammonia N-NH3 and pH were evaluated. Differences were not observed in the dry matter (DM), crude protein (CP), neuter detergent fiber (NDF) and acid detergent fiber (ADF) intake, however, the extruded treatments presented larger ingestion of ether extract (EE). We can still mention that the association between the extrusion and the lignosulfonato in the canola seeds stimulated the consumption of Non-Structural Carbohydrates (NSC). Another parameter that suffered alterations was the apparent digestibility, which was smaller for the animals fed with CGCL in comparison with the CGC. However, it was not observed alterations in the DM, OM, CP, ADF, NDF and NSC digestibility for EGC and EGCL treatments. For rumen components, such like N-NH3 (mg/dL), VFA and pH and also blood parameters, any alteration was not observed in the cows fed with GC, EC, GCL and ECL treatments. In conclusion that there are not differences that justify the canola seeds processing or it lignosulfonate treatment, since the results obtained with the crushed canola didn't differ a lot of the other treatments, even if the processing of the canola would have an extra cost for the producer. The project was supported by the Fundaçao Araucaria, Curitiba, PR and CNPq, Brasilia, DF, Brazil Key words: extrusion, fat, heat treatment, chemical treatment

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24 XXV. Jubilee World Buiatrics Congress 2008

480 Mineral Metabolism during Late Pregnancy and Calcium Status after Parturition in Dairy Cows M. Casalone , C. Cannizzo , A. Stefani , L. Moro , 1 1 M. Gianesella , M. Morgante 1 Universita'degli Studi di Padova, Dipartimento di Scienze Cliniche Veterinarie, Legnaro (PD), Italy 2 Istituto Zooprofilattico Sperimentale delle Venezie, Legnaro (PD), Italy Milk fever represent one of the most important economic losses in dairy cows management, therefore to found some indicators (blood parameters or functional test) before calving should be very important for the prevention of hypocalcaemia and related disorders. The objective of this work was to study some blood and urinary excretion parameters during late pregnancy that could be related with the calcium status after parturition. 60 dairy cows from a farm located in the North of Italy with high incidence of milk fever were monitored 7 (±2) days before calving and 24h after calving: blood and urine samples were simultaneously collected for creatinine (Cr), calcium (Ca), phosphorus (P), magnesium (Mg), sodium (Na) and potassium (K) determination. Fractional excretion (%CrX) of electrolytes were calculated by the following formula: %CrX = Xu/Xp x Crp/Cru x 100 where "u" and "p" were the urinary and plasmatic concentration, respectively. Results were subject to ANOVA and correlation analysis using SIGMA STAT 3.05. Blood level of Ca after parturition show only a significant (P < 0.05) correlation (+ 0.490) with the blood level of Mg during late pregnancy but the results indicated that there were several significant correlation between the urinary level of Ca after parturition and some parameters during late pregnancy: in particular with the blood level of P (+ 0.378), with the urinary level of Ca (+ 0.384), P (+ 0.401) and Mg (+ 0.277) and with the fractional excretion of Ca (+ 0.360) and P (+ 0.374). These data suggest that modifications of some blood and urinary parameters during late pregnancy could be related with the calcium metabolism after parturition. Although no animal developed any hypocalcaemia status post partum, this data suggest that it could be very important to do further studies especially on Ca and P excretion before calving to found useful indicators for risk evaluation of single cows regarding the probability to develop hypocalcaemia and related disorders after parturition. 481 Effect of Glycerol-containing Preparation on Changes in Selected Parameters of Dairy Cows in Experimental and Field Conditions Z. Farkasova , P. Reichel , K. Kovacocyova , M. Fabini , 2 1 A. Bobus , G. Kovac 1 University of Veterinary Medicine, Department of Internal Medicine, Kosice, Slovakia 2 Polychem s r. o., polychem, Prievidza, Slovakia The aim of our study was to determine the effects of glycerol-containing preparation on changes in selected parameters in rumen fluid (volatile fatty acids - VFA, acidity - pH), blood serum (glucose, total cholesterol - TCH, total lipids - TL, triglycerides - TG, nonesterified fatty acid - NEFA, betahydroxybutyrate - BHB), and milk (fat, protein, urea) of dairy cows. The observation was aimed at prevention of lipomobilisation syndrome of cows during peri-partal period. In experimental conditions we determined optimal dose of glycerol from three selected concentrations. We used 150, 200, 300 g/day/cow in 24 dairy cows. In the third experimental group (300g) the positive effects were recorded. The following observations were done in herd of 210 Holstein-Friesian dairy cows kept in loose housing with annual milk yield of 8500L. Glycerol bound to powder vehicle was added to the diet at the dose of 320g per animal and day for 35 days (3 weeks before and 3 weeks after parturition). To compare glycerol effects on health and production, 12 experimental cows were selected and divided retrospectively by date of calving into two groups E1 (n = 6) and E2 (n = 6), and by the period of glycerol intake before and after parturition (E1<25 days a. p. and 10 days p. p.; E2<8 days a. p. and 27 days p. p.). The results of analyses were compared with control group (n = 6) fed the same diet without glycerol addition (C<7 days a. p. and 28 days p. p.). Samples (blood, rumen fluid, milk) were collected three times: I. 3 weeks a. p., II. th th th 7 - 10 day p. p., III. after 30 day of glycerol feeding. We expected the positive effect of glycerol (as a glucoplastic substance) on changes of pH, as well as on decrease of acetate to propionate ratio, the NEFA and BHB concentrations, and increase in glucose concentrations in blood serum. The results indicated antilipolytic, hepatoprotective, and glucoplastic effects of glycerol, particularly in the group with longer glycerol administration in

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post-partal period (E2) compared with the control group. Glycerol addition to the diet showed also favourable effects on quantitative and qualitative parameters of milk production and milk profile. Key words: dairy cows, glycerol, milk production, prevention of lipomobilisation syndrome 482 Preventive Diagnosis in Dried Cows on Two Farms with the Monitoring of Clinically Manifested Problems during the Lactation Phase. V. Petrovic, G. Kovac, P. Reichel, P. Mudron, O. Nagy, H. Seidel, R. Link, I. Pavlikova, J. Novotny, V. Hisira University of Veterinary Medicine, Department of Cattle and Sheep Diseases, Kosice, Slovakia The aim of this study was to compare some parameters obtained from preventive diagnosis of Slovak spotted cows (Ssc) and HolsteinFriesian cows (HFc) in dry period with the monitoring of clinically manifested problems during the lactation phase. The housing, nutrition, zootechnical conditions, prevention and treatment of locomotion and reproductive problems were similar on both farms but milk yields were up to 7 l higher on a farm with HF herd. Ten healthy cows in dry period were preventively diagnosed in each herd and clinically monitored in lactation phase to intercept the productive and reproductive problems. The preventive diagnosis included enzymatic, hepatic, protein, energy, vitamin, element, rumen, urine profiles. The lower AST, GMT activities were found in Ssc only and increased LDH activity in 80% of Ssc vs 20% of HFc. The increased albumin was determined in 40% of Ssc only, whereas increased TIg in all vs half of animals in a herd. Lower concentrations of vitamins and Ca were found in cows on both farms, but with up to 50% of HFc. Concentration of Fe was decreased in probably half and Cu, Se in all of animals on both farms. Disproportion of Mg and Zn was found in more then 50% of Ssc only. Plasma Glu was higher in 80% vs 30% and NEFA in 20% of animals in both herds, whereas amount of TL was higher in 20% of Ssc only. Rumen pH was increased in 60% vs 20% of animals in a herd as well as butyrate. Lower propionate was assessed in all cows, but 80% of Ssc had NEFA beyond the physiological range. Acetate and lactate concentrations were higher in HFc only. The C2:C3 ratio was enhanced in rumen fluid of all tested cows. Urine pH was elevated in more then 60% of animals, but the incidence of increased level of NABE was found 3 times higher in herd of HFc. Elevated concentrations of Ca, Na, K and Mg in excreted urine were found in up to 50% of Ssc vs 20% of HFc for each parameter. The clinical monitoring of cows in lactation phase revealed the frequent occurrence of retained placenta with subsequent endometritis in cows on both farms. Higher concentration of BHB was determined in 20% of HFc only, but without any signs of clinical manifestation of acute ketosis. In conclusions, the comparison of parameters from preventive diagnosis on both farms revealed better physiological status of HFc in the dry period but the high production of milk, as mentioned above, resulted in subclinical ketosis during the lactation phase. Key words: dried cows, preventive diagnosis 483 Study the Effects of Dry Period Length on Reproductive Indexes after Following Parturition M. Mohammadsadegh , S. Lotfolllahzadeh , AR. Bahonar , 1 M. Gahry 1 Islamic Azad University, Garmsar branch, Large Animal Clinic for Theriogenology and Ambulatory Services, Garmsar, Iran 2 Islamic Azad University, Garmsar branch, Large Animal Clinical Sciences, Garmsar, Iran 3 Facutly of Veterinary Medicine, University of Tehran, Food Hygiene, Tehran, Iran Most of the literature indicates that a dry period (DP) of 40 to 60 d is needed to achieve maximum milk yield during the following lactation (Coppock et al., 1974; Sorensen and Enevoldsen, 1991; reviewed by Bachman and Schairer, 2003). Other research indicates that a DP of 30 d is sufficient to maintain milk (Lotan and Adler, 1976; Bachman, 2002; Gulay et al., 2003). Nevertheless, a shortened DP seemed to reduce milk production in primiparous, but not multiparous, cows in a recent study (Annen et al., 2004) and our recent study found somewhat of a reduction in milk production per day, but not in FCM production (Rastani et al., 2005). Information relating the effects of DP to subsequent reproductive performance of

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lactating cows is sparse. Lotan and Adler (1976) reported that days open, number of inseminations, and pregnancy rates for 18 pairs of dairy cows having DP of 30 and 60 d were numerically similar. Remond et al. (1997) reported that BW of cows with no planned DP increased 24 kg in the first 60 DIM, whereas BW of cows with a 60d DP lost 28 kg during same time period. We speculated that cows with a DP of 56 d would be in more negative EB than cows with no planned DP due to reduced milk production in cows with no planned DP. Thus, cows with no planned DP were expected to ovulate earlier than cows with a 56-d DP; whereas cows with a 28-d DP were expected to be intermediate in days to first ovulation based on an expected intermediate level of negative EB. At the base of the importance of dry period management on the following milk production, mastitis rates, reproductive indexes and normal life of cows in after following parturition, the effects of different lengths of dry period on reproductive indexes were to be evaluated .One hundred Holstein dairy cows from 10/5/85 to 25/6/86 were selected from Bonyad Janbazan Shohada dairy farm in Razan Hamadan (Iran). In depended variable was the length of dry period and depended variables were reproductive indexes (1- parturition to first service, 2- days open, 3- numbers of service to pregnancy, 4mastitis rate on the first month of lactation). Control variables were 1- pick of milk production, 2- BCS at the breeding, 3- number of lactation. Fisher Exact test and General Linear Model were used to statistical analysis. It is concluded that there was no correlation between dry period length and reproductive indices. Key words: dry period length, reproductive indices, cattle 484 The Use of Calcium Chloride and Calcium Sulfate in an Oral Bolus as a Supplement for Post-partum Dairy Cows J. Spain , J. Sampson , L. Carstensen , C. Jones 1 University of Missouri, Department of Animal Science, Columbia, United States 2 Boehringer Ingelheim Denmark, Division Vetmedica, Copenhagen, Denmark 3 Boehringer Ingelheim Vetmedica Inc, St. Joseph, United States Objective: The onset of lactation results in a sudden demand for calcium and imposes challenges to calcium homeostasis in cows. Cows unable to adapt to the calcium demand, develop hypocalcemia (incidences 23-39 %), which may manifest itself as milk fever (5-10%). This also increases the risk of periparturient disorders. With systemic acidification, PTH sensitivity is increased resulting in enhancing calcium mobilization from bone to blood. When the systemic pH is effectively decreased, urine pH values (pH 7.8) will decrease to a range between 6.2 and 6.8. The objective of this study was to investigate the effects of Bovikalc® (Boehringer Ingelheim, St. Joseph, USA), an oral calcium supplement containing CaCl and CaSo4, on calcium homeostasis during the first 24 h after calving. Materials and Methods: Multiparous Holsteins (n = 20) were blocked by parity and day of calving and randomly assigned to either Control (C) group receiving no calcium supplement or treated (B) group receiving one bolus directly after calving and a second bolus 12 h later. Blood samples were drawn at 48 and 24 h before calving. Ionized blood calcium (iCa) was measured using an IDEXX Vet Stat Analyzer. Cows with an iCa level of 1.10 mmol/L or less were included in the study. Blood and urine samples were collected at 0, 1, 6, 12, 13, and 24 h post partum. Blood iCa and pH and urine pH were measured. Results: No differences in iCa levels were found between C and B groups (1.24 vs. 1.22; 1.19 vs. 1.18 mmol/L) at -48 and -24 h, respectively. iCa levels were similar at calving (0h, 0.95 vs. 0.94 mmol/L) for C and B, respectively (p = 0.84). Thereafter, iCa levels were significantly higher in cows given the bolus as described (p = 0.02). Urine pH differed with treatment (p = 0.002) and by treatment over time (p = 0.003). It decreased in B cows from 7.58 (0h) to 6.79 (24h) compared to 8.00 (0h) to 8.09 (24h) for C cows. Blood pH did not change (p = 0.27) which reflects a tight homeostatic regulation of blood pH. Calcium supplementation with Bovikalc® after calving decreased urine pH and increased iCa levels compared to control cows. Conclusions: By 2 administrations of Bovikalc® with its two calcium components it was possible to increase the iCa level in blood of hypocalcemic cows significantly. Additionally it was shown that Bovikalc® had a systemic acidifying effect which most likely enhances mobilization of the cow's own calcium. Key words: dairy cows, hypocalcemia, calcium supplement

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485 The Microelements (MCEs) Supplementation by Mineral Lick (ML) and the Level of MCEs in the Blood, Colostrum and Milk of Dairy Cows S. Krys, A. Podhorsky, E. Lokajova, L. Pavlata University of Veterinary and Pharmaceutical Sciencies Brno, Clinic for Ruminants, Brno, Czech Republic The aim of this experiment was to verify the option of providing MCEs supplementation to dairy cows (DC) in the dry period through supplemented ML, and to then compare the content of MCEs in the blood, colostrum and milk of supplemented and of non-supplemented DC. The experiment was carried out on Czech Pied DC. We formed an experimental group (E, n=12), which was supplied with ML ad lib. enhanced with organic forms of MCEs (Se, Zn, Cu, Mn) for 4 to 5 weeks before parturition and 1 week after parturition (a.p.). The control group (C, n=10) did not get any ML. Samples of blood were th th taken 14 days before calving, on the calving day, 4 and 20 day a.p. Samples of milk and colostrum were taken on the day of parturition (day 0) and then on days 1, 2, 3, 4 and 20 a.p. Reception of ML by DC was very uneven, its total consumption ranged from 0 to 0.25 kg/animal/day. ML feeding did not result in evidential increase of concentrations of MCEs in DC blood. Only a trend to increased concentrations of Se and Cu in blood of E group was observed, where the average Se concentration increased by 60 µg/l (Cu by 9 µmol/l), while in C group, Se increased only by 30 µg/l (Cu by 6 µmol/l). Neither concentration of monitored MCEs in colostrum and milk pointed to a conclusive difference between groups (average concentration of MCEs in first colostrum in groups E and C was as follows: Se 115 vs. 102 µg/l; Cu 2.0 vs. 1.6 µmol/l; Zn 317 vs. 363 µmol/l; Mn 0.6 vs. 0.4 µmol/l). The concentration of MCEs in colostrum a.p. dropped quickly. Significant drops were determined for Se, Cu and Zn on the first (for Mn on the second) day a.p. Apart from Mn, all monitored MCEs showed significantly lower (p<0.05) concentration in milk (in groups E and C, concentrations were as follows: Se 21 vs. 22 µg/l; Cu 0.6 vs. 0.75 µmol/l; Zn 79 vs. 78 µmol/l; Mn 0.6 vs. 0.44 µmol/l) than in the first colostrum. Supplementation of MCEs to DC through ML thus does not appear to be a proper method of supplementation, as the intake of ML is very uneven and, in the E group, it did not ensure a conclusive increase in concentrations of the MCEs in blood, colostrum and milk. Se concentration in the first colostrum is 5 times higher than in milk, Cu 3 times, Zn 4 times and Mn 1.3 times higher. The work was supported by the Research Project MSM6215712403. 486 Preliminary Results from an Ongoing Field Study on Study on the Effect of Catosal in the Treatment of Sublinical Ketosis in Cows P. Sarasola , U. Aramendi , U. Spiecker-Hauser , B. Schmidt 1 Ondax Scientific, CRO, Hondarribia, Spain 2 Bayer HealthCare, R&D, Monheim, Germany

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Objective: The aim of the present investigation was to assess the efficacy and safety of Catosal® in the treatment of subclinical ketosis in cows. Materials and Methods: At the time of writing this abstract, there is an ongoing multi-centre, placebo-controlled, randomised and masked field study being conducted to evaluate the effects of Catosal® in the treatment of subclinical ketosis in dairy cows. In this study, cows from several dairy farms in Spain are screened by milk test (Ketolac BHB®, Biolab GmbH) within two weeks after calving. Cows with a positive response (i.e. beta-hydroxybutyrate (BHB) milk levels >200 µmol/l) and absence of clinical signs of ketosis are enrolled in the study and are randomized to one of three treatment groups: Two injections of Catosal® (T1), one injection of Catosal® and one of saline (T2), two injections of saline (T3), administered intravenously on Days 0 and 3, respectively, at a dose volume of 5 ml/100 kg bodyweight. Blood samples for assessments of changes in BHB, glucose, glutamate dehydrogenase (GLDH), and cholesterol are collected on Days 0, 3, and 6, respectively. From this study, a subgroup of 17 animals showing baseline serum BHB levels of >3.2 mmol/l and absence of clinical signs of ketosis was selected for an interim analysis. Results: The primary efficacy criterion was the treatment success, defined as the proportion of animals with serum BHB levels of >1.2 mmol/l on days 3 and 6 in each treatment group. Results showed that 30% and 60% of the animals in T1 were classified as treatment successes on days 3 and 6,

26 XXV. Jubilee World Buiatrics Congress 2008

respectively. In contrast, none of the animals in T2 and T3 was classified as treatment successes. This effect was confirmed by statistically significant (p<0.01) treatment differences on the evolution of serum BHB and glucose levels over time. Serum levels of GLDH and cholesterol did not show treatment-dependent changes in this preliminary analysis. All treatments were well tolerated. No treatment-related adverse events were observed. Conclusion: These preliminary results indicate that two injections of Catosal® given twice at an interval of 3 days is safe and effective in the treatment of subclinical ketosis in cows. Key words: Catosal, subclinical ketosis 487 Comparison of Different Forms of Glycerol Additives and their Effects on Selected Metabolic Parameters of Dairy Cows P. Reichel , Z. Farkasova , K. Kovacocyova , M. Fabini , 2 1 A. Bobus , G. Kovac 1 University of Veterinary Medicine, Department of Internal Medicine, Kosice, Slovakia 2 Polychem s r. o., Polychem, Prievidza, Slovakia Glycerol as a glucoplastic substance is used for the prevention of nutritional deficiencies in dairy cows, predominantly during transition period till the top of lactation. The experiments were performed on two farms with dairy cows during transition period. We studied the effects of long-term feeding of different forms of glycerol additive on prevention of lipomobilisation syndrome of dairy cows. The selected metabolic parameters in blood serum and rumen fluid, and their effects on milk production and feed intake were evaluated: in the rumen fluid (volatile fatty acids - VFA, acidity pH, acetic acid, propionic acid, acetate to propionate ratio), in the blood serum (glucose, total cholesterol - TCH, total lipids - TL, triglycerides - TG, nonesterified fatty acid - NEFA, betahydroxybutyrate - BHB), and milk (milk-yield, fat, protein, urea). The experiment was done for 35 days (3 weeks before and 3 weeks after parturition). Forty-two dairy cows were divided in two experiments. In the first experiment with liquid form of glycerol additives 24 cows were divided in the control (n=12) and experimental group (n=12). In the experiment with powder form of glycerol 18 cows were used: control (n=6), first experimental (n=6) and second experimental group (n=6). The liquid form of glycerol additive was poured on to the food directly (300ml/two times of the day/cow) whereas the glycerol supplement in powder form was admixed to diet (320g/cow/day). We found positive antilipolytic, glucoplastic properties and the beneficial effects on health of cows (significantly reduced negative energetic balance, increased food intake), as well as on quantitative and qualitative parameters of milk production. In conclusion, the both forms of glycerol additive had similar positive effect on the selected metabolic parameters in the blood serum and rumen fluid, on milk production, and feed intake. The powder form of glycerol is better for its positive properties, practical application, manipulation, and buffer capacity of zeolite, which is interesting for practical exploitation in field conditions. Key words: glycerol, dairy cows, prevention of ketosis, lipomobilisation syndrome 488 Comparation of Clinical and Biochemical Obseravations in Cows with Physiological and Pathologic Course of Postpartal Period J. Bires, M. Huska University of Veterinary Medicine, Department of Cattle and Sheep Diseases, Kosice, Slovakia In the work there is a comparison of the results of clinical and biochemical observations in cows during prepartal and postpartal period with physiological and pathologic course of postpartal period. Examinations were carried out on 21 cows in time of 14 days before expected parturition to 10 days of lactation. Clinical picture including conditional state of cows before and after parturition and laboratory examinations (enzymatic, protein, energetic, hepatic and mineral profile) was evaluated. The results of observations determinated diagnostic value of clinical and laboratory examinations in determination of course postpartal period in regard to production and reproduction activities, disease of cows, resp. They also showed on limitation their biological interpretation in common field praxis, which could lead to incorrect results during application of metabolic profile test and subsequent non-adequate recommendation in the farm of cows. Key words: dairy cows, parturition, clinical and biochemical observation

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489 Non-Surgical Treatment of Abomasal Displacements with Zeolite in Cattle G. Aksoy, H. Biricik, I. Çamkerten, A. Da_ Harran University, Faculty of Veterinary Medicine, Sanliurfa, Turkey In this study; clinical efficacy of zeolite mineral in the medical treatment of left and right abomasal displacements was investigated. Seven cows (five with right displaced abomasum, two with left) were used. Diagnosis was based on routine clinical examinations. Laboratory findings before treatment were hypochloremia, hypokalemia, and hemoconcentration in three cows with right displaced abomasum. Zeolite was administered orally for 3 days (twice a day, 2 g/kg) via a stomach tube. Patients were clinically examined including ping effect, appetite and ruminal movements following zeolite administration. Blood samples for biochemical analysis were collected at 3rd day of the treatment. Furthermore, zeolite (50 g/day) was added to the ratio during 1.5 months to prevent recurrence. The ping effects on the flank had disappeared completely following fourth application in six cases. Clear rumen motility increased after the second application and the cows started consuming food. Biochemical parameters and defecation returned normal 72 hours after the treatment. Any recurrence was encountered during 1.5 months. In a cow with right displaced abomasum, treatment was unsuccessful because of adhesion to the right abdominal wall. As a result, zeolite mineral may be part of the therapy for displaced abomasums. Addition of zeolite to the ratio can prevent recurrence of displacements. Further studies need to investigate the efficacy of zeolite mineral on medical treatment of abomasal displacements. Key words: zeolite, abomasal displacement, cattle 490 Control of Ruminal pH in an Acute Acidosis Model J. Keane , L. McLaughlin , P. Thompson , G. Greenwood , I. Bruce 1 Pfizer Animal Health, Veterinary Medical Research and Development, Sandwich, United Kingdom 2 Pfizer Animal Health, Veterinary Medical Research and Development, Kalamazoo, Michigan, United States

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Sub acute ruminal acidosis (SARA) in beef cattle is associated with decreased productivity, including decreased growth rate. Metabolism of high carbohydrate rations increases the concentration of ruminal VFAs and ruminal lactate resulting in ruminal pH below 5.5 (SARA) or 4.5 (acute acidosis). Acarbose is a pseudo-tetrasaccharide that inhibits -amylase and glucosidase enzymes thus reducing the rate of carbohydrate digestion and was hypothesized to prevent acidosis in an acute challenge. Acute acidosis was induced in ruminally fistulated steers by intraruminal administration of a high carbohydrate load (12.5 g/kg liveweight; 48.4% ground corn, 48.4% corn starch, 2.1% sodium caseinate and 1.1% urea) mixed in 2-3 gallons of water. Ruminal pH was measured every 15 min after the challenge using a continuous recorder for the subsequent 24 hrs and rumen fluid samples were collected at 3 hour intervals for 12 hours. When acute acidosis, defined as ruminal pH <4.5 and lactate >50 mM, was observed, ruminal contents were removed, the rumen was washed out and then inoculated with rumen contents from a donor animal. If acidosis was not observed within 24 hours of the first challenge, a second challenge was performed and the same procedure was followed for the subsequent 24 hours. When administered in the carbohydrate load at a dose of 1.1 mg/kg body weight, acarbose protected against acidosis in 6/7 animals compared with 3/7 control 2 animals (c = 2.80, P<0.10). The equivalent amount of acarbose mixed in a small amount of total mixed ration consumed voluntarily, protected against acidosis induced by the carbohydrate load in 4/6 2 animals compared with 1/7 control animals (c = 3.75, P<0.10). In both experiments, acarbose decreased ruminal total VFA concentrations and percent propionate. In a third experiment using the same model, the efficacy of acarbose (1.1 mg/kg bodyweight, n=8) was compared with that of monensin (12 mg/kg dry matter intake, n=8), sodium bicarbonate (1% dry matter intake, n=7) and negative control (n=7). Acute acidosis was induced in all control, monensin and sodium bicarbonate treated groups whereas acarbose protected against acidosis in 5 of 8 animals (P=0.03 for all comparisons to acarbose). It was concluded that acarbose effectively controlled ruminal pH in this acute challenge model and was more efficacious than monensin or sodium bicarbonate.

Nutrition and Metabolic Disorders 27

491 Effect of Acarbose on Milk Yield and Composition in Early Lactation Dairy Cattle Fed a Ration to Induce Sub-acute Ruminal Acidosis (SARA) J. Keane , L. McLaughlin , P. Thompson , G. Greenwood , I. Bruce 1 Pfizer Animal Health, Veterinary Medicine Research and Development, Sandwich, United Kingdom 2 Pfizer Animal Health, Veterinary Medicine Research and Development, Kalamazoo, Michigan, United States

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SARA in dairy cattle is associated with decreased productivity, including decreased milk yield and milk fat. Metabolism of high carbohydrate rations increases the concentration of ruminal VFAs and, in severe cases, ruminal lactate resulting in ruminal pH below 5.5 (SARA) or 4.5 (acute acidosis). Acarbose is a pseudotetrasaccharide that inhibits -amylase and glucosidase enzymes thus reducing the rate of carbohydrate digestion and has been demonstrated to maintain higher pH values by decreasing total VFAs and % propionate in an acute model of acidosis. To evaluate the efficacy of acarbose in managing low milk fat associated with SARA thirty early lactation Holstein cattle (average 101 days in milk) were fed a ration consisting of ground corn, soybean meal and corn silage (70:30 concentrate: fiber). The ration was shown to result in pH < 5.5 for at least 4 hr/day for 80% of the days tested. Cattle were trained to feed from Calan gates and were assigned to control or 0.75g/animal/day acarbose added to the total mixed ration (TMR) according to a cross-over design. Animals were fed the assigned test ration for 13 days and daily feed intake, milk yield and composition were measured for the subsequent 7 days of each 20-day cross-over period. Percent and yield of milk fat were increased in the acarbosetreated groups (3.33% vs. 3.04%, P<0.01 and 1158g/day vs. 1035g/day, P=0.01, respectively). While milk yield was not affected, 3.5% fat-corrected milk (FCM) yield was increased in animals receiving acarbose (33.7 kg/d vs. 31.7 kg/d, P<0.01). In addition, dry matter feed intake was increased in the animals receiving acarbose (22.2 kg/d vs. 20.7 kg/d, P<0.01). In a second experiment using the same design and ration, thirty animals approximately 80 days in milk were encouraged to consume a large meal by being fasted for 3 hrs prior to the daily feeding. Under these conditions, percent and yield of milk fat were increased (3.16% vs. 2.67%, P<0.01 and 1229g/d vs. 1047g/d, P<0.01, respectively) in animals receiving acarbose. Although milk yield was not affected, 3.5% FCM yield was increased in animals receiving acarbose (36.9kg/d vs. 33.9kg/d, P<0.01). In addition, dry matter feed intake was increased (22.5kg/d vs. 21.8kg/d, P<0.04). It was concluded that acarbose corrected the decreased milk fat associated with SARA-inducing rations and increased 3.5% FCM. 492 Could Phytotherapy Heal Microhematuria Caused by Chronical Ingestion of Pteridium Aquilinum? R. Machado de Oliveira, F. Kloster, E. Bueno, C. Molli, A. Possamai, S. Cirio, M. Mangrich-Rocha, R. Ollhoff Pontifícia Universidade Católica do Paraná, Ciencia Animal, Sao José dos Pinhais, Brazil Objectives: Bovine enzootic hematuria (BEH) is still a common problem in Brazil mainly for small farmers in regions not suited for animal production. BEH has no available treatment only being recommended the early slaughter of affected cattle and the eradication of bracken fern. The present study aimed to observe the therapeutic effect over the symptom microhematuria of BEH using the aqueous extract of sawdust of Tabebuia avellanedae, a native Brazilian tree. Material and methods: 15 Nelore heifers with microhematuria evaluated by Combur-Test® stripes with microhematuria (1 or 2 +) were randomly assigned to two groups. Group A (n=10) treated and group B (n=5) untreated control. They came from a 173 beef cattle herd of the municipality of Rio Branco do Sul- PR® Brazil where 53,2% of them had different degrees of hematuria and the farm is severely infested with bracken fern (Pteridium aquilinum). The animals were kept in a stockyard of the Ruminant Hospital of PUCPR and treated SID subcutaneously with 100ml of an aqueous extract of Tabebuia avellanedae for 30 days. Previous to the experiment the animals were kept at the hospital for one month for adaptation and received all the same balanced feed based on maize silage. Microhematuria was tested immediately before and at the end of the experiment. Blood parameters

as urea, creatinine, ,,GT and transaminases were evaluated as was the local injection site. Results and discussion: Group A initiated with a mean of microhematuria of 1,1-0,1 against 1,6-0,6 of group B without difference. After the end of the experiment the values were respectively 0,6-0,3 against 2,0-0,9 with a tendency (p = 0,08) for the treatment group. Six heifers of group A had no microhematuria after the 30 days period, but in the control group also two animals showed no signs of microhematuria, whereas two other animals worsened. The hepatic and the renal function of both groups were not altered. Five heifers developed abscesses at the injection points, due to probably secondary contamination. As the extract is rich of lapachol an naphtoquinone with antitumoral activity, we suppose that there could be a direct effect over tumoral vascularization in BEH. Conclusion: The phytoterapic extracted from Tabebuia avellanedae had a tendency of healing animals with microhematuria due to ingestion of bracken fern. Further studies must avaliate if BEH at his initial stage could be treated. Key words: phytotherappy, Pteridium aquilinum, lapachol; hematuria 493 Clinico-therapeutic Studies on Post-parturient Hemoglobinuria in Buffaloes S. Sridhar Harayana. Agriculture University, India, Veterinary Medicine, Hisar, India Study was conducted on 15 clinical cases of post parturient hemoglobinuria in buffaloes from and around Hisar district of Haryana (India). For comparison same number of healthy buffaloes from similar habitat and locations were kept as control. These clinical cases had recently calved and were in their third to sixth lactation. They exhibited clinical signs like red or coffee coloured urine, anorexia, anemia and laboured breathing. A typical feature of clinical syndrome was straining during defecation. Hematological examination in these cases revealed lower levels of hemoglobin, packed cell volume and total erythrocyte count. Hypophosphatemia was a consistent finding in serum biochemical analyses of these cases which also showed increased levels of metabolite malondialdehyde and enzymes super oxide dismutase, catalase and glutathione peroxidase. All but two of these cases recovered fully when treated with antioxidant ascorbic acid @ 7.5 g dissolved in 5% dextrose 500 ml intravenously along with supportive therapy with mineral mixture @ 100g orally and vitamin B complex injection once a day till recovery. On an average 3 days of therapy was needed for complete recovery. Unrecovered animals died probably due to their advanced disease state. Iron containing hematinics were prescribed only after the hemolysis stopped since iron is reported to increase oxidative stress further. It is hereby concluded that oxidative stress on erythrocytes is probably the precipitating cause of intravascular hemolysis seen in this disease. Key words: hemoglobinuria, oxidative stress, buffaloes 494 The Effect of Cu, Se, Mn and Zn Supplementation on Tissue and Organ Concentration of Copper in Goats O. Langová, A. Pechová, L. Mi_urová, L. Pavlata, R. Dvorák University of Veterinary and Pharmaceutical Sciencies Brno, Clinic of Ruminants, Brno, Czech Republic The aim of the experiment was to find out whether there are any interactions during peroral supplementation with several trace elements in their inorganic form. We monitored interaction between copper (Cu) and zinc (Zn), manganese (Mn) or selenium (Se) in adult white shorthaired goats. Twenty-two goats were enrolled in the trial and divided to four experimental groups. All groups were supplemented with 3 mg of Cu per animal and day in the form of CuSO4. Group A (n=6) was supplemented only with Cu; animals of group B (n=6) were given Cu together with Zn (72 mg per animal and day) in the form of ZnSO4; group C (n=5) was supplemented with Cu together with Mn (54 mg per animal and day) in the form of MnSO4; and in group D (n=5) we administered Cu together with Se (0.54 mg per animal and day) in the form of Na2SeO4. The basic feed ration was identical for all groups and consisted of hay (ad libitum) and wheat flour (300 g) supplemented with respective trace elements. Animals were supplemented with different trace elements for three months. Finally, the goats were slaughtered and samples of individual tissues were taken. These were mineralized and analyzed for Cu concentration by the method of atomic absorption spectrophotometry. No significant Cu concentration differences between groups in any tissues or

28 XXV. Jubilee World Buiatrics Congress 2008

organs were identified. However, the results varied a lot within individual groups. Mean values of Cu concentration in individual groups ranged as follows (tissues are ordered in a descending line): liver (9.77-12.95 mg/kg of fresh tissue), heart (3.90-4.41 mg/kg), kidney (2.49-2.92 mg/kg), lungs (2.45-2.66 mg/kg), tongue (1.81-1.94 mg/kg), diaphragm (1.39-1.61 mg/kg), pancreas (1.25-1.56 mg/kg), spleen (1.24-1.52 mg/kg), thigh musculature (0.85-1.03 mg/kg) and neck musculature (0.77-0.87 mg/kg). On the basis of our results Cu supplementation with concurrent administration of Zn, Mn and Se in higher doses does not cause different concentrations of Cu in individual tissues and organs of the goats. The work was realized with the support of MSM Project No. 6215712403. Key words: microelements, liver, kidney, heart, lungs 495 Evaluation of Copper Status in Dairy Cows Living in Region Irrigated with Potato Utility Refuses M. Kleczkowski, W. Kluci_ski, T. Jakubowski, K. Dembele Warsaw University of Life Sciences, Faculty of Veterinary Medicine, Clinical Sciences, Warsaw, Poland Copper status (CS) can be determined by 5 blood parameters: total plasma copper (Cu) zinc (Zn), molybdenum (Mo) as well as ceruloplasmin (Cp), and superoxide dismutase (SOD) activities. The aim of experiment was to evaluate CS from the region irrigated with potato utility refuses. The experiment lasted for 6 months, and it comprised 45 cows of the Black-White breed, at the 3 - 6 years. Cows were chosen from the 3 districts in the valley of Narew River, each group (1, 2 and 3) with a different irrigation method. Cows, from 1 and 2- experimental groups, inhabited areas irrigated with utility refuses from potato industry company. Area inhabited with cows from 1 group was irrigated with utility refuses by using a flow method. Area inhabited with cows from 2 group was irrigated with utility refuses by using a sprinkle method. Cows from 3 group were control. Mineral analysis of utility refuses had been made before experiment started. Blood samples from all cows were collected 3 times: 1- one week before calving, two-three days after calving, and 3- six weeks after calving. Cow blood parameters that were determined are: Cu, Zn, Mo concentration, and Cp, SOD activity. Average value of particular 3 parameters for cows from group 1 was: Cu- 8.99 µmol/ dm , Mo -0.24 3 3 µmol/ dm , Zn- 11.46 µmol/ dm , SOD- 70.11 U/gHb, Cp- 1.37 µmol/ 3 3 dm , for cows from group 2 was: Cu- 7.23 µmol/ dm , Mo- 0.21 µmol/ 3 3 3 dm , Zn- 9.82 µmol/ dm SOD- 54.22 U/gHb, Cp- 0.89 µmol/ dm , for 3 cows from control group 3 was: Cu- 12.07 µmol/ dm , Mo- 0.26 µmol/ 3 3 3 dm , Zn- 14.55 µmol/ dm , SOD- 81.36 U/gHb, Cp- 1.66 µmol/ dm . The analysis of results showed that nutrition of dairy cows produce a decrease of copper status and Cu-dependent antioxidant activity in blood of cows. Key words: cows, copper, antioxidants, utility refuses 496 Diurnal Variation of Acid Base Status in Cows Fed Different Anionic Salts Once a Day Only C. Gelfert , A. Löptien , N. Montag , R. Staufenbiel 1 University of Veterianary Medicine, Clinic for ruminants, Vienna, Austria 2 Faculty of Veterinary Medicine, Freie Universitaet Berlin, Clinic for Ruminants and Pigs, Berlin, Germany Eleven non-lactating, non-pregnant, Holstein-Friesian-Crossbreed cows received 2,000 meq of either calcium chloride (CaCl2) or calcium sulphate (CaSO4) once a day for a period of one week. At day seven, blood and urine samples were taken every four hours starting at 6 o'clock before feeding and ending the same time the next day. Before feeding, urine analysis of the CaCl2-cows showed a compensated acidosis due to an reduced pH and low levels of net acid-base excretion (NABE). Feeding AS changed acid-base status (ABS) noticeably in both groups over time. All cows developed a strong metabolic acidosis. The strongest acid load was visible 18 hours after the morning feeding. Changes were greater in CaCl2-cows and NABE differed significantly (p=0.037) between both anionic salt groups. Nadir of NABE was 41mmol7l in CaCl2-cows and +4mmol/l in the CaSO4-cows. The fractional clearance of calcium was higher (p=0.027) in the CaCl2cows due to a higher calcium excretion via the kidneys. Serum calcium concentration did not show significant differences between both groups (p>0.05) but changed significantly during the day (p<0.001). During the night, the acidosis became weaker and initial values were reached

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the next morning. Contrary to the CaSO4-cows, CaCl2-cows still showed signs of a compensated metabolic acidosis in blood and urine. The results of the study showed that CaCl2 had a stronger impact than CaSO4 and that feeding AS once a day only encloses the risk of an interrupted effect of the AS during the night. 497 Influence of a Starter with Rape Cake on the State of Health in Calves R. Dvorak, L. Pavlata, A. Pechova, J. Filipek, E. Gopfert University of Veterinary and Pharmaceutical Sciences, Faculty of Veterinary Medicine, Clinic of Ruminants, Brno, Czech Republic After rape oil removal valuable remains are left and may be used for feeding purposes. Full utilisation of products made of rape is interfered by the presence of antinutritional substances, i.e. glucosinolates (GSL). The total content of glucosinolates has been decreased to 25 µmol/g in cultivated varieties, the so-called double zero "00" Canola ones, or even to 10 -15 µmol/g in the best ones. This study was aimed at examining a new starter Grant (G) with 10% content of rape cake processed using our proposed technology (GLS, max. 3 µmol/g). A total of 45 calves immediately after delivery weighing 44.5 kg were included into the experiment. Three equal groups of 15 calves were formed. Control group S was fed only by the Standard starter containing common components. The other Telstar (T) starter is commonly used in calf rearing in the Czech Republic. The experimental starter G contained 10% of treated rape cake. The experiment lasted 90 days. State of health was checked using clinical examination and selected biochemical parameters in blood and ruminal fluid. The overall consumption of starters in groups S, T and G amounted to 120.6, 154.8 and 130.5 kg, respectively. Selected biochemical parameters ranged within reference values for the whole experiment. There were not significant differences in iodine levels of groups S and T (208.90±71.40; 152.10±43.00 µg/l) when compared with group G (184.80±34.80 µg/l). Likewise, hormone levels of T3 and T4 of groups S and T(4.02±0.98; 3.51±1.00; 77.80±26.30; 71.50±30.00) were not different from group G (3.75±1.00; 63.00±24.30 nmol/l). Mean weights of thyroid glands of 6 calves from each group S,T and G sacrified at the end of experiment were 16.5±2.8, 11.7±2.2 and 15.5±4.8 g, respectively. Histology revealed no changes between individual groups in the structure, size of follicles and colloid content. Feeding the starter containing 10 % of rape cake low in GSL content had not any negative effect on growth and development of calves. The experimental calves showed no significant changes in parameters of ruminal fermentation, nitrogen, energy, lipid and mineral metabolism, liver function as well as thyroid gland function. Supported by Ministry of Education, Youth and Sports of the Czech Republic (Project MSM6215712403). Key words: calf, nutrition, rape cake 498 The Effect of Vitamin E Supplementation on Oxidative Status of Dairy Cows R. Bouwstra, P. Dobbelaar, M. Nielen, T. Werven Faculty of Veterinary Medicine, Utrecht University, Animal Health Department, Utrecht, The Netherlands Objective: The effect of oral vitamin E supplementation during the dry period on oxidative status biomarkers in serum was evaluated in an observational field study. Materials and Methods: Ninety-eight cows were selected from 26 herds in The Netherlands. Fifty-two received a daily vitamin E supplement of 1.000 international units; forty-six were not supplemented. A blood sample was taken between 0 and 28 days before calving. Serum was analyzed for ·-tocopherol, cholesterol, triglycerides, superoxide dismutase (SOD), glutathione peroxidise (GSHpx), uric acid, albumin, protein sulfhydryl oxidation level, reactive oxygen metabolites (ROM), Fe, the ferric reducing ability (FRAP) and malondialdehyde (MDA). Results: Alfa-tocopherol was significantly higher in the supplemented group, as was the ratio ·-tocopherol: cholesterol. MDA was significantly lower in the supplemented group, suggesting lower levels of lipidperoxidation. Results of the mixed model also showed that supplementation lowered MDA serum levels. Conclusion: In conclusion, vitamin E supplementation increases · tocopherol and lowers MDA serum levels under field conditions. Key words: vitamin E, field study, oxidative status

Nutrition and Metabolic Disorders 29

499 Secondary Hepatogenous Photosensitization in Buffalos due to the Excessive Accumulation of Hepatic Copper A. Minervino , R. Barreto júnior , F. Rodrigues , L. Reis , 1 3 1 R. Ferreira , S. Headley , E. Ortolani 1 College of Veterinary Medicine, University of Sao Paulo, Department of Clinical Sciences, Sao Paulo, Brazil 2 Rural Federal University of Semi-Arid, Mossoró, Brazil 3 Faculty of Veterinary Medicine, University of Helsinki, Helsinki, Finland Four 1.5 yr-old, male, murrah buffaloes, maintained during six months with high Cu intake and without direct solar exposure, were transported to regular farming conditions. Shortly after sunlight exposure, all buffalos demonstrated clinical manifestations characteristic of photosensitization: anorexia, apathy and severe cutaneous lesions. Blood samples were obtained before solar exposure (T0), during the clinical manifestations (T1), and after recovery (T2) to evaluate haematological (PCV, Hb, RBC and WBC) and biochemical (total protein, albumin, g-glutamil transferase (gGT), aspartate amino transferase (AST), creatine kinase (CK), urea, and creatinine) parameters. Skin samples obtained by incision biopsy at T0 and T1 were processed for routine histopathology. Treatment consisted of removal from sunlight exposure and aqueous Zinc oxide solution. Three animals recovered fullly and 1 died. The hepatic Cu concentrations and gGT and AST activities before solar exposure were above normal values. For haematological parameters, only RBC demonstrated significant difference, being reduced at T1 and returning to normality at T2. A similar pattern was observed with the total protein and albumin; but an opposite trend occurred with CK activity. The buffalo that succumbed had the highest liver of Cu concentration and more extensive macroscopic skin lesions. At T0 the buffalos had generalized alopecia, but insignificant histological alterations (acantholysis and neovascularization). However, at T1 histopathological examination revealed hepatic photosensitization characterized by diffused, moderate parakeratotic hyperkeratosis, multifocal moderate acantholysis, degeneration and necrosis of squamous epithelial cells, sweat gland atrophy, sebaceous gland hyperplasia, and dermal edema. Although the buffalos demonstrated a liver insufficiency at T0, the skin lesions indicative of secondary hepatogenous photosensitization were only triggered after the solar exposure. In this case, a high Cu intake induced hepatic lesions that may have caused inefficient metabolization of phylloerythrin resulting in secondary photosensitization after the sunlight exposure. Research supported by FAPESP Key words: buffaloes, hyperkeratosis, photosensitization, hepatic lesions 500 Trace Elements in Cattle: Copper and it`s "Friends" A. Mueller, B. Freude, M. Scholz, M. Weiss IDEXX, Vet Med Labor GmbH, Ludwigsburg, Germany Prophylactic procedures to prevent diseases become more and more important in cattle rearing. Adequate supply in trace elements is required. Copper (Cu) deficiency is regarded as the second most common mineral deficiency of cattle in the World (Telfer et al, 1996; Black und French, 2004). Two different copper deficiencies are known: a primary copper deficiency induced by a simple deficiency of copper in the diet, and a secondary, resulting from the reduction in copper absorption or utilization by the antagonistic effects of molybdenum (Mo) and iron (Fe). 340 cattle samples were analysed as serum/plasma pairs regarding the concentration of Cu, Mo and Fe with ICP-AES (Inductively Coupled Plasma Atomic Emission Spectroscopy) during July 2006 until March 2007. The Cu-plasma analyses showed that only 24% of these samples were below the reference value for Cu (80 Ïg/dL). Compared to these results, 84% of the corresponding serum samples showed a Cu deficiency (below 80 µg/dL). Serum clotting reduces Cu by up to 80% compared to the plasma values (in confirmation to Laven & Livesey, 2006). In contrast to copper the analyses of Mo in bovine plasma and serum resulted in comparable values. 4500 bovine serum and plasma samples were analysed for Mo. 80% of Mo-concentrations were below 26 µg/L. Only a few samples (< 1%) showed higher values (max. 3300 µg/L). A correlation between low Cu and high Mo values in plasma, or low Cu values with high Fe concentrations could not be found. Additionally, hair analyses of Cu (N = 173) and Mo (N = 99) were performed. 85 % of the hair samples (31 2 1 1

times analyses of each hair sample) showed Cu values between 5 and 20 mg/kg (49%: 6,6-10,4 mg/kg; 4 %: below 6,6 mg/kg), 15 % of the samples showed higher values up to 100 mg/kg). The Mo concentrations in the hair samples varied between 0 up to 800 µg/kg or even higher). The comparison between Cu and Mo in hair (N = 99), or Cu hair/plasma (N = 107) and Mo hair/plasma (N = 70) showed no correlation. The analysed cattle samples with low copper plasma concentrations showed mainly a primary copper deficiency. A secondary copper deficiency caused by a molybdenum intoxication could not be found in this set of samples. 501 Effect of chronic glucose infusion on lactation performance and metabolic profiles in dairy cows B. Al-Trad , K. Reisberg , T. Wittek , A. Alkaassem , G. Gäbel , 2 1 M. Fürll , J.R. Aschenbach 1 Faculty of veterinary medicine, University of Leipzig, Institute of Animal Physiology, Leipzig, Germany 2 Faculty of veterinary medicine, University of Leipzig, Medizinische Tierklinik, Leipzig, Germany Objective: To characterize the metabolic adaptations that occur during gradual increases of glucose supply via prolonged i.v. glucose infusions in dairy cows. Material & methods: Twelve midlactation dairy cows were assigned randomly to continuous i.v. infusions of either saline (control group, n=6) or 40% glucose solutions (treatment group, n=6) for 28 d. The infusion dose started at 1.25% of the daily energy (NEL) requirement and then gradually increased until a maximum dose of 30% NEL requirement was achieved at d 23. Then infusion dose was maintained at 30% for 5 d. The treatment (feed plus infusion) was hypercaloric but isonitrogenous. Liver and skeletal muscle biopsies were taken on d 0, 8, 16, 24, and 32. Blood samples were taken every 2 d at 10:00 a.m. and additional 24-h blood samples (6-h intervals) were taken before each biopsy. Milk and urine samples were also taken on the biopsies days. Results: No changes occurred in daily feed intake and milk production, milk lactose and urea concentrations. Milk protein percentage and yield tended to increase during the high infusion dose (i.e. 20 & 30% NEL requirement). Decrease in milk fat percentage and yields were observed in both groups; however, the decrease was numerically higher in the treatment cows. Body weight and BFT increased in the treated group. Small and occasional increases in blood glucose and insulin concentrations were observed in the blood samples taken at 10:00 a.m. every 2 d. However, at infusion of 30% NEL requirement, five out of six treated cows had blood glucose concentrations >8 mM at 16:00 p.m., i.e. 1 h postprandially. Glucose infusion decreased serum NEFA, BHBA and BUN concentrations. Serum liver enzymes, cholesterol, bilirubin, K and Ca concentrations were not affected by infusion. Less than 2% changes were observed in serum P, Cl and Na concentrations. Glucoseuria was detected during the maximum infusion dose. Liver glycogen increased gradually to reach plateau on d 16 before it fell back to baseline values on d 32. Skeletal muscle glycogen increased and liver total lipid tended to increase during the high dosage of glucose infusion. Conclusions: Dairy cows on an energy-balanced diet do not direct excess glucose to increased lactation performance. Excess glucose is transiently stored as glycogen in the liver and, predominantly, transferred to body fat. At high dosages, glucose is also stored in skeletal muscle as glycogen and excreted via the urine. 502 Biochemical Profile of Cattle with Induced Hypocalcaemia and Subsequently Treated with an Enriched Calcium Solution R. Barreto júnior , A. Minervino , F. Rodrigues , E. Meira júnior , 2 3 2 2 R. Ferreira , A. Lima , C. Mori , E. Ortolani 1 Rural Federal University of Semi-Arid, Mossoró, Brazil 2 College of Veterinary Medicine, University of Sao Paulo, Department of Clinical Science, Sao Paulo, Brazil 3 Vallée S.A., Sao Paulo, Brazil Twelve two years-old Holstein heifers were used to study the biochemical profile during the induction and recovery of hypocalcaemia. The picture was induced by continuous infusion of a EDTA solution (5% and pH 7.4) into the jugular vein at the speed of 220 mL/h until the animals presented definitive clinical signs of hypocalcaemia such as sternal or lateral recumbency; then the infusion

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30 XXV. Jubilee World Buiatrics Congress 2008

was immediately stopped and the heifers were treated, during 30 min, with 1 mL/kg BW of a calcium solution (ValléeCálcio®) with 2.44 g of calcium /100 mL, from three sources: Gluconate, Lactate and Sacarate, enriched with 5 g Glucose, 0.47 g phosphorus and 0.19 g magnesium per 100 mL. Venous blood samples were taken before infusion (T0), when the muscle tremors started (T1), at the end of the infusion (T2), at the end of the treatment (T3) and 24 h later (T4) for serum analysis of calcium, ionized calcium, phosphorus and magnesium, plasma glucose levels and blood activity (30 °C) of g-glutamyl transferase (gGT), aspartate aminotransferase (AST) and creatine kinase (CK). The induction generated a similar clinical picture as seen in the natural course of hypocalcemia and the treatment with the calcium solution promoted total recovery. The induction generated a significant decrease in calcium, ionized calcium, phosphorus and magnesium levels, principally at T2, but the treatment transiently increased the calcium and ionized calcium at T3 and all of the variables returned to normality 24 h later. The plasma glucose levels increased only after the treatment, but returned to normal at T4. There was no change in the gGT activity throughout the trial, but a significant increase of AST and CK at T4, indicating that some muscle damage occurred during the process. Key words: hypocalcaemia, calcium, phosphorus, treatment 503 Clinical Picture of Induced Hypocalcaemia in Holstein Heifers Subsequently Treated with an Enriched Calcium Solution R. Barreto júnior , A. Minervino , F. Rodrigues , E. Meira júnior , 2 3 2 2 R. Ferreira , A. Lima , C. Mori , E. Ortolani 1 Rural Federal University of Semi-Arid, Mossoró, Brazil 2 College of Veterinary Medicine, University of Sao Paulo, Department of Clinical Sciences, Sao Paulo, Brazil 3 Vallée S.A., Sao Paulo, Brazil Twelve two years-old Holstein heifers were used to study the clinical picture during the induction and recovery of hypocalcaemia. The picture was induced by continuous infusion of a EDTA solution (5% and pH 7.4) into the jugular vein at the speed of 220 mL/h until the animals presented definitive clinical signs of hypocalcaemia, such as sternal or lateral recumbency; then the infusion was immediately stopped and the heifers were treated, during 30 min, with 1 mL/kg BW of a calcium solution (ValléeCálcio®) with 2.44 g Ca (gluconate, lactate, and saharate), 5 g Glucose, 0.47g phosphorus and 0.19 g magnesium per 100 mL. A clinical examination was performed before infusion (T0), when the muscle tremors started (T1), at the end of the infusion (T2), at the end of the treatment (T3) and 24 h later (T4). The induction generated a similar clinical picture as seen in the natural course of hypocalcemia, but a constant and involuntary kinetic tongue movement cleaning the nostril and nose and causing a lip sound. Limb stiffness and uneasiness preceded the muscle tremor, and from this moment on the animals refused food. Then apathy, staggering and mydriasis became evident. These symptoms were followed by a rapid sternal and in most cases lateral recumbency. Even in this last position the heifers kept the head turned to the flank. At this moment the skin temperature at the lower limb and anal reflex were decreased; anuria and somnolence were evident. The heart rate and hypophonesis increased as higher was the EDTA infusion. Conversely, rumen stasis occurred in most cases at T1. Most animals had gas bloat. The treatment with calcium rapidly reversed the clinical picture, restored the heart rate and the rumen tonus and did not cause any adverse reaction within 48 hours after treatment. Key words: hypocalvaemia, symptoms, treatment, recovery 504 Evaluation of Faecal Quality as a Diagnostic Tool in SARA Diagnosis in Dairy Cattle J. Tajik1, Mg. Nadalian1, A. Raoofi1, Gr. Mohamadi2, A. Bahonar1 1 University of Tehran, Clinical Sciences, Internal Medicine, Tehran, Iran 2 University of Tehran, Clinical Sciences, Tehran, Iran Objectives: Sub acute ruminal acidosis (SARA) is characterized by daily episodes of low ruminal pH between 5.5 and 5.0. This digestive disorder is the consequence of feeding high grain diets to dairy cows, which are adapted to digest predominantly forage diets. SARA may be a common and economically important problem in dairy herds. There is Limited knowledge of the symptomatology of SARA. Changes in faecal consistency and structure of SARA affected cows have been

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Scores 1 Parameter Faecal Very consistency dry, (FCS) lumpy

2

3

4

5

Circular, moist raised pat, symmetrical rings surrounding a dipped centre Faecal No few particles undigested particles particles clearly feed particles visible (FFP) Faecal gas No few Massive bubbles bubbles bubbles presence (FBS) of bubbles

Dry, stiff, semi formed pats

Flat, Liquid loose, pools of thinly faeces spread

Massive presence of particles

described. Considering the fact that SARA has to be understood as a transient condition with no clinical consequences visible at the time of insult, it seems that faecal changes rather apply to more severe states of ruminal acidosis than to SARA. This study was designed to evaluate the validation of faecal quality as a diagnostic tool in SARA diagnosis in dairy cattle. Materials and Methods: Ruminal fluid by means of rumenocentesis (Nordlund& Oetzel 1995) was collected from 117 cows (37 early lactation and 80 mid lactation cows) from 10 dairies, 2-4 hours following morning total mixed ration feeding. Additionally scoring of faecal quality (consistency, presence of undigested feed particles and gas bubbles) was done (table1). chi square and fisher exact tests were used in statistical analysis of data results. Results: There were no significant differences between individuals experiencing SARA (ruminal pH< 5.5) and the rest of population (ruminal pH>5.8) in FCS (P=0.908), FBS (P=0.577) and FFP (P=0.288). Also, no significant difference was found between animals with SARA and a marginal pH (5.6<pH <5.8) and the rest of the study population in comparison of FCS (P=0.985), FBS (P=0.375) and FFP (P=0.456). Any significant differences was found between early lactation and mid lactation cows in FCS (P=0.97), FBS (P=0.91) and FFP (P=0.11). Conclusions: According to results of this study faecal quality changes are not significant signs for SARA detection in dairy cattle. Key words: SARA, dairiy cattle, faecal quality 505 Occurrence of Primary Pregnancy Toxemia of Ewes due to a Misdiagnosis of Pregnancy via Ultrasonography N. Alidadi, S. Rafia, S. Moadab, M. Eslampour Faculty of Veterinary Medicine, University of Tehran, Clinical Sciences, Tehran, Iran In a seven day period, four ewes were referred to the Veterinary Research and Teaching Hospital of University of Tehran, with the chief complaints being the recumbency and profound depression. They came from a flock of 80 sheep with "Shaal" breed which had been naturally bred 4.5 months before. The pregnant ewes had been separated from non-pregnant in about 2 months after the breeding by using the transabdominal ultrasonography. The referred ewes were all from the "non-pregnant diagnosed" group and had been put on a low energy diet of mainly straws for the maintenance. They were all multiparous with low body condition scores, the mental state was depressed and the ewes were sternally recumbent at the time of examination, which became lateral terminally. They were oblivious to their surroundings and had muscular twitches over the thorax and flank. Champing of the jaws were present with one ewe showing a persistent involuntary movements of the mandible. The animals were not blind although the Menace Response was significantly slow. The rectal temperatures were normal to hypothermic especially in terminal stages. Tachycardia and polypnea were constant findings in all four ewes. Mucous membranes were pale and it was cyanotic in one. While examining the abdominal region, surprisingly, the pregnancy with twin lambs was detected. Blood and urine samples were submitted to laboratory for further investigations. The results showed hypoglycemia in two cases while the other two being euglycemic. Blood calcium levels were normal in all of them. Ketonuria was a persistent finding in all of the ewes. The haemogram showed a stress Leukon in 3 ewes. None of the ewes could survive and

Nutrition and Metabolic Disorders 31

they all died in a two to three day course. Of the necropsy findings, the severe fatty change of the liver seemed to be the most important. The diagnosis of "Primary pregnancy toxemia of Sheep" was made upon the gathered data. With this, the remaining ewes in the "non-pregnant diagnosed" group were rechecked for the presence of pregnancy and those found pregnant, were moved to a higher energy diet with nutritional supports and eventually further cases and losses were prevented. From these, it has been concluded that a small error in early pregnancy diagnosis via Ultrasonography can lead to significant losses in sheep flocks. So it is important to know, along with modernizing the techniques in animal husbandry, it is of great importance to implement the methods meticulously to prevent such problems. Key words: sheep, pregnancy toxemia, metabolic disorders 506 The Use of Urinary pH Measured in Pooled Urine Samples for Monitoring the Use of Anionic Salts in Dairy Cow Herds C. Gelfert , S. Römer , N. Brand , M. Goebbels , R. Staufenbiel 1 University of Veterianary Medicine, Clinic for ruminants, Vienna, Austria 2 Faculty of Veterinary Medicine, Freie Universitaet Berlin, Clinic for Ruminants and Pigs, Berlin, Germany The use of pooled urine samples of dairy cows was evaluated in the monitoring of the use of anionic salts (AS). In a first step, it was evaluated, whether the pH of pooled samples reflect sufficiently the calculated mean of the pH measured in the single samples, which form the pooled one. Five large dairy farms in Germany were visited monthl, and 10 cows each from four different stages of lactation were randomly selected. From each cow, a urine sample was taken by using a urinal catheter and a pooled sample was produced for each stage of lactation by merging equal amounts of each single sample. The urinal pH was measured in the single samples and the pooled samples (xp) with a portal pH meter. After measuring the pH in the single samples, the mean pH of each lactational group (xs) was calculated. The evaluation of xp followed the procedure of Bland and Altmann (1986). The mean of xs was 8.22 and the mean of xp was 8.29. The urinary pH of Xp was slightly higher than xs. The mean difference was 0.07. These small increases of pH only bore a negligible risk of false diagnosis of urinary acid-base status. In the second step, the urinary pH of pooled samples was tested in reflecting the use of AS in dairy cow herds. 26 dairy farms were visited on which AS were used. Urine samples from 10 randomly selected healthy transition cows were taken, and a feed sample from the total mix ration including the anionic salts was taken to measure the concentrations of the following minerals: sodium, potassium, calcium, chloride, and sulfate, and DCAD was calculated. Although all farms used AS, a high variation of urinary pH was visible, ranging from 5.90 to 8.64. DCAD also showed a high variation ranging from -393 mEq/kg DM to 303 mEq/kg DM. A DCAD below zero was no guarantee for a decrease of urinary pH. Therefore, the farms were grouped according the diagnostic result of urinary pH (cut off point 7.8). DCAD tends (p=0.070) to result in lower values when urinary pH decreased (-1.96mEq&kg DM/114mEq/kg DM). Pooling samples of cows of the same stage of lactation minimizes laboratory work and resulting costs and the measured value corresponds sufficiently to the calculated mean of the single samples. The use of AS in the prevention of periparturient hypocalcaemia has to be monitored by urine analysis. Measuring DCAD may lead to misinterpretation when other faults in feed management exist. 507 Effect of Anionic Salts Supplement in Last Two Months Pregnant on Metabolic Disorders and Blood Minerals of Holstein Cow M. Ganjkhanlou, A. Nikkah, A. Zali, M. Motlagh Animal Sciences Group, animal science, Karaj, Iran In order to, investigate the effect of feeding cow with different of dietary cation anion balance (DCAB) in far-off and close-up period, on metabolic disorders and blood composition of Holstein cow in pregnancy and fresh period this experiment was carried out. In this study 18 Holstein pregnant cows (220-225 d) were fed individually, a base diet with tree different DCAB (+13(control), 0,-13 mEq [(Na+K)(Cl+S)]/100gr dry matter) for 60±5 days. For decrease DCAB two anion salts such as ammonium chloride and ammonium sulphate were used. The result indicated milk fever percentage were 0, 33, 33 for diet -13, 0,+13 mEq/100gr DM, respectively. The rate of retained placenta was 9.37, 4.62, 4.61 in cows received diet +13,0,-13 mEq/100gr DM

1 2 2 2 2

respectively. Blood pH, Na, K, Mg and Ca were unaffected by dietary cation-anion balance. Urinary PH of cows was affected by diets. Urinary pH increased linearly with increasing DCAB. The mean urinary PH was 7.99 (for diet +13), 6.81 (for diet 0) and 6.11 (for diet -13). Key words: DCAB, ammonium chloride, ammonium sulphate, metabolic disorders 508 The Effect of Sodium Selenite and Sel-Plex on Selenium Deficiency Prevention in Beef Cattle J. Illek , M. Matejicek , D. Kumprechtova , M. Hulik , T. Berka 1 Veterinary and Pharmaceutical University Brno, Clinic of Ruminant Diseases, Brno, Czech Republic 2 VVS, Vermerovice, Czech Republic 3 Alltech, Alltech CZ, Praha, Czech Republic Objective: The aim was to investigate effects of dietary inclusion of different selenium (Se) forms and AA cows in the grazing season. Material and methods: 3 Se deficient herds, grazed at Se deficient pastures (bellow 0.1 mg Se/mg DM). Herd A got a complete mineral lick with sodium selenite (30 mg Se/kg). Herd B got mineral feed with sodium selenite (30 mg Se/kg). Herd C got mineral feed with organic Se (Sel-Plex) (30 mg Se/kg). Grazing season was April - November. There were groups by 40 cows, along with calves and a bull. They fed ad libitum on pasture forage and the mineral lick or mineral feeds only. In the end of the pasture season, 10 cows from each herd were withdrawn blood samples. Whole blood Se levels and glutathionperoxidase (GPx) activity were measured, metabolic profile parameters analysed in serum. In the end of grazing season, all the cows were pregnancy tested. Results: Se (ug/l) and GPx (ukat/l) in blood Herd A Se 52.0aA 4.5 8.7 GPx 320.8A 97.8 30.5 B Se 61.4bA 3.9 6.3 GPx 494.6B 44.2 8.9 C Se 84.4B 11.8 14.0 GPx 796.0C 71.0 8.0

1 2 1 3 3

Mean S.D. C.V.

a,b,c P < 0.05; A,B,C P < 0.001 Whole blood Se levels and GPx activity in AA cows at end grazing season were markedly different between the herds. Herd A showed the lowest Se levels and GPx activity, i.e. pronounced Se deficiency. Herd B showed higher Se level and GPx activity, but still deficient. Herd C showed significantly higher Se and GPx levels, within the physiological ranges. Differences in body condition between the herds were not observed, but 2 cows died in Herd A. Herds B and C had no deaths. Death causes were not identified. There were some differences in numbers of pregnant cows after natural mating. 75%, 80% and 95% conceived in Herds A, B and C, respectively. Conclusion: Organic Se favourably influenced Se levels and GPx activity in blood and fertility. Mineral licks with sodium selenite seem to be worst source of Se. The mineral licks did not remove Se deficiency, which was reflected in very low Se levels and GPx activity and worse fertility results. The results support the previous findings that the utilization of Se from inorganic sources by cattle is poorer than that from organic sources. The differences between Herds A and B were also due to a lower intake of the lick than tha of the mineral feed (meal). The study was carried out within the research project no. 1G46086 (NAZV). Key words: beef cows, organic Se, Se deficiency, metabolic profile 509 Effects of Various Anti-Stress Additives in Daily Diets on the Microelements of the Heat Stress Cows H. Yuan, De. He, J. Wu, Li. Wen, Li. Yuan, R. Li, M. Zhang Hunan Agricultural University, College of Veterinary Medicine, Changsha, Hunan, China Twenty healthy and lactating cows with similar weight, corporeity, appearance, and productive capability, were selected from the cattle farm of Hunan Agricultural University, in the hot season on August 1-30, 2007. These cows were classified into 4 groups using a method of single-factor comparison in order to evaluate the effects of various anti-stress additives on the microelement in blood, milk, and hair of the heat stress cows. The results showed that the concentration of zinc in blood was significantly

32 XXV. Jubilee World Buiatrics Congress 2008

increased in the cows fed with the diets supplemented with vitamin C at a dosage of 1100-1500 mg/day compared with the control group (P<0.01), and their concentration of chromium and selenium were obviously enhanced when compared with the control group (P<0.05). The concentration of iron and chromium in blood was significantly increased in the cows fed with the diets supplemented with a combination of antiheat stress additives (provided from the institute of Baite Bio-technology) at a dosage of 500 g/day compared with the control group (P<0.05 and P<0.01, respectively), whereas their concentration of cobalt was decreased in comparison with the control group (P<0.01). The concentration of iron and selenium in blood was remarkably higher in the cows fed with the diets with a supplement of the Chinese tradition medicine as an anti-heat stress additive (Four Chinese tradition medicines were minced, filtered with 40 µm mesh and combined with a designed ratio, and then the cows were fed at a dosage of 64 g/day) than that in the control group (P<0.05), and their concentration of chromium in hair was 2 times higher than that in the control group (P<0.01). In contrast, the concentration of zinc and chromium in milk were lower in the cows treated with Chinese tradition medicines than those in the control group (P<0.01). The concentration of chromium in hair was increased in the cows with the Chinese tradition medicine treatment, but their selenium, manganese, and cobalt were obviously decreased (P<0.05). Key words: cow, heat stress, anti-heat stress additives, microelement 510 The Comparison of Some Blood Minerals of Najdi Cattle in Hot and Cold Seasons A. Rasooli , Gh. Khadjeh 1 School of Veterinary Medicine,, Clinical sciences, Internal Medicine, Ahvaz, Iran 2 School of Veterinary Medicine,, Clinical Sciences, Ahvaz, Iran One of the native cattle in Khoozestan province of Iran is Najdi breed, that is characterized by a hump in the withers area. There was no information about blood minerals of this breed in Khoozestan province, therefore this study was carried out in order to determination of blood minerals of Najdi cattle in hot and cold seasons with the attention to age and sex. In this study, blood minerals of 180 Najdi cattle in different age and sex groups were determined by conventional laboratory methods. The result of this study showed significant differences between calcium, phosphorus, sodium, potassium and chloride levels in hot and cold seasons. The levels of phosphorus and chloride were higher in cold season and the levels of calcium, sodium and potassium were higher in hot season. The levels of phosphorus, sodium, potassium and chloride have significant differences between different age groups. There was no significant difference between male and female groups. Key words: cattle, Najdi, minerals, blood 511 An Investigation on Subclinical Ketosis in a Dairy Farm of Guangdong Province K. Wang, J. Xu Foshan Science and Technology College, Department of life Science, Foshan, Guangdong, China An investigation on ketone body of a farm in Foshan in Guangdong province was performed, we use the adopt-glucose-oxidase and the concentrations to analysis the blood, milk and urine of 27 holstein cows which derived from k group and I group. The results showed that the blood ketone was 1.155±0.511 mmol/L, the milk ketone was 2.904±1.311 mmol/L, the urine ketone was 3.127±1.814 mmol/L, and two groups cows contain recessive ketone disease (milk ketone and urine ketone greater than 1.722mmol/L). There were 6 cows indicated subclinical ketosis, the coefficient correlation of milk ketone and urine ketone was 0.59038. Key word: cow, blood ketone, milk ketone, urine ketone 512 Cold Pressed and Extruded Rapeseed Oilcake for Dairy Cows Feeding J. Dulbinskis, V. Sterna, D. Ikauniece, I. Zitare Research Institute of Biotechnology and Veterinary Medicine Sigra of the Latvia University of Agriculture, Sigulda, Latvia Rapeseed oilcake is one of richer source of protein and partly fat for cows. From publications known that heating increased nutrition value

1 2

of rapeseed oilcake. Rapeseed oilcake can be produced with cold pressure method and extrusion by heating in different temperatures from 110 °C to 140 °C.Bacteriological and mycological analyses of the rapeseed oilcake samples done by all accepted methods of microbiology, microorganisms differentiated with BBL Crystal GPID.The count somatic cells of milk were detected with Bentley Somacount 500 Combi (USA). The content of fatty acids in rapeseed oilcake was detected with gas chromatograph Varian 3400. Additionally to base feed (maize silage, hay etc.) 2 months to trial groups of cows were fed out 2 kg rapeseed oilcake, heated at 110 °C, heated at 140 °C or sunflower seed oilcake. Rate of microbial contamination of rapeseed oilcake produced with cold pressure method and heated in different temperatures from 110 °C to 140 °C is depended from used temperature. Temperature of extrusion influenced also isolated species. By heating at the 110 °C are killed yeasts, lactic microorganisms significantly decreased amount of moulds. Treatment at the 140 °C caused that microorganisms produced butiric acid were not detected. More often was isolated Mucor sp., Penicillium sp. and Aspergillus sp. moulds. Content of fatty acids after heating in different temperatures was not significantly changed (p>0.05). Content of dry matter in heated rapeseed oilcake is by 2.5-3.5% higher than in unheated and degradable protein increased by 4.68-5.3g per kg. In the groups where in cows diet was included rapeseed oilcake 2kg per day, heated at 110 °C or 140 °C, decreased content of saturated fatty acids and increased content of unsaturated oleinic and miristoleic acid, and amount of produced milk increased at the same time. In these groups decreased fat content of milk (p<0.05). Investigations results showed that feeding of heating rapeseed oilcake decreased somatic cells count by 10-50% in comparison with initial level. Extruded rapeseed oilcake heated at 110 °C and 140 °C can be fed out to dairy cows as additional source of protein. Key words: cows, rapeseed oilcake, protein, fatty acids, milk 513 Cations and Anions Contained in Diets and its Effect on Indices of Acid-Base Balance of Dairy Cows J. Twardon , R. Bodarski , S. Kinal , S. Krzywiecki , J. Pres , 3 R. Mordak 1 Wroclaw University of Environmental and Life Sciences, Department and Clinic of Obsterics, Ruminant Diseases and Animal Health, Wroc_aw, Poland 2 Wroclaw University of Environmental and Life Sciences, Department of Animal Nutrition and Feed Science, Wroclaw, Poland 3 Wroclaw University of Environmental and Life Sciences, Department of Internal and Parasitic Diseases with Clinic for Horses, Wroclaw, Poland Objectives of study: Some scientific data indicated that dietary cationanion balances (DCAB) values for cows in lactation have an influence on the yield and milk composition, as well as on some blood biochemical indices of acid-base balance. The aim of presented study was determination of DCAB values of Polish feeds and diets. Materials and Methods: The monitoring studies were conducted on the seven dairy cows herds from South-East Poland region. In feed samples the content of Na, K (AAS method), S (nefelometrically) and Cl (titrate method) was determined. On the base of obtained data the DCAB values of diets from all farms were calculated. In blood samples taken from cows some indices of acid-base balance were determined. Results: The highest content of K was stated for grass and legumes silages. In maize silages levels of K were very various. The concentrates contained medium levels of K, whereas brewery grains and beet pulp silage were characterized by low concentration of this element. The content of Na was low in grass and legumes silages, as well as in bran and maize. The concentrates contained slightly higher level of Na. The highest Cl content was stated for grass, legumes and maize silages. In bran and rape seed meal the level of Cl was low. The highest content of S was observed for concentrates, brewery grains and for grass and legumes silages. Maize silages contained the low levels of S. The DCAB levels of diets oscillated between +66 and +228 meq/kg DM and were lower then optimal - +300 meq/kg DM. This situation could be explained by high share of maize silage in diet and simultaneous low share of grass and legumes silages. Obtained BE indices were higher than reference value that stated about alkalosis occurrence. The high level of BE indices was linked with act. HCO3 values, which were higher than normal concentration. The increase of ionized calcium concentration was associated with high BE

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Nutrition and Metabolic Disorders 33

and HCO3 level. The concentration of K ions holds in reference values. The concentration of Na ions was various, but still was on proper levels. Conclusions: The DCAB levels of diets oscillated between +66 and +228 meq/kg DM and were lower then optimal. Obtained BE indices were higher than reference value, that stated about alkalosis occurrence. Key words: dietary cation-anion balance, feeds, dairy cows 514 GH Serum Values in Dairy Cows with Clinical Primary Ketosis A. Cuomo , L. Scaletta , G. Bertoni , L. Esposito 1 Faculty of Veterinary Medicine, Dipartimento di Scienze Cliniche Veterinarie, Teramo, Italy 2 University Cattolica Sacro Cuore, Istituto di Zootecnica, Piacenza, Italy 3 Faculty of Veterinary Medicine, Dip. Scienze Zootecniche ed Isp. Alimenti, Napoli, Italy The objective of the present study was to investigate the GH values in dairy cows with spontaneous clinical ketosis. Eight ketotic Holstein Fresian cows (KT group) within the sixty days after parturition, with ßhydroxybutyrate (BHBA) values higher than 1.2 mmol/L and nonesterified fatty acid higher than 0.6 mmol/L, were evaluated for metabolic profile, plus RIA for insulin and GH. Before sampling the clinical diagnosis of ketosis was confirmed by portable dry analyzer Medisense-Optium Abbott. A control group of six normal no lactating cows (CT group) was evaluated in the same way. Any subject was treated by drugs before sampling and the samples were collected at 09.00 a.m. in both groups. Statistical analysis was performed respectively in the groups and between the groups, by software GraphPad Prism 5. In the ab group KT mean value was respectively for GH 2.473 ng/ml (SD bcA bC 1.038), insulinemia, 4.570 µU/ml (SD 2.101), glycaemia 46.00 ab mg/dl and (SD 7.746), BHBA 2.213 mmol/L (SD1.606) and NEFA ac 1.796 mmol/L (SD 0.5299). In the CT group mean value was a bB respectively for GH 2.162 ng/ml (SD 0.9184) insulinemia 8.445 ABC µU/ml (SD 1.082) glycaemia 71.50 mg/dl (SD 8.264) BHBA a a 0.283333 mmol/L (SD 0.075277) and NEFA 0.1904 µmol/L (SD 0.1692). All the results are in accordance with the literature except the GH in KT group. Usually in cow 60 days post-parturition the GH value ranges around 5-10 ng/ml, significantly different respect our data in ketotic cows for the same period of lactation. Statistical analysis shows between the groups significance for insulinemia, glycaemia, BHBA and NEFA values as expected, at difference of GH that is not significant. No correlation has been observed between NEFA and GH values in the KT group, at difference of positive correlation described in literature for ketosis-free lactating cow. We hypothesize, that low GH concentration in dairy cow affected by primary ketosis, could have a protective effects against more pronounced fats mobilization during NEB. Key words: cow, ketosis, GH 515 Romifidine as Treatment for Primitive Ketosis in Dairy Cow A. Cuomo , L. Scaletta , G. Bertoni , L. Esposito 1 Faculty of Veterinary Medicine, Scienze Cliniche Veterinarie, Teramo, Italy 2 University Cattolica Sacro Cuore, Istituto di Zootecnica, Piacenza, Italy 3 Faculty of Veterinary Medicine, Dip. Scienze Zootecniche ed Isp. Alimenti, Napoli, Italy Romifidine (RMF) is an a-2 agonist utilized like analgesic-sedative drug in dog and horse, and rarely in cow. The a-2 agonist drugs induce a transient anabolic effect with hyperglycaemia, biphasic hypohyperinsulinemia and a rise of GH concentration. The objective of this work was to evaluate the effectiveness of Romifidine in the treatment of ketosis and its metabolic effect in dairy cows. Eight dairy cows affected by clinical ketosis (KT group), with ß-hydroxybutyrate (BHBA) values higher than 1,2 mmol/L and nonesterified fatty acid (NEFA) higher than 0.6 mmol/L, were included in the group. All the animal displayed the syndrome in the sixty days after parturition and do not received treatment before our study. Six not lactating cows with normal BHBA values were enrolled in the control group (CT). Both groups were evaluated, for metabolic profile plus GH and insulin, to study the RMF effects in a negative energy balance mode (NEB) like group KT and in a positive energy balance (PEB) like group CT. In KT group RMF was injected at

1 1 2 3 1 1 2 3

09.00 a.m. for 5 days at dose of 7.5 ϵg/Kg/i.m, after collection of basal blood sample and evaluation of levels of BHBA, by portable dry analyzer Medisense-Optum Abbott. Adjunctive sample was collected at day 1 after 2.5 hours from RMF injection. In CT group the samples were collected before injection and at 1; 1.5; 2; 2.5;3; 24 hours after it to evaluate a complete daily curve. Each sample was processed for metabolic profile plus insulin and GH and data were statistically processed. The sedative RMF at 7.5 µg/Kg/i.m. induces a significant increase in GH and glycaemia values and a biphasic hypo-hyperinsulinemia in both groups, without any sedation effect. GH mean value shows significant differences between KT and CT groups, before and after 2.5 hour RMF injection ab A respectively of 2.473 (SD 1,038) and 9.225 ng/ml (SD 8.774) in KT a a and 2.162 (SD 0.9184) and 3.022 ng/ml (SD 1.206). In the group KT BHBA mean value falls down at 1 mmol/L (SD 0.7838) and food intake normalizes within 5 days. RMF is effective as unique treatment for primitive ketosis in dairy cow. We hypothesize, to explain this action, a role of described hyperglycaemia and transient hypo-hyperinsulinemia, plus the normalization of GH values, without the predictable increase values of NEFA, wich is typical for GH action in NEB cow. For this reason we could hypothesize some additionally, direct actions of RMF not correlated with GH activity. Key words: cow, ketosis, G, romifidine 516 Mycotoxin Screening in Healthy Cows and Cows with Abomasal Displacement A. Alkaassem , M. Fürll , S. Dänicke 1 University of Leipzig, Medizinische Tierklinik, Leipzig, Germany 2 Federal Agricultural Research Center (FAL), Animal Nutrition, Braunschweig, Germany Objectives: The aim of this study was a) to examine samples of fodder, blood, milk and bile for concentration of deoxynivalenol (DON), Zearalenone (ZON) and their metabolites de-epoxy-DON, a-zearalenol (ZOL), ß-ZOL, zearalanon (ZAN), a-Zearalanol (ZAL) as well as ßZAL, b) to study possible relations between the toxin arrears and diseases. Material and methods: We checked 61 ill cows with abomasal displacement and accompanying diseases as well as 13 healthy HF cows. Results: 77.8% Patients without mycotoxins were cured. From those with mycotoxins 18 (72%) were cured without clinical reactions; 7 had to be euthanized due to severe accompanying diseases and severe haematological or clinical chemical deviations in the blood being typical for abomasal displacement. We do not observed specific clinical mycotoxins effects on health. ZON/ZOL associated changes in the ovaries and uterus could not be macroscopically determined. The transabdominale collection of bile by aspiration can be easily performed in sick cows. In healthy cows mycotoxins were not found. In the slightly contaminated fodder samples the mycotoxin concentration were 0.161 g DON /kg (0.086-0.191) and 6.35 µg/ ZON kg (4.88-7.85). None milk samples contained mycotoxins. DON (0,002 µg/ml) was found in one of 61 serum samples, de-epoxy-DON in four samples (8%). In bile de-epoxy-DON (37.6 µg/ml) was found only once. 39% of the examined bile samples were contaminated with ZON and/or its metabolites (ZON 9.85 ng/g [8.10-16.33], b-ZOL: 59.9 ng/g [5-78] and ß-ZOL: 37.6 ng/g. In all cases concentrations were within the physiological range. The metabolic parameters protein, albumin, urea, kreatinin and TEAC did not show differences in cases with or without mycotoxins. Mg, Ca, Na, K, Cl, pH, PCO2, bilirubin, glucose, FFA and cholesterol concentrations were not in physiological ranges in all cases independed on mycotoxins. The haemogram of all patients with or without mycotoxin did not show deviations. AP, GGT and CK activities of all patients were within the normal ranges. However mean AST and GLDH activities of all mycotoxin positive patients were slightly increased compared to mycotoxin negative cows. Conclusions: We concluded that no specific clinical, haematology or clinical chemical changes for mycotoxin influences (DON, ZON) could be determined. 517 Strong Electrolytes (Sodium, Potassium, Chloride) Determine Hydrogen Ion and Bicarbonate Concentrations in Urine of Ruminants H. Staempfli , P. Constable 1 Ontario Veterinary College, Department of Clinical Studies, Guelph, Canada

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34 XXV. Jubilee World Buiatrics Congress 2008

2

Purdue University, Department of Veterinary Clinical Sciences, West Lafayette, United States

Urine contains 5 strong ions (high Na, K, Cl, and low Ca, Mg) and 4 buffers over the physiologic range for urine pH (2 volatile buffers [bicarbonate and ammonia] and 2 non-volatile buffers [phosphate, creatinine]. Translating physicochemical approach to urine it is hypothesized that the primary determinants of the urinary pH are the strong electrolyte concentrations represented by the strong ion difference (SID) [(Na +K)- (Cl)]urine. The theoretical polynomial relationship between SID vs pH and SID vs HCO3 in simulated urine containing strong ions and normal pCO2 predicts very high urine bicarbonate concentrations (>200 mEq/L). Normal urine samples were collected from 5 mature Holstein dry cows and analysed on a multianalyzer system (Hitachi 911) for concentrations of Na, K, Cl, and TCO2. PH (pHmeas) was measured using an Accumet-benchtop pHmeter. The pH (pHcalc) was then calculated using SID in the Stewart 8 model equation on an excel spreadsheet (MS Office 2003) with pCO2 kept at 40 mmHg and individual SID from each urine sample (table). The high level of bicarbonate concentrations observed in urine confirms that bicarbonate is a dependent variable adjusting to concentration of SID based on dissociation equilibria and law of electroneutrality. Electrolyte intake and renal excretion are very closely correlated and alkaline pH in urine of ruminants is strongly associated with surplus strong ion intake, especially with diets high in potassium concentrations. There was good correlation between pHcalc and pHmeas in this small sample size. Electroneutrality was off, indicating measurement problems with ion specific electrodes. In conclusion pH and bicarbonate concentration (TCO2) of urine are primarily dependent on SID. Strong electrolytes, bicarbonate (TCO2) and pH in Urine of 5 dry cows Na K Cl SID pHcalc pHmeas TCO2 E(mEq/L) (mEq/L) (mEq/L) (mEq/L) (mEq/L) Neutrality (mEq/L) 10 108 129 37 95 388 276 246 306 320 143 110 204 133 178 255 274 171 210 237 8.4 8.4 8.2 8.3 8.4 8.5 8.4 8.4 8.3 8.5 229 227 91 148 218 -26 -47 -80 -62 -19

at first day and decreased while treatment. Antithrombin and factor XIII decreased in cows which died faster than in cows which were cured. The fibrin degradation products were slightly increased and were sunk while treatment. Cows, which died, had highly increased dDimers. Procalcitonin and haptoglobin were increased in ill cows and decreased during treatment. Conclusions: Dairy cattle which suffer from fat mobilization syndrome or endometritis have a poor prognosis concerning the acute phase proteins fibrinogen, haptoglobin and procalcitonin and the haemostatic parameters especially antithrombin, factor XIII and dDimers. A single dose of dexamethasone has no adverse effects to the coagulation, but shows a stabilizing effect. An additional administration of heparin is indicated. Key words: cattle, blood coagulation, fat mobilisation syndrome, coagulation disturbances 519 Oxalates Plants: Reviewed of Eco Toxicological Aspects among Farm Animals M. Abdallah , T. Abdelkrim , K. Gerard 1 Centre Universitaire d'El-Tarf, Veterinary Department, El-Tarf, Algeria 2 Université d'Annaba, Biology Sciences, Annaba, Algeria 3 Ecole Vétérinaire, Veterinary Department, Lyon, France The poisonings by oxalic acid and organic salts (the oxalates) are especially known of the veterinary practitioners as poisoning due to the ethylene glycol (antifreeze), which oxidized in oxalic acid in the liver, is at the origin, in particular, of interstitial nephritis and deadly epithelial among the domestic animals (carnivores). However, some plant poisonings to the oxalates are not diagnosed correctly, probably by ignorance. They are due to plants rich in oxalates and which diagnosis is not so difficult. The aim of our work is to bring a maximum of relative information to the eco toxicological aspect of these toxic substances composing these plants. The example of the oxalis is given, very rich oxalates plant that infests the natural grasslands as those sown. The pathological aspects that especially interest the farming activity veterinarian more that the canine practice are discussed as well as the pathogenic aspect of the poisonings in different animal species, especially in bovine species (cattle). Key words: oxalates plants, oxalic acid, eco toxicology, pathologies, cattle 520 Saponins and Pithomyces chartarum Spores Evaluation in Pastures from Southwestern of Goiás State, Brazil K. Brum , MC. Fioravanti , M. Haraguchi , LA. Silva , 4 5 1 1 1 F. Riet-Correa , JR. Borges , J. Souza , E. Campos , M. Pires , 1 A. Pinto 1 Universidade Federal de Goiás, Escola de Veterinária, Goiânia, Brazil 2 Universidade Federal do Mato Grosso do Sul, Campo Grande, Brazil 3 Intituto Biológico, Sao Paulo, Brazil 4 Universidade Federal de Campina Grande, Centro de Saude e Tecnologia Rural, PATOS, Brazil 5 Universidade de Brasília, Faculdade de Veterinária, Brasília, Brazil Brachiaria species are important forages in tropical areas. In Brazil, there are around 51 millions of hectares that are feed sources to livestock. Outbreaks of hepatogenous photosensitization have been reported in ruminants grazing B. decumbens and B. brizantha. The disease was attributed to the presence of Pithomyces chartarum spores, but recent studies suggest that the steroidal saponins contained in the forage could initiate photosensitization outbreaks. General objectives of this study were verify if the quantity of protodioscin, a steroidal saponin, and the spores quantity variation during the year; and to inquire if the saponin was present in other forages that was grazing by the animals. With two month of interval, six harvests was taken in ten farms located in Jataí, Mineiros and Perolândia cities of the southwestern region of Goiás State, Brazil. The spore count was made second a standard technique. The butanol-soluble residue containing saponin was submitted to thin layer chromatography and spectrophotometric analysis using Ehrlich reagent in 515nm. P. chartarum spores found in the pastures varied of 0 to 40.000 spores/g The number of of grass and the biggest amount was observed in the July month (dry season in Brazil). Protodioscin was isolated only in B. decumbens and B. brizantha leaves; the biggest

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Key words: acid-base, urine, strong-ions, electrolytes, pH 518 Fat Mobilization Syndrome and Blood Coagulation in Cows K. Waldau-Ringel , M. Müller , M. Fuerll 1 University of Leipzig, Department of Internal Medicine, Leipzig, Germany 2 University of Leipzig, Department of Clinical Diagnostics, Leipzig, Germany Objectives: During the fat mobilization pro- and anti-inflammatory cytokines are released from the adipose tissue. These could cause a circulatory insufficiency and the lipolysis is increased by insulin inhibition. Dystocia with placental retention and endometritis following bacterial determined endotoxin release are caused by enhanced cytokine development. Cytokines could activate the coagulation cascade, so that hemostasis disorders and a disseminated intravascular coagulopathy are found. The aim of the study was to evaluate, if dairy cattle with fat mobilization syndrome or endometritis have coagulation disorders in dependence of disease at admittance into the clinic. Material and methods: Blood was collected from 60 cows, which were admitted into the clinic because of abomasal displacement and accompanied diseases. From these 60 cows, a basal therapy was given to group 1 (n=25) and additional a single dose dexamethasone was given to group 2 (n=25). 10 cows died in spite of intensive care. Comparative 36 healthy cows were tested. The numbers of thrombocytes and leucocytes were analyzed in EDTA blood and a number of haemostatic parameters like prothrombin time (PT), fibrinogen (FIB), reptilase time (RT), Antithrombin (ATIII), fibrin degradation products (D-Dimers), factor XIII (F XIII) and the activated protein C (APC) in citrated blood. Additional as acute phase proteins haptoglobin and Procalcitonin (PCT) were tested. Results: 50 cows could discharge from clinic after 4 days. 6 cows died within 24 hours and 4 cows died after 4 days. Fibrinogen was increased

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Nutrition and Metabolic Disorders 35

amount of saponins was observed in March, in the rain season, and the lowest amount in July. These results shoed the great variation in levels of saponins during the experiment, suggesting a relation between the pluviometric precipitation and/or cycle of life in these species of Brachiaria evaluated. Acknowledgements to Coordenaçao de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and to Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) by financial support. Key words: brachiaria, spectrophotometry, sporidesmin, photosensitization, protodioscin 521 The Effect of Dry Period versus Continuous Lactation on ßcarotene, Vitamin A and Vitamin E in the Mammary Secretion F. Schweigert , A. Wesser , I. Buchholz , J. Raila 1 University of Potsdam, Department of Physiology and Pathophysiology, Potsdam-Rehbruke, Germany 2 University of Leipzig, Department of Veterinary Physiology, Leipzig, Germany ß-carotene and fat-soluble vitamins A and E are accumulated during the dry period in the mammary gland and secreted with colostrums together with the accumulated immunoglobulins. Both are of great importance for the health of the new borne calf. In this study changes in fat-soluble vitamins A, E and ß-carotene (rpHPLC) and IgG1 (immunodiffusion) in the secretion of the mammary gland during late gestation in dry dairy cows (n=12) and those which were milked until parturition (n=5) were compared over a total period of 11 weeks starting 8 before parturition. Results showed a biphasic increase in all components during the dry period. The first approx. 30 d ap and the second closely toward parturition. In animals with continuous lactation an increase towards parturition was observed as well but maximum levels were reached earlier approx. 20 ap. Absolute levels were comparable in both groups. In both groups low levels characteristic for mature milk were reached within a few days after parturition (P<0.05). The increase of fat-soluble vitamins in secretion was observed despite a continuous decrease in milk fat indicating no lipidassociated transfer of these components. Similarities between IgG1 and fat-soluble vitamins point to mechanism responsible for the accumulation of these components in milk. Finally, the accumulation in colostrums is not effected by the dry period but related to endocrine changes associated with parturition. Key words: fat-soluble vitamins, ß-carotene, dry-period, milk, cattle 522 Bovine Hepatic Function in Brachiaria Pastures from Southwestern of Goiás State, Brazil K. Brum , MC. Fioravanti , M. Haraguchi , LA. Silva , 4 5 1 1 1 F. Riet-Correa , JR. Borges , J. Souza , E. Campos , M. Pires , 1 A. Pinto 1 Universidade Federal de Goiás, Escola de Veterinária, Goiânia, Brazil 2 Universidade Federal do Mato Grosso do Sul, Campo Grande, Brazil 3 Intituto Biológico, Sao Paulo, Brazil 4 Universidade Federal de Campina Grande, Centro de Saude e Tecnologia Rural, PATOS, Brazil 5 Universidade de Brasília, Faculdade de Veterinária, Brasília, Brazil Brachiaria species being constituted by B. brizantha (60 millions ha), B. decumbens (25 millions ha) and others (10 millions ha) which are used to livestock. Outbreaks of hepatogenous photosensitization have been reported in ruminants grazing B. decumbens and B. brizantha. The disease was attributed to the presence of Pithomyces chartarum spores, but recent studies suggest that the steroidal saponins contained in the forage could initiate photosensitization outbreaks. General objectives of this study were: to determine the presence of hepatic alterations, by seric dosage of hepatic enzymes activities and bilirubin values and to verify if there was a relation between the quantities of saponin protodioscin, the number of P. chartarum spores and hepatic alterations. With two months of interval, six harvests was taken in ten farms located in Jataí, Mineiros and Perolândia cities, in the southwestern region of Goiás State. In each harvest the animals was weight and the blood samples was taken to determination the serum activity of gama glutamyltransferasis, aspartate aminotransferase and bilirrubin values. In each farm twenty bovines had been sampled. For the accomplishment of seric dosages standardized commercial reagents were used. The spore count was made second a standard technique and

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saponins were determinate to thin layer chromatography. The average of all biochemistry values had remained inside of the normality parameter. About hepatic alterations tests, only aspartate aminotransferase show statistical difference (p<0.05) on third harvest, in first month of rain period, when the greatest number of animals had increased this enzyme activity. The biochemical results also show few significant alterations indicating absence of consistent hepatic or biliary lesion, during the experimental period. Only the presence of small amounts of saponin protodioscin and a few spores it was not sufficient to develop photosensitization outbreaks in the studied bovines. Acknowledgements to Coordenaçio de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and to Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) by financial support. Key words: hepatic function, Brachiaria brizantha, Brachiaria decumbens, sporidesmin, protodioscin 523 A Novel One-Step Method for Fat-Soluble Vitamins and ßcarotene in Blood at Cow-Side F. Schweigert , R. Mothes 1 University of Potsdam, Department of Physiology and Pathophysiology, Potsdam-Rehbruke, Germany 2 BioAnalyt GmbH, Potsdam, Germany Background: Fat-soluble vitamins (FSV) and ß-carotene (BC) are important blood components associated with cell differentiation and growth. Monitoring their blood levels is important to optimise performance of highly productive animals through optimised feeding. Currently the determination of FSV and BC from blood is a time consuming and cost-intensive multi-step procedure, which needs specific equipment for extraction and centrifugation. Finally, FSV and BC are determined by spectrophotometric or HPLC methods in qualified analytical laboratories. Aim: Aim of this study was to compare the extraction efficacy of a recently introduced new and innovative one-step extraction procedure with standard methods. Methods: serum or total blood samples from cows (n=172) BC were analysed by HPLC and compared with the novel assay system for FSV and BC consisting the iExblood all-in-one extraction unit and by a hand held photometer, the iCheck. These two components enables to extract of FSV and BC from blood without prior separation of plasma in a single step at cow-side. In a subset serum was extracted either with the disposable assay system iEx blood or the standard measurements regularly used for HPLC analysis and finally analysed by HPLC. Results: BC values range from 0.32 mg/L to 15.30 mg/L. No differences were observed between both methods for individual herds or for all animals either measured in serum or in blood (HPLC serum vs iCheck® serum: 3.46±2.43 vs. 3.69±2.46 and HPLC serum vs. iCheck® blood: 3.46±2.07 vs. 3.33±2.04. Furthermore, results correlated 2 (Pearson correlation coefficient) well with HPLC analysis (r =0.98 and 0.99, serum and blood respectively, both P < 0.001). With regard to vitamin E (VE) and vitamin A (VA) levels assayed were in the range of 1.38 to 8.30 mg/L and 0.21 to 0.34 mg/L respectively. No significant differences were observed between both methods for VE and VA measured in serum (standard extraction vs iEx extraction: 4.02±1.81 2 vs. 4.29±1.96 and 0.27±0.04 vs. 0.25±0.04 (r =0.97 and 0.82, VE and VA respectively, both P < 0.001). Conclusion: Results show that with the novel test system blood levels of FSV and BC can easily be assessed within a few minutes at cow-side achieving a comparable quality as with highly sophisticated time consuming and expensive laboratory method such as HPLC. Key words: fat-soluble vitamins, ß-carotene, cow-side assay, diagnostic 524 Effect of 2-Methyl-2-Phenoxyl Sodium Propionate on Lipid Profile in Postparturient Dairy Cows J. Bouda , MA. Blanco , J. Ávila , C. Hernández , EG. Salgado 1 Universidad Nacional Autonoma de México, Patologia, MEXICO, D.F., Mexico 2 Universidad Nacional Autonoma de México, Produccion Animal: Rumiantes, MEXICO, D.F., Mexico Fatty liver and ketosis are frequent metabolic disorders in dairy cows during the first weeks postpartum. The compound 2-methyl-2phenoxyl sodium propionate (MPSP) is an activator of the peroxisome proliferator activated receptor-a involved in energy

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36 XXV. Jubilee World Buiatrics Congress 2008

metabolism. The objective of this study was to evaluate the effect of the intramuscular administration of MPSP on blood serum lipid variables and selected reproductive parameters in dairy cows. Thirty nd th six Holstein cows within the 2 and 4 lactation with body condition score between 3.5 and 4.5, 10 to 6 days before expected calving date were randomly divided into two groups. Group 1 (n=18) received three application of 50 mL (5 g of MPSP) at 20 day-intervals. The first application was 10 to 6 days prepartum, the second and the third administration were 10 d and 30 d postpartum, respectively. Group 2 (n=18) receiving no treatment. Blood samples were collected in all cows from tail vein on day 10 to 6 before expected parturition date and on day 10, 30 and 40 after calving. Serum free fatty acids (FFA), b-hydroxybutyrate (BHB), cholesterol (CHOL) and triacylglycerols (TG) were measured. The results were analyzed with repeated measures analysis of variance. A Chi-square test evaluated the proportion of cows with increased concentrations of FFA (>0.4 mmol/L) and BHB (>1.4 mmol/L). There were no significant difference in FFA and BHB between Group 1 and Group 2 (P>0.05). Before parturition, BHB concentrations were lower than those after calving (P<0.05). The concentrations of CHOL and TG were higher in Group 1 on days 10 and 40 (P<0.05). There was no difference in the proportion of cows with increased FFA between both groups after parturition (P>0.05). The proportion of cows with increased concentrations of FFA (>0.4 mmol/L) before calving was higher in Group 1 (P<0.05). The number of cases of subclinical ketosis (BHB >1.4 mmol/L) was lower in Group 1 at days 30 and 40 postpartum (P<0.05). The open days were lower in Group 1 (109.9) in comparison with Group 2 (118.5) (P<0.05). The preventive administration of MPSP reduced the frequency of subclinical ketosis and increased the serum concentration of CHOL and TG during the first 40 days postpartum. Key words: PPAR-alpha, dairy cow, lipid profile 525 Ammonia Detoxication Efficacy in Cattle Adapted or not to Dietary Urea and afterwards Undergone to Different Privation Periods N. Santos , E. Ortolani , C. Mori , A. Antonelli , M. Sucupira , 3 S. Kitamura 1 Universidade Federal Rural de Pernambuco, Veterinary Medicine, Recife, Brazil 2 Universidade de Sao Paulo, Department of Clinical Science, Sao Paulo, Brazil 3 Universidade Anhembi Morumbi, Department of Veterinary Medicine, Sao Paulo, Brazil Objectives of study: This study was carried out to investigate the recommendation stated by HUBER (1978) that there is a need to readapted cattle that were fed previously with dietary urea, but subsequently were deprived of this nutrient for more than 3 d. Materials and methods: Twenty four steers fed a low crude protein ration, deprived of urea, were randomly distributed into 5 equal groups, and were fed as follows: O (control; basal diet); the remaining groups (A,B, C and D) were fed the basal diet + urea (1% D.M.) for 21 d; afterwards the groups B,C and D were fed only the basal diet for 4, 15 and 30 d, respectively. Thereafter, all steers were challenged with an intravenous infusion of ammonium chloride (1.5 M) up to the outcome of convulsion when the infusion was stopped. Throughout the infusion and for 3 h later clinical examination was carried out; blood samples and whole urine were also collected. Results: The steers adapted to urea (A,B,C and D) needed a greater amount of ammonium chloride to cause convulsion (p < 0.001) and took longer to exhibited this symptom than the control group. As compared to the control, all the adapted groups, but the C, had a less severe clinical picture and recovered sooner, and did not require any medical treatment after the end of the experiment. This resistance was due a faster fall in the blood ammonia level, caused by a greater activity of the urea cycle, excretion of a higher excretion of ammonium and urinary volume. It was demonstrated that the higher the production and excretion urea by the kidneys, the greater the diuresis, that by its turn increased the excretion of ammonium into the urine improving the efficiency of detoxification, principally at the most critic moment of the st poisoning (from the convulsion up to 1 h later). Conclusions: It was concluded that cattle fed previously with urea and afterwards restricted of urea for up to 30 d do not require a new readaptation period, contradicting Huber's axiom. Key words: urea, cattle, ammonia, intoxication

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526 Do Cows with Subclinical Milk Fever have Low Skin Temperature? Can We Use Skin Temperature for Diagnosis of Hypocalcaemia? L. Carstensen , U. Doerfeld , J. Deitmer , L. Nielsen 1 Boehringer Ingelheim Denmark, Division Vetmedica, Copenhagen, Denmark 2 Veterinary Practice, Reinsberg, Germany 3 Boehringer Ingelheim Vetmedica GmbH, Ingelheim am Rhein, Germany Objectives: Subclinical hypocalcaemia, which may precede clinical milk fever or may stay subclinical, is characterised by low level of blood calcium without the clinical signs of milk fever. Subclinical milk fever is difficult to diagnose, but also affects cows due to reducing the rumen function and dry matter intake (1) and increasing the risk of secondary post parturient disorders (2). The aim of the present study was to investigate the association between blood ionized calcium level (iCa) and ear skin temperature (EST). A second aim was to make a model to predict iCa from EST, and to evaluate whether measurement of EST can be used as an indicator of subclinical hypocalcaemia and early diagnosis of clinical milk fever. Materials and methods: The study was conducted in a large commercial dairy herd in Germany. EST was measured in triplicate at two well defined areas at the cow ear with an infrared light thermometer in 57 Friesian Holstein cows immediately before and after calving. Blood was drawn from Vena jugularis in serum vacutainers and the iCa was measured immediately with a VetStat analyser (IDEXX). Triple measurements of room air temperature (RAT) were also recorded. Data were analysed in SAS using ANOVA with EST as response variable, and iCa, RAT, parity as independent variables. Hereafter simulations were made in order to find the optimal inverse model with iCa as response variable and EST, RAT and parity as independent variables. Results and conclusion: The results showed that the EST after parturition was dependent on iCa, parity and RAT. The effect of parity st nd was that 1 and 2 parity cows were sometimes hypocalcaemic, but did not show signs of reduced peripheral blood perfusion as measured rd th by EST. In 3 to 5 parity cows, the EST was positively correlated with iCa. There was a strong effect also of RAT, which hence should be maintained in the model. It was possible to make several models to predict iCa from EST. The sensitivity and specificity of these models for use as a diagnostic indicator of hypocalcaemia however were too low. With more data it may be possible to develop a valid predictive model. Key words: dairy cows, hypocalcaemia, skin temperature, diagnostic models 527 Effect of Feed Supplementation on Health and Milk Yield of Dairy Cows Grazing Endophyte Infested (N. lolii) Yatsyn-1 Ryegrass Pastures F. Lanuza , H. Uribe , A. Torres , E. Cisternas , O. Araya , 2 3 1 1 1 F. Wittwer , S. Moyano , C. Uribe , L. Angulo , M. Villagra 1 Instituto de Investigaciones Agropecuarias INIA-Remehue, Producción Animal, Osorno, Chile 2 Universidad Austral de Chile, Facultad de Ciencias Veterinarias, Valdivia, Chile 3 Instituto de Investigaciones Agropecuarias INIA-La Platina, Agricultura Sustentable, Santiago, Chile The aim of this trial was to evaluate the effect of adding feed supplements on dairy cows grazing N. lolli infested ryegrass and white clover permanent pastures. The feed supplements were added to minimize the presentation of the ryegrass stagger syndrome. The research used 16.2 ha of Yatsyn-1 ryegrass and white clover pasture of the dairy unit at the Remehue Regional Research Centre. The land was subdivided in 3 sections, each one of 6 to 9 paddock of the same area. Thirty-nine cows calving in spring and after lactation peak, from December 2001 to March 2002, were used. The pasture was grazed when it had a grass availability of 1,800-2,200 k DM/ha. Switching decision to a new paddock was taken when residual grass was 6 cm height measured with falling plate meter. Cows were assigned to a repeated measurements random block design into three treatments: I Grazing + 0.15 kg mineral salt supplementation; II Grazing + 30 kg silage and .15 kg mineral salt supplementation; III Grazing + 5.6 kg concentrated feed, 0.15 mineral salt and 0.2 kg sodium bicarbonate.

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Nutrition and Metabolic Disorders 37

Water was offered ad-libitum. Feed supplements were offered in the milking parlor (treatments I and III) and 2-3 hours before milking (treatment II). Stocking rate for the 86 days trial were 2.07, 2.08 and 2.57 cows/ha for treatments I, II and III, respectively. In forage samples, in average, concentration (mg/Kg DM) of Lolitrem B (0.9±0.1) was higher than Ergovaline (0.5±0.1). Daily milk yield, b ab a adjusted to 4% fat content, was 12.9 , 14.5 and 15 L. for treatments I, II and III, respectively (P<0.05). Although some forage samples had a concentration of 1.53 mg/kg DM, observation of the cows, on grazing paddocks and milking parlor, did not provide evidences of ryegrass stagger clinical signs. Serum activity of Aspartate aminotransferase (AST) and Gama-glutamiltransferase (GGT) were between reference ranges. Creatine kinase (CK) was above reference range but was at permitted range for samples taken after physical exercise. It is concluded that feed supplementation has a dilution effect on toxic intake; however, in weather conditions with low rainfall and high humidity, leaving residual grass of 6 cm, did not allow us to observe clinical signs of ryegrass stagger syndrome in non supplemented grazing cows. 99C5AT FDI-CORFO/INIA Project Key words: supplementation, grazing dairy cow, endophyte 528 Assessment of Three Sources Used for Magnesium Supplementation in Grazing Dairy Cows G. Wittwer, D. Scandolo, H. Chihuailaf, L. Böhmwald, A. Contreras Universidad Austral de Chile, Facultad Ciencias Veterinarias, Valdivia, Chile Objective: Evaluate the effect of three magnesium (Mg) sources supplementation on plasma and urinary excretion of (Mg) in dairy cows at pasture consuming grass with high potassium content. Material and Methods: Experimental animals were 27 Frisian lactating cows, grazing on pastures, mostly Lollium sp, ( Mg= 0,17%; K= 2,68% DMB). Cows were distributed in 4 groups and supplement with 30 g/d p.o as MgSO4, MgO or MgHPO4 administered in one dose th after the morning milking during 5 consecutive days. The 4 group remained as control. Blood and urine samples were obtained on days 0, 1, 5 and 12 (7 days after supplementation). Plasma and urine Mg concentrations were determined by AAS. The clearance of urinary Mg was also calculated (CUM = Mg-U/creatinina-U). Differences between periods were determined by repeated measures ANOVA test with a significance of 95% Results: Mg supplementation with the three sources had not significant effect (p>0.05) in the plasma Mg values. CUM values increases (p<0.05) when MgHPO4 was used, also in the MgO group, but in a less and shorter extension (Table 1). The morning ruminal environment of cows grazing high K content pastures could allow a better utilization of Mg when phosphate is used compared to oxide; and sulphate seems not to be a efficient way to supplement Mg. CUM determination has a better sensitivity to evaluate Mg balance than plasma Mg concentration Table 1. Plasma magnesium concentration and clearance of urinary magnesium, CUM (mean ± SD), before, during (days 1 and 5) and after (day 12) a supplementation with 30 g of magnesium as sulphate, oxide or phosphate. Plasma magnesium (mmol/L) Days MgSO4 (n=7) MgO (n=7) MgHPO4 (n=7) Control (n=6) 0 0,61±0,06 0,54±0,17 0,60±0,08 0,47±0,11 1 0,62±0,06 0,54±0,17 0,60±0,12 0,42±0,06 5 0,65±0,07 0,60±0,16 0,70±0,17 0,46±0,10 12 0,66±0,07 0,55±0,09 0,62±0,09 0,49±0,12

529 Evaluation of Some Commonly Used Nutrition Software for Predicting Dry Matter Intake of Holstein Cows during the Mid-Lactation M. Taghinejad roudbaneh , A. Nikkhah , M. Kazemi 1 Islamic Azad University Tabriz, Animal Science Department, Tabriz, Iran 2 University of Tehran, Animal Science Department, Karaj, Iran Diets are typically formulated based on nutrient density. Because nutrient intake is a function of dry matter intake (DMI), predicting DMI precisely and accurately is important to prepare balanced rations when on farm estimates of feed intake are not available (Hayirli et al., 2003). Lack of accuracy in prediction of feed intake can affect on animal performance, animal health or environmental impacts of dairy industry (Fox et al., 2004). There are different software based on scientific models to formulating dairy cows' rations that predicted DMI can be different among them. The objective of this investigation was to evaluate the 2001 Nutrient Requirements of Dairy Cattle, CNCPS Cornell Net Carbohydrate and Protein System, CPM-Dairy Cornell Pennsylvania Miner and Spartan dairy ration balancer soft wares in predicting DMI of lactating dairy cows in mid-lactation. 32 lactating Holstein cows averaging BW=632±29kg, DIM=55±12d and MY=32±0.85kg were randomly assigned in a CRB design that each treatment has 4 cows in second and 4 cows in third lactation period. Four rations using the same feedstuff and similar inputs (BW=650kg, DIM=60d, MY=32kg, Milk fat=3.5% and Milk protein=3.2%) were formulated by each software for each group of cows. The formulated rations fed individually for 90 days three times a day to allow 10% feed refusal. Daily DMI and other trait were recorded. Data were analyzed using GLM procedure of SAS (1999). Differences for MY and milk composition were not significant between four groups. The variables used to evaluate DMI prediction were MSPE, MPE and RPE as explained follow: MSPE = mean square prediction error (kg2/d), MPE = mean prediction error (kg/d); RPE =relative prediction error (%). By increasing the RPE rate, accuracy of the DMI prediction decrease. The RPE for NRC2001, Spartan, CNCPS and CPM-Dairy were respectively 7.16%, 8.10%, 10.80% and 12.63%. The results of the present study showed that NRC 2001 predicts DMI more accurately than Spartan, CNCPS and CPM-Dairy. DMI prediction in NRC 2001 and Spartan is base of empirical equations. So they have more conformity to actual DMI monitored in farm. Key words: nutrition software, dry matter intake 530 Atypical Interstitial Pneumonia Associated with Sweet Potato (Ipomea batatas) Poisoning in Adult Beef Cows in the UK I. Mawhinney , S. Trickey , N. Woodger , J. Payne 1 Veterinary Laboratories Agency, Bury St. Edmonds, United Kingdom 2 Veterinary Practice, Norwich, United Kingdom Sweet potato (Ipomea batatas) poisoning in adult cattle causes acute respiratory distress and atypical interstitial pneumonia, and is frequently associated with a high case mortality rate. It has been described in various countries worldwide, such as USA, Japan, Australia and Brazil, and is caused by ipomeanol toxins associated with certain fungal infections of damaged tubers. In the case reported here, six beef cows died, and one heifer and a bull recovered, after acute respiratory distress, 8-12 days after introduction of sweet potato tubers at 7kg per head per day. None of the seven unweaned suckler calves showed signs of toxicity, nor did 3 other in-calf heifers with access to the tubers. Animals did not seem to respond to treatments and deaths occured within hours of first clinical signs. Two of the cows were necropised at one of the VLA's regional laborotories. Pathological changes, confirmed by histopathology, were characteristic of an atypical interstitial pneumonia. Samples of the tubers that were fed were tested for fungal contamination. Fusarium solani mould was found in the tubers, which were purchased as damaged stock from a wholesale vegetable market. Sweet potatoes have a high energy concentration and are palatable but they have been associated with a variety of health effects, one of which is acute respiratory distress characterised by development of an atypical interstitial pneumonia clinically and pathologically indistinguishable from fog fever. The toxic compounds in sweet potato toxicity have been extensively investigated by American and Japanese workers. Damaged tubers produce 3-substitued furans compounds such as ipomeamarone, itself a hepatotoxin, which are then metabolised by various fungi into other related toxins such as ipomeanine, 1-ipomeanol,

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CUM (mmol/L) MgSO4 (n=7) MgO (n=7) MgHPO4 (n=7) Control (n=6) 0,99±0,11

a, b, c a 1,44±0,35 1,41±0,38ab 1,43±0,45a a

1,71±0,28a 1,66±0,34ab 2,07±0,27b 0,81±0,19a

1,82±0,50a 1,85±0,53b 2,35±0,47b 1,06±0,18a

1,32±0,44a 1,24±0,46a 1,37±0,44a 0,84±0,33a

= P<0.05 between periods; a, b, c = P<0.05 between periods

Conclusions: The MgHPO4 present a better bioavailability than MgO and MgSO4 seems ineffective when used as supplement for Mg in cows at pasture with high K content. Key words: cows, magnesium

38 XXV. Jubilee World Buiatrics Congress 2008

4-ipomeanol and 1,4 ipomeanol. These compounds when absorbed into the animal are further converted by microsomal activity in pneumocytes to a toxic lung oedema factor, which is responsible for the pulmonary pathology. The increased use of sweet potatoes for human consumption in the UK was noted as early as 1975 by Coxon and others. They are now grown in Europe and increasingly fed as a by-products to cattle. This is believed to be the first such case published in the UK. Key words: sweet-potato, toxicity, respiratory, interstitial, pneumonia 531 Key Hepatic Gluconeogenesis Enzymes Activity in Response to Long-term Glucose Infusion in Dairy Cows B. Al-Trad , K. Reisberg , T. Wittek , G. Gäbel , M. Fürll , 1 J.R. Aschenbach 1 Faculty of Veterinary Medicine, University of Leipzig, Institute of Animal Physiology, Leipzig, Germany 2 Faculty of Veterinary Medicine, University of Leipzig, Medizinische Tierklinik, Leipzig, Germany Background and Aims: Periparturient ketosis in dairy cows is characterized by an imbalance of glucose and lipid metabolism. Insufficient gluconeogenesis is thought to be primarily caused by a lack of glucogenic precursors due to negative energy balance. However, it is not clear whether insufficient catalytic capacity of gluconeogenic enzymes might contribute to insufficient glucose production postpartum. The purpose of the present study was to investigate (1) whether a positive energy balance due to chronic glucose infusion has a depressive effect on selected gluconeogenesis enzymes activity in dairy cows and (2) how quick such depressive effect might be reversed after stopping surplus glucose supply. Material and Methods: Twelve midlactation dairy cows were maintained over a 28 d experimental period on continuous intravenous infusion of either saline (control group, six cows) or 40% glucose solutions (treatment group, six cows). The infusion dose was calculated as a percentage of daily energy (NEL) requirements, starting at 1.25% and increasing by 1.25%/d until a maximum dose of 30% was reached. Dose was then maintained at 30% for 5 d. No infusions were made between d 29-32. Blood samples were taken every 2 d and liver biopsies were taken on d 0, 8, 16, 24, and 32. Results: Blood glucose and insulin concentrations showed only slight and occasional increases while NEFA and BUN concentrations decreased by glucose infusion. Liver glycogen increased progressively during glucose infusion until reaching plateau between d 16 and 24. Liver glycogen fell back to baseline values on d 32 after cessation of glucose infusion. During the infusion period, no significant changes were observed in the activities of phosphoenolpyruvate carboxykinase and glucose 6-phosphatase. Fructose 1,6-bisphosphatase decreased and pyruvate carboxylase tended to decrease in the treatment group during the high dose of glucose infusion (i.e. infusion dose of 20% & 30% NEL requirements) but quickly returned to baseline after stopping the infusion. Conclusion: The activity of key hepatic gluconeogenesis enzymes in lactating dairy cows is not significantly affected by long-term increases in glucose supply. Excessive glucose improves body tissues energy status without impairing the role of liver as the main source of glucose production in dairy cows. Thus, dysregulation of gluconeogenic enzymes activity by a positive energy balance pre-partum is likely not a major cause of postpartum metabolic disturbances. 532 Milk Drop Syndrome in Dairy Ewes: Clinical and Diagnostic Approach N. Giadinis , G. Arsenos , G. Fthenakis , P. Tsakos , C. Dovas 1 School of Veterinary Medicine, Aristotle University, Thessaloniki, Greece, Greece 2 University of Thessaly, Thessaloniki, Greece 3 Institute of Infectious and Parasitic Diseases, Centre of Veterinary Institutes of Thessaloniki, Thessaloniki, Greece "Milk drop syndrome" in dairy sheep is a problem with considerable financial implications. However, it has not been thoroughly investigated and research data are scarce. In this paper, we present the results of a study, carried out from 2003 to 2007, into the factors involved in milk drop syndrome reported in 18 flocks of dairy sheep in Northern Greece. The appropriate clinical and diagnostic approach is also discussed. In each flock, five adult ewes were selected randomly among those with reported milk drop. Samples of blood, milk and faecal samples were collected from these animals throughout a milking period. Their feeding

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regime was also evaluated. Evaluation of milk yield was based on data of previous milking records. In 10/18 flocks, the subclinical mastitis, caused by coagulase negative Staphylococcus spp. or Streptococcus spp, was the reason for the reduced milk yield. In other flocks the reasons were contagious agalactia confirmed by isolation of Mycoplasma agalactiae in milk cultures (1 flock), inappropriate and unbalanced nutrition with a ration containing suboptimal protein (1 flock), chronic ruminal acidosis (1 flock) or Maedi-Visna confirmed by PCR, serum test for antibodies and histological findings in mammary glands (2 flocks). A combination of the above factors was observed in 3/18 flocks, while in one flock these factors were coupled with intense gastrointestinal parasitism. The results confirm that subcilincal mastitis is the major reason for reduced milk yield. The role of different pathogens and the need for proper management of animals in individual flocks imply that the clinical and diagnostic approach should involve a farm investigation before any measures are taken. Key words: dairy ewes, milk, mastitis 533 The Relation of ante Partum Backfat Thickness with Metabolic Disorders and Fertility Problems in Dairy Cows - a Long Term Study H. Pothmann-Reichl , F. Zimmer , I. Sommerfeld-Stur , C. Iben 1 University of Veterinary Medicine Vienna, Teaching and Research Farm, Vienna, Austria 2 Veterinary Clinic, Private Veterinary Practice, Oberkirchen, Germany 3 University of Veterinary Medicine Vienna, Clinical Department of Animal Breeding and Reproduction, Vienna, Austria 4 University of Veterinary Medicine Vienna, Department of Veterinary Public Health and Food Sciences, Vienna, Austria ObjectivesIn this retrospective study the influence of post partum change of back fat thickness (BFT) in cattle on fertility, metabolic problems, mastitis and milk yield was investigated. It is hypothesised that a higher decrease of back fat thickness post partum (p.p) has a negative impact on fertility and fat metabolism. Animals, materials and methodsFive dairy farms in Germany (292 cows) and two farms in Austria (111 cows) participated in this trial and 1045 lactation periods were analysed. The back fat thickness was measured with an ultrasound (3.5 MHz) between the tuber coxae and the tuber ischiadicum at the following days: 8 weeks ante partum (a.p.), 42 days a.p., day of parturition, 42 days and 100 days p.p. The programme "Inter-Herd"(Interagri, Earley Gate, UK) was used to record the data. Statistic evaluation was done by SPSS for Windows 14.0.1 using odds ratio (OR >1 increased risk, OR <1 decreased risk) as risk parameter and a significance level of p<0.05 and a confidence interval of 95%. ResultsA significant increased risk for cycling problems (OR 0.56), for mastitis (OR 0.48) and for abomasal displacement (OR 0.07) was found in cows with a BFT loss over 35 % within 42 days p.p. The relative chance for cows with BFT loss less than 35% within 42 days p.p. to develop displaced abomasum is very low compared to those who lost more than 35 % BFT. The same limit of BFT loss was found for cows within 100 days p.p. concerning mastitis (OR 0.49) and ketosis (OR 0.37). For cows within 42 days p.p. and a BFT loss over 40% the results showed a significant higher risk for developing orthopaedic problems (OR 0.29). ConclusionIn general body condition scoring is an important instrument to monitor feeding management. It is a practical tool for veterinarians as well as for farmers. Measuring BFT is an even more objective method to evaluate body condition and a key for professional herd health management. Feeding strategies that aim for a reduced loss of BFT (<35%) after parturition have a positive impact on herd health. Key words: cattle, backfat, ultrasound, metabolic, fertility 534 Trace Element Deficiency in the Azorean Cattle: an Animal Production, Health and Economical Constraint C. Pinto , J. Viana , P. Aranha 1 Serviço de Desenvolvimento Agrário de Sao Miguel, Veterinária, Ponta Delgada, Portugal 2 Sociedade Açoreana de Sabnes, Veterinária, Ponta Delgada, Portugal Dairy cattle production represents the principal industry of the Azores 5 archipelago, responsible for 27% (4,5x10 tonnes) of all Portuguese milk production. Cattle are reared all-year-round on intensive grazing systems; soils are of volcanic origin with some natural trace element (TE) deficiencies. In order to identify TE deficiencies in cattle (Zn, Cu,

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Nutrition and Metabolic Disorders 39

Se, I and Co) more than 500 blood samples were analyzed from 70 herds, several of them with problems of infertility, abortions, premature calving, weak calves, enzootic haematuria and high somatic cell counts. The majority (91%) of the analysis were performed in Holstein-Friesian (HF) cattle, of which 284 were milking cows (MC), 129 heifers (H), 13 calves (C), 43 bullfight cattle (BF) and 12 beef cattle (B). The results of the TE survey indicate that 92% of the animals had both low levels of plasmatic iodine and thyroxin (T4) 84% of the herds had at least one animal with Co deficiency (75% BF cattle, 54% B cattle, 51% H and 37% MC). About 31% of the MC had low levels of Se (glutathione peroxidase, GSH-Px1) and serum Cu, as well as 38% of serum Zn. The most severe deficiencies were identified in the heifers group. To complement these data, 7 clinical trials were developed using HF reared on pasture grazing systems. In each trial there were two randomly constituted groups: a control group and a group have given extra feed supplements. The individual weights of animals were measured every 14 days. Blood samples were taken at the beginning and end of the trials for TE levels and blood cell counts. Different supplementation protocols and products were implemented in each trial. The clinical trial results indicate that TE supplemented yearlings had significantly higher productive output and hematological parameters than the controls. The least laborious, most consistent and reliable supplementation system studied, was obtained with the intraruminal slow releasing bolus (SRB) from Alltrace, Agrimin (UK). However, similar results were obtained when Se and I were added to the drinking water. A TE survey should be included in the sanitary and management plans for each herd in order to enhance animal performance and health, as well as farm profitability. Key words: cattle, trace elements, mineral deficiency, azores 535 Influence of Postpartum Propylene Glycol Supplementation on Metabolism, Animal Health, Reproductive Performance and Production in Dairy Cows M. Iwersen, U. Falkenberg, R. Voigtsberger, W. Padberg, A. Mertin, W. Heuwieser Freie Universität Berlin, Tierklinik für Fortpflanzung, Berlin, Germany The effect of postpartum (pp) supplementation of 250g propylene glycol (Propy) per cow and day via a total mixed ration (TMR) on metabolism, animal health, reproductive performance and milk production was studied on a commercial dairy farm in Germany. A total of 258 multiparous Holstein-Friesian cows were enrolled in a randomized and controlled field study. On the basis of expected calving date and previous milk production animals were blocked and allocated to one of the two treatment groups. The TMR fed in group 1 (Control, n=131) was supplemented with Propy before offering it to group 2 (Study, n=127). The TMR was mixed and offered twice daily for a period of 40 days pp. Blood samples were collected within 12 hours at the day of parturition, on day four, ten, twenty and forty. Samples were centrifuged and serum was stored at 20 °C until analysis. Body Condition Score (BCS) and Back Fat Measurement (BFT) values were recorded on the same days. Serum samples were shipped to one commercial laboratory for the determination of Nonesterified fatty acids (NEFA), Betahydroxybutyrate (BHBA), Triglycerides, Ketone bodies, Bilirubin, Glucose and activities of Aspartate-aminotransferase (AST), Glutamate-dehydrogenase (GLDH), Alanine-aminotansferase (ALT) and g-glutamyl-transferase (GGT). Metabolic parameters were analysed by a 6-step UNIANOVA with repeated measurements using SPSS®-Software. ALT, GGT, GLDH, Insulin, Bilirubin and Glucose were comparable between the two groups. Concentrations of AST, Triglycerides, BHBA and Ketone bodies were significantly (p<0.05) influenced by treatment. Concentration of AST, Triglycerides, BHBA and Ketone bodies indicated a better metabolic situation, especially on day four, ten and twenty pp for cows treated with propylene glycol. Referring to the test day results from the local dairy herd improvement association the total yield of fat corrected milk (FCM, 4 %) did not differ between the two treatment groups in a period of 305 days pp. Also milk composition was not affected by treatment. Neither reproductive performance nor changes of BCS or BFT during the study period were significantly influenced by treatment. Although indicators of metabolic status were improved by pp use of propylene glycol, economic benefits are questionable for dairy farms with good management programs. As most important economic factors milk production and milk composition were not influenced supplementing Propy in a TMR fed pp.

536 The Effect of Time of Selenium Application before the Slaughter on their Concentration in Blood and Selected Tissues in Goats L. Sevcikova, A. Pechova, L. Misurova, L. Pavlata, R. Dvorak University of Veterinary and Pharmaceutical Sciences Brno, Clinic of Ruminants, Brno, Czech Republic The aim of the experiment was to evaluate selenium distribution in ruminant organism at different time intervals after the parenteral application. Twenty white shorthaired goats aged 6 months and weighing 29.3 kg on average were enrolled in the trial and divided to four groups. Animals of each group were given Selevit inj. ad us. vet. BIOTIKA (natrii selenis 2.2 mg/ml, tocoferoli acetas 25 mg/ml) intramuscularly in neck musculature in the dose of 0.2 ml/ kg live weight. Selevit was applied in the first group (D7, n=5) 7 days, in the second group (D3, n=5) 3 days and in the third group (D1, n=5) 1 day before the slaughter. On the day of the slaughter, blood samples were taken by puncturing vena jugularis. Immediately after the slaughter, tissue samples of liver, spleen, pancreas, kidneys, testicles, lungs and heart, diaphragm, tongue, thigh and neck musculature were taken. The samples were mineralized in microwave oven MILESTONE MLS 1200 and the concentration of selenium was determined by AAS hydride technique using SOLAAR M 6 (UNICAM) appliance. Blood concentration of selenium was measured in µg/l; tissue concentration was measured in µg/kg of fresh tissue. The results show that selenium concentration in blood and all the examined organs and tissues increased following Selevit application. As compared with control group (C), all experimental groups (D1, D3, D7) showed significantly higher selenium concentrations: blood (133.0, 150.4, 177.6 vs. 93.7), plasma (84.6, 86.6, 113.1 vs. 46.3), liver (616.95, 485.91, 539.5 vs. 170), spleen (171.3, 170.9, 194.4 vs. 133. 5), pancreas (203.0, 222.8, 250.3, vs. 158.6), myocardium (128.6, 135.4, 153.5 vs. 88.4), diaphragm musculature (66.5, 66.2, 78.4 vs. 51.4), tongue musculature (128.6, 135.4, 153.5 vs. 88.4), neck musculature (63.4, 68.5, 149.6 vs. 43.0). As far as selenium distribution dynamics is concerned, the highest concentrations were found in blood, plasma, spleen, pancreas, lungs, and heart, diaphragm and thigh musculature 1 day after the application. Later on, the values kept decreasing. On the contrary, selenium concentration in liver and kidneys were highest 7 days after the application. This indicates that selenium deposited th slower in these organs and its concentration was increasing until the 7 day after the application. The work was realized with the support of MSM Project No. 6215712403 537 Correlations between Milk Fat Content and Fatty Acid Composition in Cows Receiving Different Lipid Supplements at Four Dietary Fiber Levels H. Fébel , J. Várhegyi , J. Schmidt , I. Várhegyi 1 Research Institute for Animal Breeding and Nutrition, Herceghalom, Hungary 2 University of West Hungary, Mosonmagyaróvár, Hungary The fat content and milk fatty acid (FA) composition are influenced by a number of interacting dietary factors including the amount of fiber, the proportion of concentrate to forage, and fat supplement. The rate of ruminal degradation of fiber in feeds for lactating cows has received limited attention. The aim of this trial was to investigate the effect of feeding total mixed ratios (TMR) of different fiber level and to study different ruminal degradation rate of neutral detergent fiber (NDF) on milk fat content and FA composition. Milk fat content and FA composition were evaluated in 4 dairy units (A, B, C and D) using lactating Holstein cows fed TMR with 40.6% (A); 37.9% (B); 36.3% (C); 32.1% (D) NDF content. Diets were markedly different in rates of ruminal degradation of NDF. The TMR in farm A had 6.5%/h; B=8.0%/h; C=6.3%/h and lowest value was in farm D, 4.9%/h. The diets differed in FA composition. The lowest proportion of saturated FA (19.5%) and the highest polyunsaturated FA content (51.1%) were in diet D. TMR in farm C contained higher proportion of oleic acid (30.4%) and linolenic acid (6.3%) and the lowest amount of linoleic acid (37.9%). Milk fat content differed among the farms, the values were 3.77%, 3.47%; 3.61%; 3.9%, respectively. The highest value (3.9%) was obtained in farm D where the cows received the lowest NDF content TMR. Cows in farm D had significantly lower percentage of trans11-18:1 and conjugated linoleic acid (cis-9 trans-11 C18:2; CLA) in milk fat. In contrast, cows in farm B

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40 XXV. Jubilee World Buiatrics Congress 2008

had greater trans11-18:1 percentage in milk fat (2.39%) this response resulted in lower milk fat (3.47%). In the farm C the percentage of CLA, the ratio of trans11-18:1 to CLA and the ratio (6:3) of FA were very beneficial according to human health demands. Milk fat content was negatively correlated to trans11-18:1 percentage (r=-0.93) and to CLA in the milk fat (r=-0.85). Our data suggest that rate of ruminal degradation of NDF influences the milk fat content. Results showed that there was a negative correlation between milk fat content and degradation rate of NDF (r=-0.91). It can be assumed, that even suboptimal fiber levels in TMR containing slower degradable fiber may be sufficient to maintain proper rumen fermentation and level of milk fat synthesis. This finding suggests that in the TMR of cows not only the quantity of fiber fractions (NDF, ADF) has to be determined, but even their degradation rate should be taken into consideration. 538 The Effect of Antioxidant and Omega-3 fatty acid Supplement on Metabolic Status and Reproductive Performance in Postpartum Dairy Cows F. Tóth , Gy. Gábor , O. Balogh , G. Koppány , H. Fébel 1 Research Institute for Animal Breeding and Nutrition, Herceghalom, Hungary 2 Vitafort Ltd., Dabas, Hungary The objective of this study was to investigate the effect of different supplements on metabolic status, the onset of cyclic ovarian and the rate of re-conception function in the postpartum (pp.) period. The study was carried out on 66 pp. Holstein cows. The cows were fed the same total mixed ration in dry period and 5-7 pp. days were divided into the control (C) (n=16) and the experimental (E) group (n=50). The feed of E was supplemented with extra beta carotene, vitamin E, inactivated yeast, selenium and microelement chelates until 50 pp. d.; from 50 pp. days onwards the supplement was only 200 g grinded linseed (W3 fatty acid supplement). The average milk yield of E cows was 3 kg/day higher than C cows from 8th to 120th lactation day. The plasma level of BHB was higher in the E group. While both groups showed elevated NEFA level on day 30, only C group had increased NEFA concentration on day 50. Plasma concentration of urea was lower in the E group on day 90 and 120 pp. Serum level of carotene in the E group was higher during the whole experiment. P4 concentrations between Days 18 and 39 after AI were greater for cows fed linseed supplementation than the control cows. According to the P4 profile more than 80 % of the E group had cyclic ovarian activity versus only 62.5% of the C group. Although, the conception at first AI (41.2 vs. 57.1 %) and the overall proportion pregnant (47.1 vs. 71.4 %) was lower in the E group. The hypothesis that feeding linseed would increase conception rate of dairy cows was not supported by the present experiment. The higher P4 level of E cows means the positive effect of omega 3 fatty acids and may also have contributed to decrease embryo mortality. The applied supplements effected elevated milk production, but this higher production led to a negative energy balance (NEB) which is confirmed by elevated levels of NEFA and BHB. This negative effect of NEB is presumably connected to lower fertility performance. This work was supported by the project of OMFB-0121 Key words: conception rate, omega-3 fatty acid, antioxidant, metabolic status 539 Use of hypertonic and isotonic saline solution and furosemide for the treatment of the ammonia poisoning in cattle S. Kitamura , A. Antonelli , R. Barreto Júnior , C. Mori , 1 1 1 M. Sucupira , B. Caputi , E. Ortolani 1 Faculdade de Medicina Veterinaria da Universidade de Sao Paulo, Clinica Medica, Sao Paulo, Brazil 2 Universidade Anhembi Morumbi, Clinica Medica, Sao Paulo, Brazil 3 Universidade Paulista - UNIP, Instituto de Ciencias da Saude, Sao Paulo, Brazil To test the efficiency of some treatments for ammonia poisoning in cattle 25 steers, never fed non-protein nitrogen before, were used. Ammonium chloride solution was infused intravenously (iv) only once in each steer until the onset of convulsive episode, and then the infusion was immediately stopped. Thereafter, the animals were randomly distributed in one of the five different groups, as follows: 1) control (infusion 300 mL isotonic saline solution - ISS); 2) infusion (iv) of ISS 30 mL/kg BW throughout the following 4 h and

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administration by stomach tube of 4 L water. (W); 3) same as group 2, plus a single dose 2 mg furosemide/kg BW (iv) (F); 4) injection of 5 mL/kg BW hypertonic saline solution (HSS) (7.2 %) (iv) throughout the first 30 min, followed by 20 mL/ kg BW of ISS and 4 L W.; 5) Same as group 4 and a single dose of F. For the next 4 h after the convulsion several clinical variables and the blood concentration of ammonia, glucose, lactate, urea, creatinine, sodium and potassium, hemogasometric profile, urine global volume and urinary excretion of ammonium, urea and creatinine were followed. The clinical picture seen just before the treatment was classical and caused a characteristic systemic metabolic acidosis, compensated by reduction in the pCO2, hyperlactemia and hyperglycemia. At the convulsion the ammonia blood levels were high and similar among th the groups, but at the 120 min the animals treated with HSS had lower values than the control. The HSS infusion caused, during the first 30 min. a marked reduction in the packet cell volume and the heart rate. On the other hand, furosemide provoked an increase in the heart rate and neither prevented the onset of lung edema, nor an increase in the total excretion of urine. Cattle treated with HSS stood up and had the return of the rumen movements earlier than other groups. The therapy with HSS still increased the urinary volume and the total urinary excretion of urea and ammonium during the critical st 1 h of treatment, while the use of ISS had intermediate results. The efficiency of the treatment with HSS or ISS as seen by some variables of clinical improvement was better than the control group. The higher the urinary urea excretion was, the greater the ammonia elimination became. Although with lower efficiency as seen in the treatment with HSS, the ISS caused an improvement in the clinical picture and at the end of the experiment generated an adequate ammonia detoxification. Key words: ammonia intoxication (treatment), hypertonic, furosemide, cattle 540 Environmental Low Phosphorus Induced the Erythrocytic Membrane Damage: a Pathogenesis of Endemic Haemoglobinuria in Dairy Cattle JL. Li, Q. Si, FQ. Shi, SW. Xu Northeast Agricultural University, College of Veterinary Medicine, Harbin, China Objectives: Dairy cattle could display haemoglobinuria when phosphorus was deficient in food, especially in environmental low phosphorus in some regions. Endemic haemoglobinuria in dairy cattle usually had been detected in some regions of Heilongjiang province in china. However, the reason of the endemic haemoglobinuria remians unclear. The aim of this study is to illustrate the pathogenesis of endemic haemoglobinuria. Materials and Methods: In this study, we tested the content of phosphorus in soil-grass and serum of 15 cows with obvious clinical symptom in Daqing and Anda regions of Heilongjiang province in April-July and October, respectively. In addition, we also detected Lipid composition, fluidity, viscosity and activity of ATPase of erythrocytic membrane of the 15 cows as well as osmotic fragility of the erythrocytes and blood routine. Results: Our results indicated that the content of soil and grass phosphorus was significantly lower (P<0.01) than national phosphorus standard and NRC feed standard in April, and serum phosphorus of the 15 cows decreased obviously. All 15 cows was found to have anaemia in April detected by the method of blood routine. Although the content of phospholipid of erythrocytic membrane decreased, the content of cholesterol increased significantly in April, indicating that the serum phosphorus was positively relative to phospholipid while was negatively relative to cholesterol. The fluidity and ATPase activity of decreased while the osmotic fragility of erythrocytic membrane increased in April. In July and October, levels of phosphorus of grass and soil were lower than standard level while the content of phosphorus in serum of 15 cows had no difference with normal level. These items of erythrocytic membrane were near to those of the negative control. It suggested that phosphorus deficiency took place maily in April instead of in July or October. Conclusions: This study indicated that environmental low phosphorus was the major factor caused endemic hypophosphatemia. Erythrocytic membrane damage induced by environmental low phosphorus may be the crucial mechanism of endemic hypophosphatemia. Key words: dairy cattle, phosphorus, erythrocytic membrane, endemic haemoglobinuria

Nutrition and Metabolic Disorders 41

541 Variability of Blood Profile and Milk Composition in Relation to Stage of Lactation and Season of the Year in Dairy Cows SM. Abdel raheem, C. Iben University of Veterinary Medicine Vienna, Nutrition, Vienna, Austria Objectives of study: The study was designed to determine the seasonal and lactational influences on blood metabolites, milk composition, BCS, and back fat thickness (BFT) in dairy cows. Materials and methods: The study was conducted from April 2006 until February 2007 using (n=35) dairy cows (27 Fleckvieh and 8 Holstein Friesian). Cattle were given 15 kg corn silage, 20 kg grass silage, and 3 kg grass hay as daily basal feed. A concentrate diet was given on the basis of the amount of milk production. Blood samples, Milk constituents, BCS and BFT were investigated every 2 months. Depending on the day postpartum of sampling, data were divided into 3 lactation stages; early (up to 75 days postpartum), mid (76 to 150 days), and late (after 151 days plus dry cows) lactation. Also, depending on the season at the time of sampling; the data were divided into 4 seasons, spring (April), summer (June and August), autumn (October) and winter (December and February). Statistical analyses were conducted with the SPSS; ANOVA was used for lactation stage and season comparison. The interaction between the two factors of the year was done by MANOVA. Results: Lactation stage had a significant influence on both blood metabolites (glucose, BHBA, blood urea nitrogen (BUN), albumen, cholesterol, Mg) and milk components ( milk fat, milk protein milk urea) in addition to BCS and BFT. No significant effect of the lactation stage was detected on total protein (TP), Ca, P, Cu and Zn. At the same time the season of the year has a significant impact on all parameters studied except BHBA, P and BFT. BCS was higher at late lactation (mean±SD) (3.36±0.41) than early (3.23±0.44) and mid (2.91±0.39) lactation. A significant interaction between stage of lactation and season of the year was detected for TP, cholesterol and milk protein. BFT correlated positively with BCS (r=0.84) and also milk urea and BUN (r=0.84) correlated positively. Milk yield and milk protein were correlated negatively(r=-0.61). Both BCS and BFT correlated negatively with cholesterol r= (-0.4). Conclusion: Stage of lactation and season influenced the biochemical parameters and metabolites in blood and milk as well as body condition. Therefore, the interaction between these two factors be taken into account when assessing the nutritional status of dairy cows in different climatic periods and in different stage of lactation. 542 Occurrence of Subclinical Metabolic Disorders and Possibility of the Prevention by Licking Block Supplementation in Beef Cows L. Könyves , V. Jurkovich , A. Tirián , L. Tegzes , O. Szenci , 2 1 Z. Szelényi , E. Brydl 1 Szent István University Faculty of Veterinary Science, Department of Animal Hygene, Herd-health and Veterinary Ethology, Budapest, Hungary 2 Szent István University Faculty of Veterinary Science, Clinic for Large Animals, Budapest, Hungary The aim of the study was to reveal the occurrence of subclinical metabolic disorders in charolais suckler herd in the winter calving period and spring breeding season. The biological effect of an experimental licking block containing protein source, molasses, minerals and vitamins was also evaluated as a possible tool of preventing metabolic disorders. Charolais cows (n=114) were selected randomly into experimental and control group (n=57-57). From these groups 12-12 cows were assigned randomly for metabolic profile tests. The experimental group was supplemented with experimental licking block (voluntary intake about 1.5 kg/cow/day) from 1 month before expected calving until the end of breeding season. Different manifestations of energy imbalance (increased fat mobilisation, hypoglicaemia, hyperketonaemia) and acid load was diagnosed with high occurrence in both groups. Protein-, phosphorous-, sodium- and carotene shortage was diagnosed in the control group. The urea, phosphorous, carotene concentration in blood plasma and sodium concentration in urine were significantly higher in experimental group compared to control. The licking block had no effect on energy metabolism, acid-base balance, Ca, Mg, K, Cu, Zn, Se and Mn supply. There was no difference between groups in mortality- and culling rate, the postnatal calves mortality, metritis and the number of veterinary

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treatments. There was 7.3 % higher conception rate in the experimental group but the detected subclinical metabolic disorders might play a remarkable role in poor reproductive performance in both group. Key words: metabolic disorders, beef cow, supplementation 543 Body Condition Score at Calving and Supplementation with Yeast Affect Blood Mineral and Liver Enzyme Concentrations in Peripheral Blood of Peri-parturient Dairy Cows L. O'Grady, R. Alibrahim, V. Gath, M. Doherty, F. Mulligan University College Dublin, School of Agriculture, Food Science and Veterinary Medicine, Dublin, Ireland The objective of this study was to determine the effect of body condition score (BCS) at calving and supplementation with a live yeast culture 1026 (Yea-Sacc , Saccharomyces cerevisiae, CBS 493.94) beginning two weeks pre-calving on the metabolic status of peri-parturient dairy cows. Forty Holstein/Friesian cows of mixed parity were randomly allocated to two groups depending on BCS approximately 60 days prior to parturition. Both groups were fed grass silage ad-libitum for the entire 60 days precalving and half of each group were supplemented with a dairy compound containing 2.5g of thermo-stable yeast supplement for the last 14 days pre-calving and during the first 70 days of lactation. Blood samples were taken via jugular venipuncture on the day of calving (12-24 hrs post calving) and on days 5, 15, 25 and 35 post-calving and analysed for Ca, Mg, P, gammaglutamate transferase (GGT), glutamate dehydrogenase (GLDH) and bilirubin (BIL). Data was analysed using ANOVA and the Proc GLM statement of SAS separately for each day of sampling as a 2 X 2 factorial with BCS group, supplement group, BCS group x supplement group and parity included in the model. Dairy cows in the high BCS group had a higher (P<0.05) Mg status than cows in the low BCS group on the day of calving (1.07±0.044 Vs 0.91±0.046 mmol/l). The high BCS group also had a lower (P=0.06) Ca status (2.09±0.064 vs 1.92 vs 0.061 mmol/l) and tended to have a lower P (P<0.10) status than cows in the low BCS group (1.44±0.087 vs 1.24±0.084) on the day of calving. Yeast supplementation had no effect (P>0.05) on Ca, Mg or P status on the day of calving. At day 5 post-calving there was a significant interaction of BCS group and supplement group on blood Ca status (P<0.05) with low BCS group cows having a higher Ca status for the yeast supplement group only. Yeast supplemented cows had lower (P<0.05) circulating concentrations of GLDH on the day of calving and 5 days post-calving. However yeast supplemented cows had a higher (P<0.05) circulating concentrations of BIL at day 35 post-calving. High BCS group cows had higher (P<0.05) circulating concentrations of BIL at 5 days post-calving and at 25 days post-calving and tended to have higher circulating BIL concentrations on day 35 post-calving. These data indicate that body condition score at calving and yeast supplementation alter major mineral and liver enzyme concentrations in the peripheral blood or peri-parturient dairy cows. 544 Serum Concentration of Minerals and 25-hydroxyvitamin D in Periparturient Dairy Cows and its Relation to Backfat Thickness G. Salgado , J. Bouda , A. Castillo , J. Doubek , S. Dávila , 4 4 J. Quintero , H. Romero 1 Facultad de Medicina Veterinaria y Zootecnia - UNAM, Patologia, México D.F., Mexico 2 Veterinary and Pharmaceutical University Brno, Physiological Sciences, Brno, Czech Republic 3 Microtec, Analisis Clinicos, México D.F., Mexico 4 Laboratorio de Analisis Clinicas y Patologia Veterinaria, Patologia Clinica, Gomez Palacio Dgo., Mexico The objective of this study was to compare concentration of Ca, P, Mg and 25-hydroxyvitamin D (25-OH D3) and its relation to backfat thickness (BFT) in periparturient primiparous and multiparous dairy cows. The study was realized in a commercial dairy farm (5000 dairy cows, milk yield 8500 kg) in the region of La Laguna Durango, Mexico. We used 30 animals, Group 1 (n=16) multiparous cows, Group 2 (n=14) primiparous cows. Blood samples were collected from tail vein on day 7 to 3 before expected calving date, 6 h postpartum (before colostrum milking and treatment) and 12 h postpartum. Calcium borogluconate was administered to multiparous cows 6 h after calving. Between 7 and 3 days before calving, body condition score (BCS) was evaluated and BFT was measured by ultrasound (Bantam Medical Imaging E.I). The

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42 XXV. Jubilee World Buiatrics Congress 2008

serum concentrations of Ca, P and Mg were determined by Randox kits in semiautomatic biochemical analyzer (Selectra Junior), the concentration of 25-OH D3 by chemiluminescence assay (DiaSorin, LIAISON). The concentrations of Ca, P, Mg and 25-OHD3 were compared by repeated measures ANOVA. BCS and BFT were compared by student´s t-test. Significant difference was found in concentration of Ca, P, Mg and 25-OH D3 with respect to time sampling (p<0.05) and between groups except for P concentration (p<0.05). No difference was found in BCS between groups (p>0.05). BFT was greater in multiparous cows (32.20±5.37 vs 24.64±3.56) (p<0.01). The correlation between BCS and concentrations of Ca, P, Mg and 25-OH D3 was low (r=-0.23, r=-0.12, r=-0.29 y r=0.18, respectively) Better correlations were found between BFT and Ca, P, Mg, 25-OH D3 (r=0.52, r=0.55, r= -0.51 y r= 0.53, respectively). The concentrations of Ca and P were decreased in both groups during the first 12 h postpartum and higher in primiparous cows before parturition. The concentration of Mg was increased in both groups 6 and 12 h postpartum, and was higher at 7 d in primiparous cows. The concentration of 25-OH D3 increased in both groups at 6 h and decreased 12 h postpartum, but was higher in multiparous cows. There was poor relationship between BFT and serum Ca, P, Mg and 25-OH D3. Key words: hypocalcemia, dairy cows, 25-OH D3 545 Relationship between Body Condition and Metabolic Parameters in Dairy Cows according to Different Days in Milk Z. Gergácz , E. Báder , E. Szûcs , A. Kovács , L. Könyves , 1 E. Brydl 1 University of Western Hungary Faculty of Food and Agriculture Science, Department of Cattle and Sheep Breeding, Mosonmagyaróvár, Hungary 2 Szent István University Faculty of Veterinary Science, Department of Animal Hygiene, Herd Health and Veterinary Ethology, Budapest, Hungary The aim of the study was to analyse the relationship between the body condition and the results of metabolic profile test have been done in dairy cows in different days in milk (DIM). Blood and urine samples were taken from 1984 clinically healthy cows, selected randomly from various groups of cows with different physiological stage of lactation and gestation, 3-5 hours after the morning feeding. The groups differed in respect of daily milk yield, stage of lactation and gestation. During the samplings body condition scoring (BCS) was done as well. The cows were kept at 49 large scale dairy farm located in different part of Hungary. In the present study the following groups of cows were tested: cows 12 days prior to expected parturition, cows average DIM 3, 18, 44, 76, 104, 133 and 218 with the n=373, 270, 566, 287, 185, 153, 42 and 108, respectively. The results of the examination can be summarised as st following: The BCS decreased from the 1 day of lactation (3.48) th onwards till the 44 day (2.65) and slightly increased till the day 216 (2.89). The haemoglobin value and the glucose concentration in blood samples were ranging within the physiological range and followed the tendency of BCS changing at P<0,001, P<0,01, respectively. There was a close negative correlation between the NEFA concentration in blood samples and the BCS changing (P<0,01). The aceto-acetic acid concentration exceeded the upper limit of the physiological range indicating hyperketonaemia at DIM 18. The AST activity value exceeded the upper limit of physiological range and followed the tendency of BCS changing (P<0.01). The urea concentration in the blood exceeded the upper limit of the physiological range in all cows. The NABE value in the urine samples indicated acid load in all groups sampled. According to the result of cluster analysis relationship was found between the BCS, days in milk, glucose, aceto-acetic acid and NEFA concentration in the blood. The results of the study confirm that the body condition scoring is a reliable tool for revealing the risk of metabolic disorders caused by malnutrition. Key words: body condition, days in milk, haemoglobin, glucose, NEFA, aceto-acetic acid, AST, blood urea, NABE 546 Selenium Blood Levels and Lymphocyte Response to Mitogenic Treatment in Cows and their Calves Supplemented with Organic and Anorganic Forms of Selenium P. Slavik , J. Sichtar , M. Brix , J. Illek , R. Rajmon , F. Jilek 1 Czech University of Life Science Prague, Department of Veterinary Sciences, Prague, Czech Republic

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2

University of Veterinary and Pharmaceutical Sciences Brno, Clinic of Ruminant Diseases, Brno, Czech Republic

The objective of our study was to determine, whether supplementations with different sources of selenium can influence its level in the blood and the mitotic activity of lymphocytes. 60 Charolais cows in late pregnancy were divided into three groups. The same feed ration of hay, silage, and premix was given to all animals ad libitum. In the two supplemented groups, the premix was enriched with an organic source of selenium (group "O")-selenised yeast (Sel-Plex50, Alltech, Nicholasville KY) or with inorganic form (group "I")-sodium selenite, 50mg of Se/kg of premix. The control group ("C") received the basic premix without any selenium supplement. The average intake of selenium in both supplemented groups was 3.5 mg Se/head and day. In week 6 of supplementation, blood samples were taken from 10 cows in each group which were just before calving. These cows gave birth 1 or 2 weeks later, and their calves were sampled at the age of 5 days. The remainders of the cows were sampled in week 12 with similar subsequent labour and the sampling of their calves. Selenium blood levels were determined, and transformation assay was performed in the entire blood culture. Non-specific mitogens - phytohaemagglutinin, concanavalin A, and pokeweed mitogen were used. The incorporation 3 of H-thymidine was measured with a microplate scintillation and luminescence counter. Lymphocyte mitotic activity was determined as the number of signals per 1000 lymphocytes for each mitogenic treatment. ANOVA and MANOVA tests were used for statistical evaluation. The "O" animals showed higher selenium blood levels at both sampling times compared to the other groups (P < 0.01). The "I" animals (cows as well as calves) increased their selenium blood levels st nd between the 1 and the 2 sampling (P < 0.01). Such a difference was not observed in the "C" or "O" groups. Mitotic activity differed in general according to the mitogenic treatment used. "C" calves showed higher activity than their mother s (P < 0.05). At least some types of mitogenic treatment induced significantly higher lymphocyte response in "O" and "I" animals as compared to the "C" ones. Similarly, some differences were observed between the "O" and "I" groups. The activity was generally higher in animals with a higher selenium blood level (P < 0.05). This work was supported by a grant No. QF 4005 and 1G46086 547 Milk Fat and Protein Contents as Additional Indicators for Prediction of the Onset of Cyclic Ovarian Activity in Dairy Cows Post Partum J. Sichtar, P. Slavik, R. Rajmon, L. Kourimska, P. Klabanova, F. Jilek Czech University of Life Science Prague, Department of Veterinary Sciences, Prague, Czech Republic The metabolic status of dairy cows influences their reproductive system as well as their milk production. The objective of this study was to verify possible relations between certain milk parameters and the reinitiating of cyclic ovarian activity in high producing dairy cows post partum (p.p.). Twenty-four Holstein cows with an average yield of 8 700 l per lactation were monitored from their giving birth th to their 60 day post partum. The animals were kept in the same stable and fed with the same feed ratio. They showed no clinical signs of any illness. The milk was sampled with an auto sampler during the afternoon milking every other day from day 6 p.p. Fat and protein contents of the milk were determined (Milcoscan, Foss Electric). Simultaneously, the ovarian activity was monitored by ultrasound examination. The first ovulation appeared before day 22 p.p. in 8 cows (i.e. 33 %), between days 22 and 60 p.p. in 10 cows (42 %) and 6 cows (25%) showed no ovulation before day 60 p.p. The milk protein generally tended to decrease temporarily, whereas the milk fat usually increased first, then dropped for some period, as well. Therefore, the fat-protein difference increased first and decreased again later. During the period between days 11 and 22 p.p., the milk protein was lower and fat as well as the fat-protein difference higher in animals, which did not ovulate before day 60 p.p., compared to both remaining groups. The differences were highly significant. The differences between animals ovulating before day 22 p.p. and those ovulating between days 22 to 60 p.p. were insignificant but showed a similar trend. When these two groups were compared on the basis of the last two milk samples before day 22 p.p., the Multiple ANOVA test confirmed the significance of the difference just above the critical level (p=0,058). When milk composition just before the first post partal ovulation (i.e. the last 2 samples) in individual cows was

Nutrition and Metabolic Disorders 43

evaluated, highly significantly greater milk protein was detected in those animals ovulating before day 22 p.p. Our study demonstrates that these indicators could be useful in estimation of the post partal onset of ovarian activity in dairy cows. A larger complex of metabolic status indicators in milk will be the subject of our subsequent studies. This work was supported by a grant No. MSM 6046070901 and 1G46086 548 Effect of Organic and Inorganic Zinc Supplementation on the Prevention of Zinc Deficiency in Aberdeen Angus Calves S. Dresler, L. Perlikova, D. Kumprechtova, J. Illek University of Veterinary and Pharmaceutical Sciences Brno, Brno, Czech Republic Objective: The objective of the study was to investigate the effect of supplementation of suckling Aberdeen Angus (AA) calves with different forms of zinc during the grazing season. Material and methods: At the beginning of the grazing season, zinc deficient was diagnosed in AA calves (n=16) at 2 to 5 weeks of age along with their mothers, into two groups (Control and Bioplex). The calves suckled milk, were given starter feed and gradually were getting used to graze. The calf starter feed consisted of crimped wheat, oats, barley and protein concentrate. The starter feed contained 36 mg zinc per 1 kg and was given ad libitum to calves from 1 week of age. After the beginning of grazing season, the zinc concentration in starter

feed was increased by 30 mg per kg (total 60 mg Zn/kg). The Control calves received starter feed with zinc sulphate, the Bioplex calves received calf starter with organic zinc (Bioplex-Zn). Calves in both groups received starter feed ad libitum. The experiment lasted 2 months. At the beginning and the end of the trial the calves were weighed and each calf was withdrawn a blood sample to measure zinc concentration and selected parameters of metabolic profile. The results were statistically analysed and evaluated. Results: Initial serum zinc concentration was 6.55±0.80 and 7.10±0.6 umol/l, serum total protein levels were 65.2 l and 66.5 g/l, and serum IgG levels were 14.8 g/l and 13.1 g/l in the Controls and Bioplex, respectively. At the end of the trial there was a significant increse in serum zinc concentrations in both groups, with Bioplex supplemented calves showing a higher increase in plasma zinc levels. Zinc concentrations were 11.87±1.77 and 15.81±0.72 µmol/l in the Control and Bioplex groups, respectively. The differences were statistically significant. The Bioplex supplemented calves showed higher IgG levels. There were significant differences in daily weight gain between the groups, too. While the Control calves showed the growth rate of 1.12±0.07 kg per calf per day, the Bioplex calves grew 1.24±0.09 per calf per day. Conclusions: The dietary zinc supplementation in calves favourably influenced serum zinc concentrations. The organic zinc supplementation resulted in a greater increase in serum zinc concentrations, serum IgG levels and weight gain. The organic form of zinc can be recommended as a useful tool to prevent zinc deficiency in calves.

Mastitis / Udder Health and Milk Quality

7 Health Status of Cows Before Parturition Associated with Concentration of GM1 & IgG in Colostrum and Reflects Resistance of Acquiring New IMI at Early Lactation G. Leitner1, O. Krifucks1, S. Jacoby2, Y. Lavi3, N. Silanikove2 1 Kimron Veterinary Institute, Bacteriology, Bet Dagan, Israel 2 Agricultural Research Ortganization, the Volcani Center, Animal Science, Bet Dagan, Israel 3 Hebrew University, Rehovot, Israel Objective: Study the association between the concentrations of IgG and Gangliosides type M1 (GM1) in colostrum of cows and IMI during first weeks postpartum. Materials and methods: Israeli-Holstein cows n=108: 38 heifers and 75 pluriparous cows. Cows were dried 60 d before next expected parturition. During the 45 d prior to the end of lactation, the cows were tested 3 times on the quarter level for bacteriology and SCC. Colostrum of cows and heifers were tested on the day of parturition and thereafter milk was tested at 10, 30 and 45 d postpartum for bacteriology and SCC. Bacterial identification and SCC were carried out according to IDF. Total IgG & GM1 (Relative values of GM1, GM1-RV) were determined as the reduction of binding of CT to PolySorb plates coated with GM1 by competition with the b-chain of cholera toxin (b-CT) were determined in the mixed colostrum. The effects of udder infection and lactation (heifers vs pluriparous) on total IgG, GM1-RV & SCC were determined by a two-way ANOVA in a random design. Linear discrimination analysis was used to distinguish between the classification variable (uninfected or infected) based on total IgG and GM1-RV levels and records of infection. Results: High levels of IgG & GM1-RV in pluriparous cows were associated with lower new incidence of IMI in the subsequent lactation. The mean IgG level in colostrum of heifers was not significantly lower than that of pluriparous cows. Infected quarters of heifers or cows had significantly lower IgG levels than uninfected quarters. The mean GM1-RV in the colostrum of infected heifers was significantly higher than that of the pluriparous cows. This difference was not found in uninfected animals. Moreover, in heifers, GM1-RV in infected quarters was not significantly higher than in uninfected ones whereas in the pluriparous cows, GM1-RV levels were significantly lower in the infected than in the uninfected quarters. Linear discriminate analysis based on these two variables yielded 70% successful classification. This analysis was less successful in heifers because a high proportion of them joined the herd already infected with bacteria. Conclusions: The results suggest that the health status of the animal at the last month before parturition is associated with colostrum quality and reflects its resistance of acquiring new IMI. While in heifers, of which high proportion joined the herd when already infected with bacteria no such association was found. 8 Cell Differentiation in Milk of Cows with Healthy Udders D. Schwarz1, K. Failing2, W. Wolter3, CP. Czerny1 1 Georg-August-Universität Göttingen, Institute of Veterinary Medicine, Göttingen, Germany 2 Justus-Liebig-Universität Gießen, Unit for Biomathematics and Data Processing, Gießen, Germany 3 State Council Gießen, Milk Control, Wetzlar, Germany Cell counts and the determination of differential cell patterns in milk samples are important criterions for the evaluation of udder health. The limits for the evaluation of udder health are well-defined by clear cut-off rates. The objective of our study was the analysis of somatic cell populations in foremilk samples to identify inflammations at a significantly lower level than given by current definitions. The milk of Holstein-Friesian cows (n=18) and German Simmental cows (n=2) with low cell counts from five Hessian dairy farms was examined cytologically. Cell counts were determined with a Fossomatic 5000 and bacteriological examinations. Milk smears were generated for each udder quarter and stained according to the method of Pappenheim. To create the differential cell patterns, 100 cells per milk smear were counted and differentiated by microscopy. The classification into different cell count groups of all foremilk samples analyzed in the years 2000-2003 in the German federal state Hesse showed the importance of very low (7%) and low (50%) cell counts. Moreover, the results of monthly cell counts developed on quarter levels allowed to suggest inflammable reactions in the cell count array lower than 100,000 cells/ml, because there were clear cell count variations visible in a little time period. All 80 udder quarters tested in our study were subdivided into groups with very low, low and medium cell counts. In each group we found epithelial cells at the differential cell patterns of a maximum of 3%. The lymphocytes dominated the `very low' group with an amount of 70%. The differential cell patterns in the `low' group consisted of ca. 40% lymphocytes and 40% macrophages. The percentage of the polymorphonuclear neutrophils clearly increased at cell counts of 43.000 cells/ml. These data indicated inflammable reactions at such low cell counts. Our findings were confirmed by monitoring the development of the monthly cell counts. The bacteriological examinations only identified S. aureus in one quarter, E.

44 XXV. Jubilee World Buiatrics Congress 2008

coli in another quarter and Coagulase-negative staphylococci in two other cases. The statistical calculations proved a significant correlation between cell counts and differential relationships of the four analysed cell types to each other. On the basis of the results of our study we detected inflammable reactions in the mammary glands of obviously healthy udders already at a level of 43.000 cells/ml. Key words: differential cell count, low somatic cell count milk, differential cell pattern 9 Understanding Mastitis Epidemiology on Farm - Determining the Importance of the Dry Period in Mastitis Control A. Bradley , M. Green 1 University of Bristol, School of Veterinary Science, Bristol, UK 2 Quality Milk Management Services Ltd., Wells, United Kingdom 3 University of Nottingham, Department of Veterinary Medicine, Nottingham, United Kingdom 4 Orchard Veterinary Group, Glastonbury, United Kingdom Objectives: Appropriately targeting mastitis controls on farm requires the clinician to determine the relative importance of the dry and lactating periods. The aim of this study was to investigate novel indices for clinical and sub-clinical mastitis, previously described by the authors, to categorise farms by the importance of the dry and lactating periods. Materials and Methods: Using somatic cell counts (SCC) the importance of the dry period was investigated by calculating; the Fresh Calver Infection Rate (FCIR - proportion of cows with SCCs >200,000cells/ml at the first recording post calving), the New Dry Period Infection Rate (DPNEW - proportion of cows with SCCs rising from < to > 200,000 cells/ml across the dry period), the Dry Period Cure Rate (DPCURE proportion of cows with SCCs falling from > to <200,000 cells/ml across the dry period). Using clinical mastitis data, the proportion of the herd affected (IRCA) was calculated and cases were attributed either to the dry period or lactation on the basis of the timing of an "index" case (1st case in lactation) in an individual cow. Apparent Dry and Lactating Period Origin Clinical Mastitis Rates (DPORATE and LPORATE respectively) were then calculated, namely the 1st 30 day incidence rate (target: <1 in 12 cows affected) and the incidence rate for the remainder of lactation (target: <2 in 12 cows affected). Finally for each farm the relative contribution of the dry period (DPCONT) was calculated. Results: Data is presented from 52 herds. Mean annual rates were calculated for each farm. These demonstrate significant variation in the indices between farms and within farms over time. The mean, median, min, max and target respectively, for each index is outlined below. FCIR: 0.23, 0.22. 0.10, 0.43, 0.10. DPNEW: 0.19, 0.19, 0.06, 0.42, 0.10. DPCURE: 0.71, 0.70, 0.51, 0.91, 0.85. ICRA: 0.44, 0.40, 0.14, 0.79, 0.25. DPORATE: 1.79, 1.74, 0.12, 4.94, 1.00. LPORATE: 3.00, 2.88, 0.97, 6.09, 2.00. DPCONT: 0.37, 0.35, 0.07, 0.75, 0.33. Conclusion: The indices outlined in this study allow farms to be categorised according to the likely importance of the dry and lactating periods in mastitis control, thereby allowing the practitioner to select mastitis interventions likely to have the largest impact on farm. Continued monitoring allows 10 Mammary Gland Immune Dysfunction in High Yielding Dairy Cows Immediately after Calving: can Dietary Vitamin C help? J. Mehrzad , M. Talebkhan Garoussi , AH. Falah Rad , 2 1 N. Farzaneh ,G. Hashemitabar 1 Ferdowsi University of Mashhad, Faculty of Veterinary, Department of Pathobiology, Mashhad, Iran 2 Ferdowsi University of Mashhad, Faculty of Veterinary, Clinical Science, Mashhad, Iran Neutrophils are the most vital part of the first line of immune defense against invading mastitis pathogens in the mammary gland of high yielding dairy cows. Their quality and quantity in the gland is important. Vitamin C is one of the most important water-soluble protective agents in mammalian cells. Although cows can synthesize vitamin C, substantial evidence suggests a link between vitamin C and immunity. We hypothesized that feeding vitamin C in dairy cows would improve milk neutrophil function via its antioxidant properties. This study was, therefore, aimed to determine whether supplemental dietary vitamin C affects on milk neutrophil quality and quantity immediately after calving. We fed Holstein cows supplemental vitamin C during the peri-partum period; at 10 d before anticipated calving, the cows were fed diets that provided 0 (n = 7) or 50 (n = 7) gm/d of supplemental vitamin C

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(phosphorylated ascorbic acid). Treatments continued until approximately 15 days in milk. The viability of milk neutrophils and somatic cells were examined after calving. Compared with non-vitamin C supplemented group, feeding vitamin C during periparturient period resulted in significant-increased neutrophils viability and decreased somatic cells in the mammary gland after calving. The reduced somatic cells in the gland from vitamin C treated cows suggest that it may reduce damage to the gland, milk and neutrophils caused by parturition-andlactation-related stress. In our previous experiments we observed that the unnecessarily high number of neutrophils and excessive somatic cells in the milk may relate to udder and milk damage as free radicals and proteases released by neutrophils appear to be actively involved in the damage. Therefore, in the current study limited, but fairly enough, SCC with high quality of neutrophils observed in cows receiving vitamin C supplementation implies a more efficient neutrophil response compared with the response in non-supplemented cows. Overall, supplemental dietary vitamin C during the peri-partum period improves neutrophil function; this potentially results in faster elimination of pathogens in the gland during mastitis. Though still remains inconclusive, the application of vitamin C would be further examined in dairy cows for prevention and treatment of clinical mastitis; because it boosts milk neutrophil viability. Key words: mastitis, milk, neutrophils, vitamin C 41 Using Polymerase Chain Reaction Techniques in the Diagnosis of Streptococcus Agalactiae P. Mansell, R. Agnew, G. Browning University of Melbourne, Veterinary Science Department, Melbourne, Australia Objectives of study: Although Streptococcus agalactiae is commonly believed to be of decreasing importance in many dairy regions, infection with this pathogen is still suspected as being a cause of mastitis in many herds. Infection with S agalactiae has serious implications for milk quality. Traditional methods of detecting S agalactiae in herds have used bacteriological culture and phenotypic identification of isolates. These methods are time consuming and have practical limitations. PCR methods offer substantially improved sensitivity over culture. PCR methods also offer considerable technical advantages when used for testing bulk tank samples. The aim of this study was to develop a PCR methodology to detect S agalactiae in vat milk samples. Materials and Methods: Vat milk samples or individual cow milk samples were obtained from 26 Australian dairy herds, either directly from the herd owner or via the veterinarian advising the herd. Herds were selected because they had had recent problems with elevated bulk milk somatic cell counts, or increased numbers of subclinical or clinical mastitis cases. Submitted milk samples were incubated in Todd Hewitt broth overnight and then processed to extract the bacterial DNA. This DNA was then amplified by PCR using S agalactiae specific primers spanning the 16S to 23S rRNA spacer region. These were subsequently separated and identified in polyacrylamide gels. Results: Nine of 24 bulk tank samples were positive for S agalactiae. For 7 herds, both bulk tank samples and individual cow samples were available. In only one of these herds did the results of the testing of the individual cow samples disagree with the result of the bulk tank test. Conclusions: These early results suggest that, contrary to common thought, S agalactiae continues to be an important mastitis pathogen in many herds. This pathogen represents a significant threat to efforts to control mastitis and to improve the quality of raw product supplied to processors. New methods such as PCR have the potential to offer an efficient method by which herds can be monitored for the presence of S agalactiae using routinely collected bulk tank samples. This allows early identification of herds at risk of having high bulk milk somatic cell counts. 42 Analytical Sensitivity of the PathoProof Mastitis PCR Assay Determined Using Two Different Experimental Approaches M. Koskinen,J. Holopainen,J. Kurkela Finnzymes Oy, Diagnostics, Espoo, Finland The PathoProof Mastitis PCR Assay is a novel polymerase chain reaction-based reagent kit for identification of all major mastitis-causing pathogens. Here, we report on the analytical sensitivity of the assay using two different experimental approaches. We first used purified DNA to prepare a standard dilution series from 50 to less than one genome copy of the assay's bacterial targets and determined how many copies of each target could be detected with 95% probability. In our second experiment,

Mastitis / Udder Health and Milk Quality 45

we measured sensitivity of the entire assay, including the DNA extraction procedure. For this experiment, ten mastitis milk samples shown to be negative for the tested targets were randomly chosen, spiked with known quantities of the target bacterial colony forming units (CFU) and analyzed using the PCR assay. When sensitivity was measured based on the dilution series of purified DNA, the analytical detection limit of the test was as low as 5.6 bacterial genomes per l. When using mastitis milk samples spiked with the bacterial targets, the detection limit was as low as 100 CFUs per one millilitre of milk. Both experiments showed variation in the sensitivity of the test between the different bacterial targets. In conclusion, the results of this study demonstrate that the PathoProof Mastitis PCR Assay can detect low quantities of bacterial DNA. This presentation also discusses some of the challenges involved in sensitivity validation of a PCR test in a controlled manner when using raw bovine milk as a sample matrix. Sensitivity of the PathoProof Mastitis PCR Assay should next be validated against the golden standard method, i.e. bacterial culture. 43 Investigating the Use of Viscosity Measurements for the Quantification of the California Mastitis Test (CMT) in Dairy Cattle J. Roberts University of Cambridge, Department for Veterinary Medicine, Cambridge, United Kingdom The California Mastitis Test (CMT) is widely used as a qualitative cow side test for the detection of subclinical mastitis in dairy cattle. The viscosity of the milk-CMT gel reaction has been characterised but at present no attempt made to determine whether the reaction can assess quantitatively the somatic cell count (SCC) in milk samples. This initial investigation looks at the milk and milk-CMT gel reaction viscosity for various SCC quarters and characterises the sample viscosity for these SCCs. 57 quarter samples were collected aseptically from 16 Holstein-Friesian cows in a commercial dairy herd within the UK. Viscosity measurements were made using a Brookfield R/S Rheometer at constant shear rate for milk alone and milk with CMT reagent. Each quarter SCC was measured at a commercial approved laboratory. The correlations between average viscosity, initial viscosity, final viscosity, and change in viscosity for both milk and milk-plus-CMT reagent against SCC are demonstrated. The removal of the milk viscosity from the milk-plus-CMT reagent viscosity may allow quantification of the SCC between animals. The milk-CMT gel reaction used qualitatively to demonstrate subclinical infection may be able to be measured quantitatively if the process of analysing viscosity is carefully controlled as shown by this initial analysis. Further work will investigate the CMT reaction at various constant shear rates and using controlled shear rates. It is anticipated that quantification of the viscosity of the CMT test may allow the development of a portable device for more accurate determination of SCC cow side and be developed for an in-line device in automated milking systems. 44 Prevalence of Intramammary Infections in Heifers around Calving in 40 Dairy Herds in the West of France. L. Durel ,C. Guidarini , P. Moroni , C. Locatelli , L. Scaccabarozzi , 4 E. Schmitt-Van de Leemput 1 Clinique Vétérinaire de Marigny, Marigny, France 2 Boehringer Ingelheim France, Technical & Development Department, Reims, France 3 Facolty di Medicina Veterinaria di Milano, Patologia animale, Igiene e Sanit'Pubblica Veterinaria, Milano, Italy 4 Clinique Vétérinaire de Villaines-la-Juhel, Villaines-la-Juhel, France Intramammary infections (IMI) in dairy heifers around calving are responsible for important economical losses. A better understanding of the prevalence, the responsible germs and the risk factors of such infections is needed to improve heifers udder health status in France. To investigate the prevalence and the risk factors of IMI in heifers, an epidemiological study has been carried out in two veterinary practices in the west of France from June to December 2007. In this abstract, the bacteriological data are reported. Sterile milk samples of the 4 individual quarters (n=1.496) were collected of dairy heifers (n=378) between 3 and 7 days post partum from 35 dairy herds and frozen until analyses. For each sample, somatic cell count was

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determinate and bacteriological analyses were performed according to the recommendations of the NMC and the IDF. 1% of the quarters (n=16) were not functional at calving time. In total, 22.3% (n=333) of the milk samples was positive in bacteriological analyses, 77.3% (n=1.156) was negative; 2.1% of the samples was contaminated. In general there was no clear relationship between elevated cell count (> 100.000 cells/ml) and positive result in bacteriological analyses. However, when the data were analyzed per type of bacteria, it showed that the presence of Streptococcus spp. (Streptococcus uberis) coincided in 77% (85%) of the cases with an elevated cell count, whereas the presence of Staphylococcus spp in 48% (S. aureus) and 64% (CNS). Among infected heifers (n=214), the number of animals with 1, 2, 3 or 4 quarters infected, were respectively, 63.0%, 22.4%, 10.3% and 4.2%. 43.4% (n=164) of the heifers didn't have any quarter infected. The frequency of presence of CNS, Staphylococcus aureus, Streptococcus spp (Streptococcus uberis) and Gram negative spp. were respectively 64.2%, 8.4%, 16.2% (9.3%) and 4%. On the herd level, the lowest prevalence of infected animals was 20% and the highest 100%, but for most of the farms (n=26) the infection rate varied from 45 to 75 %. In conclusion, IMI in heifers has a high prevalence in France and most of the herds have numerous infected animals at calving. Further analyses of the data are needed to provide more information on the economical impact and the risk factors of IMI infections of heifers. 45 Bovine Mastitis: Genotypes of Staphylococcus aureus Differ in Their Clinical Outcome after Intramammary Infection HU. Graber Clinic for Ruminants, Vetsuisse Faculty, University of Bern, Berne, Switzerland Objectives of study: Based on our clinical experience on bovine mastitis, we hypothesized that there are subtypes of Staphylococcus aureus (S. aureus) which differ in their contagious and pathogenic properties after intramammary infection (IMI). Materials and Methods: 210 strains of S. aureus were isolated from bovine milk samples collected during a representative epidemiological study on S. aureus in cow herds of Switzerland. In each of the 26 herds investigated, all lactating cows (n=449) were evaluated clinically for udder health. Milk of each quarter was then aseptically collected for bacteriological testing and analysis of somatic cell counts (SCC). The genotypes of S. aureus were determined by PCR amplification of the 16S-23S rRNA intergenic spacer (RS-PCR). Results: In total, 17 genotypes were obtained. Two of them (type B and C) were predominant (81%). Each of the remaining types (OG) occurred at a very low frequency. Considering genotype B, IMI was always present in many cows of a herd. Frequently more than 1 quarter per cow was infected. In the case of genotypes C and OG, IMI was restricted to 1 or very few cows per herd. Constantly, not more than 1 quarter per cow was infected. Cow prevalence of S. aureus was highly associated with the genotypes (p<0.001). There was a large difference between the median cow prevalences of genotypes B (median = 47.2%) and C (median = 7.1%; p<0.001) and between B and OG (median = 6.3; p<0.001). No difference, however, was observed between genotypes C and OG. On quarter level, the results were very similar. Analysis of SCC for IMI caused by monoinfection with S. aureus revealed higher mean counts for genotype B (meangeo = 7.85 x 105 cells/ml) than C (meangeo = 3.48 x 105 cells/ml; p = 0.025). There was no difference of SCC between other genotype combinations. Conclusions: The results demonstrate that there are subtypes of S. aureus which clearly differ in their clinical outcome after IMI. The corresponding genotypes are highly associated with the epidemiological and pathogenic properties of this pathogen. In particular, genotype B is associated with high contagiosity and increased pathogenicity, whereas the other genotypes (C, OG) cause sporadic infections only. Genotyping by RS-PCR represents an excellent tool and generates clinically relevant results. 76 Milk Leukocyte Population Patterns in Relation to Milk Fraction, Lactation Stage and Udder Health B. Bansal , J. Hamann , N. Grabowski , F. Reinecke , G. Klein 1 Guru Angad Dev. Veterinary and Animal Sciences University, Clinical Veterinary Medicine, Ludhiana, India

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46 XXV. Jubilee World Buiatrics Congress 2008

2

University of Veterinary Medicine Hanover, Foundation, Institute of Food Quality and Food Safety, Hanover, Germany

Total cell count in milk is used as an indicator of udder health status, with each cell type having its own specific function in the immune response. The aim of this study was to explore the bovine mammary gland defence mechanisms by determining the cellular composition of milk in relation to lactation stage, milk fractions and udder health. Total cell count (SCC) and differential cell count (DCC) of neutrophils (PMNL), lymphocytes (LY), and macrophages (MA) were determined on quarter foremilk and stripping samples of 24 dairy cows (08 each in early, mid and late lactation). SCC was measured by Fossomatic method. DCC were made on cytocentrifuge preparations of cells stained with toluidine blue (2 mg/l). Culturing of samples followed the procedure of National Mastitis Council. Udder health status was defined according to International Dairy Federation. Three grouping variables were considered in statistical analysis; milk fraction, stage of lactation, and udder health. Results revealed 10%, 5% and 21% of the quarters suffering from specific, non-specific and latent mastitis, respectively. Udder health and milk fractions showed significant effect on cell counts. Even the latent infections caused by Corynebacterium and CNS resulted in a significant increase in SCC. Thus, importance of these so-called minor pathogens in defining udder defence mechanisms may not be ignored. Investigation of interdependence of udder quarters indicated significant differences for SCC between healthy and diseased quarters of identical udders. Moreover, SCC for healthy quarters from healthy udders differed significantly from healthy quarters of udders having one or more diseased quarters, thus putting a question mark on the independence of quarters. SCC increased towards stripping, and this increase was more pronounced in diseased quarters. Proportions of PMNL, LY and MA did not change between foremilk and stripping, neither in healthy nor in mastitis quarters. Udder health showed significant affects on DCC; LY were highest in healthy quarters while PMNL in mastitis quarters. Over the lactation, in healthy quarters, PMNL and MA proportions increased, and LY decreased. While in mastitis quarters, cell proportions remained unchanged. Further, as indicated by DCC vis-a-vis SCC, significant pathological/ immunolo3 gical changes seem to occur at 100x10 cells/ml. Thus, probably, the physiological limit for SCC of healthy udders in bovines may be 3 defined even less than 100x10 cells/ml. 77 Evolution of CMSCC in Intramammary Staphylococcus aureus Infected Cows after Calving and Treated at Dry Period C. Ribeiro , J. Galí , D. Castro , P. Teixeira , A. Casas , 3 3 3 B. Fernández , E. Barrio , J. Simoes 1 Centro Veterinário de Aveiro, Aveiro, Portugal 2 Grup Tecnic Veterinari S.L., Gerona, Spain 3 University of Trás-os-Montes e Alto Douro, Veterinary Science Department, Vila Real, Portugal The aim of this work was to characterise the evolution of somatic cell count of mastitic cows infected with Staphylococcus aureus treated only at dry period. Composite milk somatic cell counts Somatic cell count (CMSCC, fossmatic) and microbiologic analysis (Plate Count Agar) were performed, at same time, in all 134 lactating cows from one dairy farm. All cows diagnosed with St. aureus infection were managed separately and only treated with betalactamic antibiotics at the onset of dry period. The CMSCC of first and second dairy report after calving was also used. Linear score (LS), bulk contribution (SCC %) of each animal and mean linear score (mLS; average of three last individual LS) were evaluated for each sample of composite milk. The LS was 3.4±0.2 (S.E.M., n=73) and 1.9± 0.2 (n=61, P<0.001) for positive and negative samples, respectively. The correlation between LS and bulk contribution (%) were r=0.93 (n=134, P<0.001). The St. aureus prevalence was 26.9% (36/134; 49.3% of total positive samples). The bulk contribution (%) by cows with detected St. aureus was 38%. The mean LS of single St. aureus isolated samples (mLS=3.3 ± 0.3, n=32) was not different than others microbiological positive samples (mLS=3.4 ± 0.3, n=41, P>0.05) and both were higher than negative sampled cows (mLS=2.0 ± 0.2, n=61, P<0.001). The mean lactation for cows without microorganisms detection (1.9 + 0.2) was lower (P<0.001) than cows with St. aureus (2.6 ± 0.3) or with others microorganisms (2.7 ± 0.3) isolation. The LS and mean LS at microbiologic diagnosis time (LS=3.7 ± 0.4 and mLS=3.5 ± 0.3) was higher than LS at first (LS=1.7 ± 0.4 and mLS=1.7 ± 0.3) and second (LS=1.9 ± 0.4 and mLS=1.7 ± 0.3, n=29, P<0.001) dairy report after calving. Three old cows remained with mean LS 4 in these first and

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second periods. In conclusion, a decrease of LS and mean LS from St. aureus mastitic cows was observed on next calving after a separated management and dry cow therapy. Key words: mastitis, St. aureus, somatic cells, dairy cattle 78 Udder Health and New Disease Rates in Holstein-Friesian Cows in Relation to their Lactation Number N. Grabowski, J. Hamann, G. Klein University of Veterinary Medicine Hanover, Foundation, Institute of Food Quality and Food Safety, Hannover, Germany Mammary secretion impairments (from mere colonisation to intramammary infection) mean marked economical losses. The epidemiological situation is known to differ between regions, and this study seeks to add data on the new disease rates (NDR) of mastitis in Northern Germany. A total of 63 HF cows (n = 252 quarters; lactation numbers [LN] 1 - 4) were sampled cyto-bacteriologically (IDF recommendation; quarter level) during the entire lactation (8 cows: 2 lactations). Sampling pattern included weekly (8x after calving & 3x before dry-off) and monthly (mid & late lactation) surveys. Udder healthstatus was established for every sampling day categorising culturenegative quarters with < 100,000 somatic cells/ml as normally-secreting, otherwise impaired. Once secretion changed from normal to impaired, the quarter was considered diseased permanently. In all LN, 3 major increases in NDR occurred: the first half of quarters fell ill by day 20, 60, 40 and 55 after calving (lactation 1, 2, 3, 4). Further, yet less marked increases in NDR occurred by week 16 to 25 and during the last two months, especially before dry-off. The amount of normally-secreting quarters at the end of the lactation dropped from 21 (LN1) to 3 % (LN4). Differences among LN were not significant. However, cumulative frequencies (CF) of diseased quarters showed that primiparous cows were affected earlier than older ones. These 3 increases are thought be to be due mainly to the negative energy balance, the endocrinological changes associated with gestation and the suspension of lactation with the changes in mammary tissues. Microbiology analysis translated into a pattern similar to the one observed for all secretion impairments. No significant differences between LN existed, although as a tendency, fewer quarters were colonised towards the end of lactation in LN 1 and LN2 than in the other ones. Coagulase-negative staphylococci (CNS) were more frequent in LN1 than in older ones, while the opposite was true for Staphylococcus aureus. Other pathogens were too infrequent. CF showed that infections with S. aureus occurred earlier in lactation than the ones with CNS did. The results confirm that the early lactation is a crucial phase in mastitis epidemiology from the first week on. However, the new disease rates also increase during lactation by another 30 to 40 %, challenging the practitioner once more. Key words: mastitis, epidemiology, lactation number, new disease rate 79 Thermography of the Udder Skin in Healthy Cows and after Experimentally Induced Escherichia coli-infection M. Metzner, A. Glas, C. Sauter-Louis, W. Petzl, D. Mehne, H. Zerbe, W. Klee University of Munich, Clinic for ruminants, Munich, Germany Objectives of the study: The aim of this study was to evaluate the course of thermographic patterns of the udder skin after experimentally infection with Escherichia coli (E. coli). Materials and Methods: In 5 healthy cows (Holstein-Frisian) thermographic pictures were taken every 2 hours during a period of 48 hours. 24 hours after beginning of the trial the right hind quarters were infected by inoculation of 2 ml of a suspension of E. coli in physiological saline (250 CFU/ml) (infection group). Another 16 healthy cows were used for evaluation of precision, diurnal cycles and influence of ambient temperature during an observational period of 24 hours (control group). Thermography was performed using a ThermaCAM® B20 HS (Flir Systems®). At each examination a series of three pictures was taken. Thermographic pictures were evaluated using polygon tools of Thermocam-Researcher (FLIR Systems, Frankfurt) to determine minimal (minT), maximal (maxT) and mean (meanT) temperatures in each picture. Results: By comparing the mean standard deviation from the coefficients of variation of series (n=544) of three consecutive pictures, precision was best for meanT (0.12 %; 0.09), compared to maxT (0.19 %; 0.14) and minT (0.29 %; 0.21). Differences between hindquarters of individual measurements (n = 272) of udder healthy cows (meanT)

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were 0.17; 0.13 °C (Max.: 0.73). During 24 hours a diurnal variation of meanT was visible with a maximum of +0.45; 0.58 °C at 9:30 pm and a minimum of -0.42 - 0.39 °C at 6:30 am compared to an individual mean temperature calculated for 24 hours. Influence of the ambient temperature was statistically significant, but its effect was minimal. After infection (pi) both hind quarters showed a significant elevation in temperature, which was more pronounced in maxT than in meanT between 13 to 17 hours pi compared to values measured 24 hours before (maxT at 13 hours pi: infected quarter: +1.98; 0.59 °C; uninfected quarter: +1.93; 0.62 °C, p = 0.04 for both). Comparing meanT between hindquarters the difference was greatest at 13 hours pi and temperature was lower (-0.90; 0.64 °C) at the infected quarter. Differences between maxT in hindquarters were not significant at any time point. Conclusions: After experimental infection of the udder with E. coli significant changes due to inflammation can be measured by thermography. 80 Economic Assessment of Selective versus Blanket Dry Cow Treatment Options Including Teat Sealer Use H. Seegers , D. Billon , P. Roussel , F. Serieys , N. Bareille 1 Veterinary School & INRA, Animal Health Management Group, Nantes, France 2 Institut de l'Elevage, Angers, France 3 Filiére Blanche, Rennes, France Blanket antibiotic dry cow treatment has become challenged. This is of course to relate with residues and resistance, but also with more questions on its necessity, especially in herds with a low prevalence of intramammary infections at drying-off and a low risk for new infections during the dry period. Moreover, the use of an inert teat sealer can help to limit antibiotics use. However, no grounded indications for selective antibiotic treatment implementation (or teat sealer use alone or in combination with antibiotics) are available. A dynamic stochastic simulation model was used to assess several selection rules (using individual milk SSC thresholds from 100,000 to 300,000 cells/mL) for cows left untreated, subjected to antibiotics or treated with teat sealer. Simulated initial epidemiological conditions in a typical 50-cow herd covered a range of variation in type of pathogens, prevalence at drying-off, and risk of new infections during the dry period. The utility function to rank the rules combined: (i) gross margin, (ii) total number of antibiotic treatments per cow-year, and (iii) number of clinical cases per cow-year. Simulations assumed that blanket antibiotic therapy was replaced by another control option and the outcomes with this new control option were compared to those obtained with keeping on blanket therapy. On short-mid term horizons (<3 years), moving to selective antibiotic treatment instead of systematic treatment was almost financially neutral (no impact on gross margin), but the number of antibiotic treatments was reduced from 30 to 70%. Some economic benefit was only gained in specific low prevalence (bulk-tank SCC <150,000) and low risk situations (especially with predominance of minor pathogens). Detrimental effects on gross margin were observed for herds with bulktank SCC >300,000 cells/ml. However, on mid-long term horizons (>3 years), detrimental evolution was usually observed, justifying sometimes a return to blanket therapy. For options including the teat sealer use, neutral economic effects were mostly found, but also some very positive economic effects for specific high prevalence and high risk situations (especially with large implication of environmental pathogens). Finally, a decision chart based on prevalence of infections at drying-off, risk for new infections during the dry period and type of pathogens predominantly involved was proposed. Key words: dairy cow, dry-cow treatment, antibiotics, teat sealer 81 Risk Factors for Bacteriological Quality of Bulk Tank Milk in Prince Edward Island Dairy Herds A. Elmoslemany, G. Keefe, I. Dohoo Atlantic Veteirnary College, Health Management, Charlottetown, Canada The objective of this study was to determine on-farm risk factors for bacteriological quality of bulk tank milk. Bulk tank raw milk quality was evaluated on all Prince Edward Island dairy herds (n=235) over a two year period (March 2005-March 2007). Biweekly total bacterial, preliminary incubation, laboratory pasteurization and coliform counts, were conducted using Petrifilm®. For the assessment of risk factors, a

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case control study was conducted from January 2006 to May 2007. Case and control herds were defined based on the last 6 results of bulk tank bacterial count prior to on farm evaluation. A total of 69 herds (39 cases and 30 controls) were evaluated. Data collection included a survey of management practices, cow, environment and equipment hygiene scoring, milking procedures and mastitis control, complete wash analysis of the milking equipment and monitoring the presence of bacterial films on equipment. Data was analyzed using multivariable logistic regression. A total of 16 variables were associated with bulk tank milk bacterial quality at p<0.10 and were offered to the multivariable model. Only predictors significant at p< 0.05 were retained in the final model. The variables that were <0.10 in the univariable models, but did not remain in the final model, were bulk tank alkaline wash fill and drain temperatures, pipeline alkaline wash end temperature, bypass score and trap out score (higher values or scores were protective). Automatic cleaning of bulk tank, pipeline detergent frequency and the use of water softener were also protective. Additionally, high milking cow udder and leg hygiene scores (i.e. dirty) and high bulk tank outlet and pipeline far inlet bioluminescence scores were risk factors. The 5 variables which remained in the final model were, clipping the hair on the cows` udder (OR=0.14), water hardness score>6 (OR=4.08), pipeline alkaline wash start temperature (OR=0.93), pipeline alkaline wash alkalinity (OR=1.005) and teat end cleanliness score (OR=4.39). The last 3 predictors were measured on continuous scale and the corresponding OR represent the change per one unit change of the predictor. From this study, it can be concluded that equipment hygiene, milking system washing solution temperature and chemistry and udder hygiene are associated with bulk tank bacteriological quality. Key words: bulk tank milk, bacteria, management 82 Effect of Early Milking of Calved Heifers and Selenium Supplementation on Incidence of Clinical Mastitis in Dairy Heifers C. Compton, S. McDougall Animal Health Centre, R&D, Hamilton, New Zealand Dairy heifers have amongst the highest age-specific incidence of clinical mastitis (CM) (up to 25%). However, few preventive programmes are available specifically for heifer mastitis. A field trial was undertake to investigate the effects of reducing the interval from calving to first milking of freshly-calved heifers and supplementation with selenium pre-calving on the incidence of CM and prevalence of intramammary infection (IMI) in early lactation. Heifers (n=559) from 4 spring-calving dairy herds were enrolled approximately 6 weeks prior to the expected start of calving for each group and randomly assigned to treatment groups in a 2 by 2 factorial arrangement. Selenium supplementation started at enrolment using a pour-on application of 7.5 mg of selenium as selenium selenate (i.e. 1.5 ml Selpor®, Ancare, N.Z Ltd) per 50 kg liveweight and continued at 3-wk intervals in un-calved heifers. Heifers were removed twice daily from the un-calved group and immediately milked (Treatment), or removed only once-daily in the morning and milked first at the afternoon milking (Control). At the first and 8th milking, milk leakage, teat canal closure and udder oedema were scored. Milk samples were taken for bacteriology from all quarters on the 4th day of milking and from quarters identified by the farmers with clinical mastitis up to 3 weeks post-calving.The mean interval (h) from calving to first milking for milking post-calving differed between treatment groups (9.8 (SD=7.3) h and 19.5 (SD=11.4) h, p<0.001). Twice-daily removal reduced the incidence of CM (8.0% and 14.5% for treatment and control groups, respectively, p=0.04). The risk of st CM increased linearly with the interval from calving to 1 milking (p=0.009). Selenium supplementation had no effect on CM incidence (12.2% and 10.3% for control and treated groups, respectively, p=0.60). The risk of severe udder oedema was reduced in early milked heifers (13% vs. 20% in treated vs. control heifers, p=0.05) at the first milking. The quarter prevalence of IMI at the 8th milking did not differ between early-milking or selenium supplement treatment groups (13.3% vs. 15.1%, p=0.27; and 14.5% vs. 13.9%, p=0.76; for treatment vs. control groups, respectively). It is concluded that early calf removal, but not selenium supplementation reduced the incidence of clinical mastitis in dairy heifers. 83 Effect of a Killed Whole-cell Streptococcus uberis Vaccine on Prevalence and Incidence of Streptococcus uberis Mastitis S. McDougall , G. Tait , J. Moffat 1 Animal Health Centre, R&D, Hamilton, New Zealand

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Schering Plough Animal Health, R&D, Wellington, New Zealand

Streptococcus uberis is common cause of clinical and subclinical mastitis. The objective of this study was to assess the efficacy of vaccination of dairy cows with a whole-cell killed vaccine containing S. uberis on the incidence and prevalence of mastitis associated with S. uberis. Cows from 3 dairy herds were either vaccinated with a vaccine containing S. uberis (n=695) or injected with the vaccine vehicle (n=711 cows) on 2 occasions approximately 4 wk apart with the 2nd vaccination occurring on average 55 days before calving. Milk samples were collected from all 4 glands from a randomly selected 2/3rds of the enrolled cows between 0 and 7 days after calving, and milk samples were collected from cows with clinical mastitis (CM) within 21 days of calving. The prevalence of intramammary infection associated with S. uberis and the incidence of CM associated with S. uberis were analysed at cow and gland level using multivariable models which accounted for the clustering of gland with cow and cow with herd and adjusted for the effect of potential confounders such as age and breed. Vaccination tended to reduce the prevalence of cows infected with S. uberis in any gland within seven days of calving (0.12 (0.09-0.16) vs. 0.16 (0.130.21) for vaccinated vs. control cows, respectively; p=0.057). However, there was no difference in the prevalence of S. uberis at gland level (0.043 (0.031-0.059) vs. 0.055 (0.042-0.072) for vaccinated vs. control cows, respectively; p=0.18; RR=0.785 (0.548-1.124)) or in incidence of CM associated with S. uberis between the groups at either cow (43/670 (6.4%) vs. 44/693 (6.3%) in vaccinated vs. unvaccinated cows, respectively; p=0.95) or gland (53/2628 (2.0%) vs. 48/2724 (1.7%) in vaccinated vs. control cows, respectively; p=0.50) level. Vaccination reduced the number of S. uberis present in milk in glands from which it was isolated (940 (532-1661) vs. 2476 (1200...) geometric mean colony forming units/ml for vaccinated vs. control cows, respectively; p=0.013). There was no difference in SCC at first herd test between vaccinated and control cows (71 (64-78) vs. 75 (6883) geometric estimated marginal mean (95% CI) x 103 SCC/ml for vaccinated and control cows, respectively; p=0.32). It was concluded that the whole-cell killed S. uberis vaccine evaluated in this study was not effective in reducing the prevalence and incidence of mastitis associated with S. uberis. 84 Prototheca blaschkeae Associated with Bovine Mastitis S. Marques , E. Silva , C. Kraft , V. Huss , G. Thompson 1 Universidade do Porto - ICBAS, Department of Veterinary Clinics Laboratory of Infectious Diseases, Vila do Conde, Portugal 2 Friedrich-Alexander - Universität Erlangen-Nürnberg, Department für Biologie, Molekulare Pflanzenphysiologie, Erlangen, Germany Objectives of study: Algae of the genus Prototheca and more rarely Chlorella are the only known plants that cause infectious diseases in humans and animals. Prototheca are ubiquitous, generally saprophytes, and can be isolated from a great variety of environmental sources. Until recently only two species were known to be associated with disease in animals and humans, Prototheca zopfii and P. wickerhamii, respectively. In 2006, a new species P. blaschkeae was described, which has been isolated from a case of disease in men. The prevalent form of protothecosis in animals is bovine mastitis, which generally occurs in a chronic subclinical or a mild clinical inflammatory process of the udder affecting cows that do not respond to routine therapy. The objective of the present work was to characterize Prototheca spp. isolated from bovine mastitis. We provide evidence for the first time, that P. blaschkeae is associated with bovine mastitis. Material and methods: Forty one Prototheca isolates obtained from mastitic milk from dairy cows belonging to 22 different farms in the North of Portugal were characterized by conventional microbiological methods and by phylogenetic analysis of their 18S rDNA sequences. The methods employed were morphological data analysis, biochemistry profile determination by API 20C Aux and BBL Crystal Kits (Enteric Nonfermenter and Gram positive) systems, PCR amplification of the 18S rDNA, amplified rDNA restriction analysis with HaeIII, and phylogenetic analysis of the 18S rDNA sequences. Results: Macroscopical and essentially microscopical analyses revealed two groups of algae, one that contained the majority of the isolates and the other with only four isolates. Both groups were confirmed by their biochemical profiles. PCR amplification of 18S rDNA and phylogenetic analyses showed, that most of the isolates were identified as P. zopfii var. hydrocarbonea, whereas the four isolates of the other group were identified as P. blaschkeae.

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Conclusion: Although the type of mastitis induced by all Prototheca isolates had similar clinical manifestations, we show for the first time, that besides P. zopfii also P. blaschkeae may be involved in bovine mastitis. Key words: bovine mastitis, Prototheca, genotyping 127 Controlling Highly Prevalent Staphylococcus aureus Mastitis from the Dairy Farm H. Nagahata , H. Maruta , S. Matsuki , H. Higuchi , K. Kawai , 3 A. Anri 1 Rakuno Gakuen University, Department of Animal Health, Ebetsu, Japan 2 Tokachi NOSAI, Seibu veterinary clinical center, Obihiro, Hokkaido, Japan 3 Hokkaido NOSAI, Unit of Animal Health Management, Ebetsu, Japan Objective: Staphylococcus aureus, a contagious pathogen that spreads easily from cow to cow, usually manifests as subclinical mastitis. A dairy farm with increasingly elevated somatic cell counts (SCC) in the bulk milk and a high rate of S. aureus-mastitis was referred. Our study monitored the herd for 3.5 years after we instituted a mastitis control program. In addition, to evaluate whether a high prevalence of S. aureus infection in lactating cows affects the occurrence of S. aureus infection in heifers in the herd, bacteriological findings in mammary secretions from heifers were examined. Methods: Dairy farm had 57 lactating Holstein cows were housed in freestall barns. A total of 15 heifers, 23-26months old, were also used for the survey of microbiological findings in mammary secretions. Quarter milk samples from lactating cows were cultured, and pathogens were identified. Mastitis control program: To determine the contributory causes of S. aureus infection, we checked the milking procedures and implemented the following mastitis control measures: (a) maintenance of the correct milking order, (b) recommended milking procedures, (c) post-milking teat dipping with approved disinfectants, and (d) culling of cows infected with S. aureus in 2 or more quarters or cows having chronic S. aureus infection. Results: S. aureus was detected in 24 (41.2%) of the 54 lactating cows and in 29 (12.8%) of 227 quarters of the 57 milking cows in the herd. The S. aureus infection rate decreased steadily from 42.1% at the time of our first visit to 10.5% seven months after start of the control program, then decreased to 4% at 14 months, and no isolate of S. aureus was found in milk samples at 18 months. In the course of the 3.5-year monitoring period, 24 lactating cows infected with S. aureus were culled. The average 4 SCC in the bulk tank milk decreased gradually from 52.3 x 10 cells/ml 4 4 4 (21 to 94 x 10 cells/ml ) to 14.5 x 10 cells/ml (4 to 21 x 10 cells/ml), as observed 18 months after start of the control program. No S. aureus isolate was detected in 58 mammary secretions from 15 heifers at 4-5 weeks before parturition. Conclusion: The program implemented is effective in controlling S. aureus mastitis on the dairy herd. A high S. aureus prevalence in the herd may not necessarily be a decisive factor for S. aureus infection in heifers. Key words: long-term monitoring, mastitis control program, S. aureus 128 The Recoveries from Acute Mastitis and Fertility Indices of Cows Treated with Antibiotics and Additional Medicine E. Malinowski, S. Smulski, H. Markiewicz National Veterinary Research Institute, Department of Pathophysiology of Reproduction and Mammary Gland, Bydgoszcz, Poland The aim of studies was to establish the effect of antioxidants, immunomodulator or NSAID single injection on results of mastitis therapy and fertility indices of mastitic cows treated intramammarily with antibiotics. Examinations were performed in 4 farms on 240 dairy cows th that showed acute form of mastitis as only clinical disease between 10 th and 100 day after parturition and on 60 healthy cows calved at the same time. Mastitic cows were treated with approved intramammary products in doses recommended by manufacturers. Animals included to I group received only intramammary therapy (IMT). Cows from group II, III and IV, apart from IMT, were once injected with antioxidants (vit. C, vit. E, bcarotene, Se), lysozyme dimer) or flunixin meglumine, respectively. Healthy cows from group V consisted the control for fertility indicators of mastitic animals. In average, 55.6% of cows recovered as an effect of the first course of the intramammary treatment. A single injection of antioxidants or lysozyme dimer increased recovery rates. Fertility indices: calving to the first service, conception rate, insemination index and

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Mastitis / Udder Health and Milk Quality 49

calving pregnancy period were the best in the control group. The single injection of the additional medicine increased fertility indices if compared to cows treated intramammary only. Fertility indices were better in cows recovered as the effect of the first treatment and worse in cows that became chronically inflamed and required additional therapy of mastitis. It can be concluded: acute mastitis between 10th and 100th day postpartum elongates the period from calving to first insemination, decreases the conception rate, and increases the insemination index, calving pregnancy period and culling rate of dairy cows. More detrimental are the chronic cases than acute, successfully treated cases. The single i.m. injection of antioxidants (vit. C, vit. E, b-carotene, Se) or the lysozyme dimer increases the efficacy of intramammary antibiotic treatment of mastitis caused by Streptococcus spp., CNS, E. coli and even Staph. aureus. It seems the i.m. injection of flunixin meglumine increases the efficacy of coliform-mastitis therapy with antibiotics. However injection of additional medicine to intramammarily treated cows sick with acute mastitis improves their fertility indicators. Key words: cows, mastitis, antibiotics, antioxidants, lysozyme dimer, NSAID, fertility 129 The Contagious Staphylococcus aureus Genotype B Affects the Presence of Other Mastitis-Relevant Bacteria in Dairy Herds A. Michel, A. Steiner, HU. Graber Clinic for Ruminants, Vetsuisse Faculty, University of Bern, Berne, Switzerland Objectives of study: Staphylococcus aureus (S. aureus) is globally one of the most important pathogen of subclinical mastitis and causes large economic losses. In the present study, the effect of the contagious S. aureus genotype B on the presence of other mastitis-associated pathogens (coagulase-negative staphylococci (CNS), Streptococcus spp. (non-agalactiae streptococci), and Corynebacterium spp.) was investigated in Swiss dairy herds. Materials and methods: Sample herds (SH): 1057 milk samples from 265 cows (10 farms) with diagnoses of S. aureus genotype B problems were collected. As control herds (CH), 257 cows (20 farms with 1021 milk samples) were selected with at least 1 S. aureus isolate of non-B genotype (genotypes A, C, H, E, D, R). All the samples were bacteriologically analyzed according to standard guidelines (NMC). Genotyping was done by ribosomal spacer-PCR. For the SH and CH, the prevalences on cow and quarter levels of S. aureus, Streptococcus spp., CNS and Corynebacterium spp. were compared using nonparametric statistics. Binary logistic regression was used to select those variables which best classified SH and CH. Results: Considering S. aureus, the cow and quarter prevalences clearly differed between SH and CH (p<0.001). In the case of SH, the median prevalences were 44.0% (cow) and 25.2% (quarter), respectively. For CH, the medians were 9.5% and 2.8%. The cow prevalences for CNS (27.5%) and Streptococcus spp. (15.6%) were much lower (p<0.001) in the SH than in the CH group (CNS: 76.4%; Streptococcus spp.: 32.7%). The same results were obtained on quarter level. Cow and quarter level prevalences for Corynebacterium spp. were similar in SH and CH, but the variation was different. In the case of SH, the quarter prevalence ranged between 10.9% and 45.5%, in the case of CH between 0.0% and 79.5%. SH were best discriminated from CH by a high S. aureus cow prevalence, the presence of genotype B, and a low CNS cow prevalence. Conclusions: In dairy herds, the contagious genotype B was associated with high cow and quarter prevalences of S. aureus together with low prevalences of CNS and Streptococcus spp. The prevalence of Corynebacterium spp., however, was not related to the presence of specific genotypes of S. aureus. The reasons for this newly observed phenomenon remains to be elucidated. Corynebacterium spp. was very contagious and was observed in all the SH. 130 Epidemiology of Infectious Mastitis: a PCR Evaluation of Staphylococcus aureus Carrier Mammary Glands F. Bispo , C. Rey , S. Matarazzo , J. Arcaro , E. Costa 1 Universidade de Sao Paulo - Instituto de Ciencias Biomedicas, Departamento de Microbiologia, Sao Paulo, Brazil 2 Instituto de Zootecnia, Bovinocultura Leiteira, Nova Odessa, Brazil Mastitis is an inflammatory reaction of the mammary gland, usually caused by bacterial infection, and is economically the most

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important disease in dairy milk production worldwide. A relevant aspect is the evaluation of asymptomatic mammary glands carrying pathogenic microorganisms, without showing evident signs of inflammation or detectable alterations in the milk secretion. Staphylococcus aureus (S. aureus) is one of the main causative agents of mastitis in dairy cows. A better understanding of these epidemiological relationships may lead to new approaches for the control of this disease. The aim of this study was to evaluate the diversity of S. aureus isolated from asymptomatic bovine carriers, by PCR amplification of the variable 3'-end region of the coagulase (coa) gene, and contribute to the elucidation of the carrier role in the epidemiology of infectious bovine mastitis. In a single dairy herd, mammary glands from eighty-three cross-breed cows were examined. S. aureus were isolated from milk samples of individual quarters from animals showing negative California Mastitis Test (CMT) reaction and quarter somatic cells count (QSCC)<200.000 cells/mL. Each morning, from these same mammary glands, during a 16 day-follow up period, milk samples were collected, after CMT testing, and submitted to microbiological examination and QSCC evaluation. Thirty-one S. aureus isolates were classified by morphology and biochemical tests. Total S. aureus DNA was extracted and the coa gene was amplified by PCR using specific primers. PCR products were visualized in ethidium bromide stained agarose gel after electrophoresis. Molecular typing of the coa gene of the 31 isolates revealed just a single fragment size of 800 pb. The genetic similarity presented by S. aureus isolates during the observed period from the same mammary glands demonstrated the carrier status. Key words: carrier status, mammary gland, S. aureus, PCR, bovine 166 Staphylococcus aureus Mastitis in Fresh Calved Heifers on a Dairy Farm in Hungary A. Tirian , M. Kovács , E. Brydl , O. Szenci , L. Könyves , 1 4 V. Jurkovich , E. Ungvári 1 Szent István University Faculty of Veterinary Science, Department of Animal Hygiene, Herd Health and Veterinary Ethology, Budapest, Hungary 2 University of Kaposvár, Department of Animal Hygiene, Herd Health and Veterinary Ethology, Kaposvár, Hungary 3 Szent István University Faculty of Veterinary Science, Large Animal Clinic, Budapest, Hungary 4 National Center for Epidemiology, Budapest, Hungary Objectives of the study: The goal of the study was to determine the prevalence of S. aureus mastitis in fresh calved first parity cows and compare strains in the producing and heifer population. The incidence of infection was 26% in the producing population. Heifer calves received the milk of the mother in the first 5 days after birth, in the following 2 months milk of treated masitic cows and high SCC group was fed to calves. Materials and Methods: Quartermilk samples of the investigated heifers (n=75) were taken 1­5 days after calving. Samples of the producing cows were taken in the past 16 months, those of the mother cows were taken during the lactation following the birth of the heifers. The samples were streaked on Columbia blood agar, for identification of bacteria Staphylase test (Oxid Diagnostic Reagents) was used. Typing of strains was done via Pulse-Field Gel Electroforesis (PFGE) by using contour-clamped homogenous electric field (CHEF-DR II) system (Bio-Rad Laboratories). Results: The prevalence of S. aureus mastitis in the first calved cows was 21,3%. One of the heifers had 3 infected quarters, 3 had 2 infected quarters and 13 had only 1 S. aureus positive quarter. The samples of the heifers belonged to 6 different PFGE types (A, B, C, F, G, H). The strains of the producing cows belonged to 7 types (A, B, C, D, E, F, G). The PFGE investigation showed close relationship (>95% similarity) of types A, B and C; D and E; F and G. All of the 7 types were >89% similar, only type H showed larger difference. In 6 pairs were possible to compare the types of the heifer and mother. The pathogens were homologue in only 1 pair, in 5 pairs were heifers' S. aureus strains different from the strains causing mastitis in the mother cow. Conclusions: The risk of freshening heifer population with S. aureus mastitis in a heavily infected herd is high. Sources of infection are not easy to determine but beside feeding infected colostrum also other management related risks like feeding of contaminated milk may play a major role in the spreading of infection among heifers. Key words: mastitis, S. aureus, transmission

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50 XXV. Jubilee World Buiatrics Congress 2008

167 Genotyping of Staphylococcus aureus Isolated from Bovine Mastitis Cases and Asymptomatic Carrier Mammary Glands F. Bispo , I. Rivera , J. Arcaro , E. Mamizuka , C. Mendonça , 1 1 P. Melville , E. Costa 1 University of Sao Paulo - Institute of Biomedical Sciences, Microbiology, sao Paulo, Brazil 2 Instituto de Zootecnia, Bovinocultura Leiteira, Nova Odessa, Brazil 3 Universidade Federal Rural de Pernambuco, Departamento de Medicina Veterinária Preventiva, Recife, Brazil The mastitis caused by Staphylococcus aureus is one of the main diseases in dairy herds and is responsible for chemical, physical, and microbiological alterations in milk, resulting in serious economical losses and public health risk. Chronically infected cows are the main reservoir of infection in a herd; however, there are other contamination sources and reservoirs, such as milking unit liners, and human and bovine skin. Additionally, the carrier status is characterized by mammary glands without clinical signs of inflammation, negative CMT reaction, and 3 SCC<200x10 cells/mL, and positive results in microbiological tests. Accordingly, the aim of this study was to genotype S. aureus isolates of clinical and subclinical mastitis cases and asymptomatic carrier mammary glands, from four different Brazilian dairy farms, in order to elucidate the epidemiological importance of carrier animals as a source of infection. Asymptomatic carrier mammary glands, identified by negative CMT 3 reaction and SCC<200x10 cells/mL, were followed during a 16 day period with every day sampling for CMT, SCC and microbiological tests. S. aureus isolates were characterized by morphology and biochemical tests. Macrorestrictions of chromossomal DNA of the isolates were performed by PFGE using SmaI endonuclease. The pulsotypes were then analyzed using Bionumerics software. Eleven different pulsotypes were identified by PFGE and grouped in eight different lineages. Isolates from clinical and subclinical cases and asymptomatic carrier mammary glands were classified in the same lineage. Also, the same pulsotype was observed in the same mammary gland for a period of 16 days. In conclusion, it was not possible to associate different pulsotypes to varied degrees of mammary gland inflammation or carrier status, and the existence of S. aureus asymptomatic carrier during a 16 day follow up period is suggested. Key words: carrier status, mammary gland, S. aureus, PCR, bovine mastitis 168 SCC Monitoring in my Practice I. Király , T. Barkóczi 1 Pet[fi Agricultural Ltd, Ostffyasszonyfa, Hungary 2 Dairy Service, Veresegyház, Hungary The goal of this study was SCC monitoring in our practice. Our milking parlour - Bou-Matic system of 40 stalls - started its operation on the 01.03.1999. The average number of cows in milk used to be 480-580/ day since 01.03.1999. There is a possibility to acquire data from the system. During the last seven years I have been collecting the following data: milk production, ID and time of attachment, activity, milk conductivity, milking time. Milk conductivity - like on other dairy farms in the World - is an inadequate measurement for mastitis detection. However it is suitable for estimation of bulk milk SCC. During every milking, milk conductivity of all milked cows is measured. By means of a competent computer program bulk milk SCC can be estimated well. Further analyis of collected data makes documentation of total milking. This documentation is suitable to be built in into any ISO system as average milk conductivity in the different milking groups. Key words: SCC, bulk milk 169 The Hiperkeratosis of the Teat Channel in Portuguese Dairy Farms. General Causes and Microbiological Effects J. Sousa , C. Gomes , A. Pereira , H. Madeira , J. Niza-Ribeiro 1 Segalab - Laboratório de Sanidade Animal e Segurança Alimentar, S. A., SASA, Matosinhos, Portugal 2 Universidade de Porto, ICBAS, Instituto de Ciencias Biomedicas de Abel Salazar, Porto, Portugal Hyperkeratosis is a known perceived problem in dairy cows and it has been related in recent studies with increased vulnerability of the udder to new intramammary infections. Many environmental, management and

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individual determinants may be associated with high levels of hyperkeratosis. The objective of our study was 1) to characterize the hyperkeratosis in 17 dairy farms, representative of Portuguese reality, 2) to record putative risk factors present in these farms and relate them with the observed teat condition, 3) to investigate the relation between hyperkeratosis and mastitis and 4) the existence of associations between the teat condition and the bacterial pathogens in the milk of the observed cows. The 17 farms were visited twice in consecutive weeks: in the first visit, composite milk samples from each cow were collected for bacterial analysis and the udder health was checked for clinical mastitis; Californian Mastitis Test of all quarters was also carried; in the second visit the teat hyperkeratosis and possible risk factors were observed and recorded. A logistic regression analysis was performed. Of the 17 dairy farms, 71% showed more than 20% of cows with high levels of hyperkeratosis. The prevalence of cows with high levels of hyperkeratosis in the dairy farms visited ranged from 5% to 43%. The risk factors associated with the presence of hyperkeratosis were, the teat shape (OR=,3,45 p<0,00) and position (OR= 1,26, p<0,05), the teat length (OR= 1,68, p<0,00), cow's parity (OR= 2,72, p<0,00) and the days in milk (OR= 2,08, p<0,00). The milking vacuum in the liners, the existence of automatic liner detachers and the disinfection of the teats after milking were also statistically related with hyperkeratosis but their effect is not yet clear. Teats with hyperkeratosis showed a higher risk of mastitis and the risk increase with the severity of hyperkeratosis. The teats with hyperkeratosis had a greater risk of infection with Enterococcus spp (OR= 2,02, p<0,05), Enterobactereaceae (OR= 1,73, p<0,05) and Streptococcus agalactiae (OR= 2,76, p<0,05) and cows with low levels of hyperkeratosis were in at lesser risk of having udder infection (OR= 2,42, p<0,00). The study raises the hypothesis that hyperkeratosis may weaken the protective function of the teat channel and it may increase risk of mastitis the observed dairy farms; its prevention can play a key role in mastitis prevention and control. Key words: hyperkeratosis, odds ratio (OR), mastitis 170 Teat Stenoses and Injuries are an Important Aspect of the Daily Work in Dairies. Surgical and Non-surgical Procedures which are Possible in the Cowshed R. Ebner , F. Schlederer 1 Tierärztliche Gemeinschaftspraxis Dr. Rupert Ebner / Dr. Thomas Würgau (Fachtierarzt für Gynäkologie, Ingolstadt, Germany 2 Tierarztpraxis Schlederer, Arndorf, Austria The optimal machine milking procedure is only possible with a healthy functional teat. When a malfunction of the teat occurs, because of injury, treatment is often carried out by non-professionals with the result that the problem worsens. The cow then becomes generally unwell which in the worst case scenario could result in the loss of the cow. Modern methods, such as "lateral theloscopy" which was initially shown at the XXII World Buiatrics Congress 2002 in Hannover, Germany, demonstrated a clinical procedure for the first time in front of an international audience. This demonstration gave the opportunity to view the injuries, which were previously hidden, from a completely new perspective. This method also presents the opportunity to view the damage caused by mistreatment. The success and plausibility of this method has shown that teats can be successfully treated in the cowshed. The further development of this method has simplified it, and furthermore, has lead to a trusting relationship between the farmer and the veterinarian. The most important alternatives to treat teat injuries: amputation; easy closure of perforations (all structures with one suture); and of course, lateral theloscopy will be shown, complete with photographs taken during the procedure in the cowshed. The measures taken before and after surgery, which are as important as the surgery itself, are also explained and displayed in different tables. All the above mentioned methods to treat injured teats can reduce the high level of replacement in dairy herds. This is of paramount importance to the farmers and their economic situation. Not least, is the fact that this treatment improves animal welfare and reduces the risk to the farmer of being kicked by the cow. Key words: teat stenoses, surgical and non surgical teat treatment, lateral theloscopy, injured teats 171 Investigation of an Extended Therapy Regime using Penethamate Hydriodide for the Treatment of Persistent Intramammary Infection in U.K. Dairy Cows J. Breen , M. Green , A. Bradley 1 University of Bristol, School of Veterinary Science, Bristol, UK

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University of Nottingham, Department of Veterinary Medicine, Nottingham, United Kingdom 3 Orchard Veterinary Group, Glastonbury, United Kingdom 4 Quality Milk Management Services Ltd., Wells, United Kingdom Objectives: The primary objective of the study was to determine the impact of the duration of treatment on the efficacy of penethamate hydriodide (Mamyzin® Injection) in the cure of persistent bovine intramammary infection in U.K. dairy cows. Materials and Methods: Cows with a history of a high somatic cell count (SCC), four functional quarters, no significant teat lesions and in good general health were enrolled into a treatment study (n=95). Two different regimes were assessed using penethamate hydriodide, which was administered either as a four-day ("on-label") course or a seven-day ("extended") course. Intra-mammary infection status was monitored using both bacteriology and quarter somatic cell counting from 1-week pre-treatment until 4-weeks post-treatment. Multi-level statistical models were specified and interpreted within a Bayesian framework to estimate the probability of a difference between the two treatment groups (expressed as an odds ratio), using a flat, normal prior distribution. Results: Preliminary results have shown that both treatment groups experienced a significant reduction in the prevalence of gram-positive major pathogens from before until after treatment accompanied by a fall in somatic cell counts at both the quarter and cow level. There was a reduced odds of identifying a major pathogen after treatment in quarters treated for 7 days compared to quarters treated for 4 days (mean odds ratio = 0.57) and a posterior probability of 93% that treatment for 7 days resulted in fewer quarters with major pathogens than treatment for 4 days. There was an increased odds that quarters would remain below 200,000 cells/ml for 28 days after treatment for 7 days compared to treatment for 4 days (odds ratio = 1.87) and a posterior probability of 79% that treatment for 7days resulted in fewer quarters > 200,000 cells/ml than treatment for 4 days. Conclusion: This study demonstrates the efficacy of Mamyzin® Injection in the treatment of persistent IMI in dairy cows in the U.K. and has identified an increased probability of a cure when using a 7day, extended therapy regime, compared to 4-day, on-label treatment regime. In addition, the results can be used to inform clinical decisionmaking for the treatment of high SCC cows. Key words: somatic cell count, penethamate hydriodide, extended therapy, persistent infection, streptococcus uberis 172 Challenge with Streptococcus uberis to Evaluate the Efficacy of Teat Sealers in Preventing Mastitis N. Williamson, K. Petrovski, C. Fernandez Massey University, Institute of Veterinary, Animal and Biomedical Sciences, Palmerston North, New Zealand External teat sealers based on tetrahydrofuran (DryFlex (DF)) and a wound seal for humans 2-octylcyanoacrylate (Band-Aid (BA)) to prevent dry period mastitis were investigated. Streptococcus uberis has the highest incidence in the dry period and early lactation in New Zealand. It is a ubiquitous environmental pathogen, is normally easy to cure using antibiotics and thus was an appropriate challenge organism. Quarter milk samples were taken for bacteriological culture from 175 cows, 4 and 1 days before dry-off and twice within 4 days after calving. After the last milking, 88 cows received DF and 87 cows received BA. Pairs of contralateral quarters were randomly assigned to the treated or the control group. All quarters were dipped in a Strep. uberis broth 2 and 4 days after dry-off and were assessed daily for clinical mastitis. Clinical mastitis iwas greatest 6-11 days after dry-off in both groups. Strep. uberis was the most prevalent bacterium isolated. In the DF group, 35 cases of clinical mastitis occurred in 176 treated quarters, compared with 83 in the 176 control quarters. In the BA group, 67 cases of clinical mastitis occurred in 174 treated quarters, compared with 64 in 174 control quarters. DryFlex reduced mastitis after dry-off but not after calving. Octylcyanoacrylate did not reduce clinical mastitis after dry-off. A DryFlex type teat sealer showed potential in preventing mastitis in the early dry period, but has now been withdrawn worldwide. The effect of experimentally induced clinical mastitis in the early dry period on the subsequent lactation was examined. Cows (127) developing clinical mastitis after challenge were treated with antibiotic. Yields of milk, fat, protein and milk solids were analysed with respect to treatment group. No significant effect of treatment group on lactation yields of milk, fat, protein or milk solids occurred. No difference in production occurred (milk yield, 5126 vs. 5010 litres; fat yield, 267 vs. 264 kg; protein yield, 182 vs. 179 kg) between cows that had mastitis and

those that did not. Promptly treated clinical mastitis due to Strep. uberis did not affect production in the subsequent lactation. The ability to demonstrate differences in efficacy and absence of production effects in the subsequent lactation mean this challenge model appears suitable for evaluating teat sealers. Key words: mastits, Streptococcus uberis, teat sealer 173 Use of Leukocyte Differential Patterns to Optimize Diagnosis of Bovine Intramammary Infections: Evaluation of Partitioned vs. Non-partitioned Testing A. Rivas , K. Anderson , G. Leitner , M. Chaffer , O. Krifucks , 1 R. Rodriguez 1 North Carolina State University, College of Veterinary Medicine, Raleigh, NC, United States 2 Kimron Veterinary Institute, Bacteriology,Beit-Dagan, Israel Objectives: To explore the use of leukocyte patterns in diagnosing infectious mastitis. Materials and methods: Quarter milk samples were investigated in 3 cross-sectional studies conducted in the US (study I, n=120; and study III, n=500) and Israel (study II, n=484). The somatic cell count (SCC), milk microbiological cultures, and the percentages, total counts and relative proportions of lymphocytes, macrophages and neutrophils were established. Non-periparturient (studies I and III) and periparturient (study II) cows were assessed. Samples were collected without (studies I and III) and with previous determination of health status (study II). Disease prevalence was >40% (study I) and <20% (studies II and III). An algorithm was used to group (or separate) similar (o dissimilar) leukocyte patterns. Microbial species were classified as either major (e.g., Staphylococcus aureus) or minor (e.g., CNS) pathogens. Milk culture results were used to establish the sensitivity and specificity of both SCC and leukocyte patterns. Results: Using leukocyte indicators (e.g., the neutrophil %, the lymphocyte %, the ratio of phagocytes/lymphocytes), 3 major patterns were observed: "no mastitis" (NM), "early mastitis" (EM), and "late mastitis" (LM). Significant differences in the major/minor (M/m) pathogen ratio were noticed: for example, EM showed a high M/m pathogen ratio, while NM displayed a low M/m ratio. A higher proportion of both disease-negative and disease-positive samples was correctly diagnosed by "partitioned" testing (based on samples subsets characterized by leukocyte differential patterns) than by the (SCCbased) "non-partitioned" testing method: 81.7 vs. 75.8% (study I), 98.6 vs. 98.1% (study II), and 92.6 vs. 84.0% (study III), respectively. Conclusions: "Partitioned" testing may (a) provide more information, (b) yield higher diagnostic accuracy, and (c) detect disease subsets likely to be missed by the SCC alone. Further studies, aimed at establishing the prognostic value, generalizability, and applicability of these findings are recommended. 174 Efficacy of a 5-Day Extended Lactational Therapy with Cephapirin Sodium in Dairy Cows Chronically Infected with Staphylococcus aureus JP. Roy , L. DesCôteaux , D. DuTremblay , F. Beaudry 1 Université de Montréal, Department of Clinical Science, St-Hyacinthe, Canada 2 Université de Montréal, Veterinary & Biomedical Sciences, St-Hyacinthe, Canada Objective: To determine the efficacy of a 5-day extended therapy with cephapirin sodium in dairy cows chronically infected with S. aureus. Materials and methods: The pre-selection of the chronically infected cows was based on anterior positive milk culture. The infection status was confirmed by bacteriological culture of individual quarter milk samples taken at 4 and 2 weeks before treatment. A cow was considered infected if 1 of the 2 milk samples was positive for S. aureus. The selected cows were blocked by parity (1, 2, 3 +), by DIM (<101, 101 to 200, >200) and were randomly allocated to treatment groups: 1) 40 cows treated for 5 consecutive days with 1 tube of CefaLak® per quarter BID (200 mg of cephapirin) or 2) 30 untreated control cows. Milk samples were taken for bacterial culture at 10, 24 and 31 d after the end of the treatment period. Bacteriological cure was confirmed by 3 negative bacterial cultures. Chi-square test was used at a significant level of 0.05. Results: Seventy-five (75) cows from 14 dairy herds in St-Hyacinthe (Québec, Canada) were enrolled in the study and 61 cows completed it. A

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52 XXV. Jubilee World Buiatrics Congress 2008

total of 31 dairy cows were treated and 30 cows were used as negative controls. The overall cow cure rate was 25.8% (8/31) in the treated group and 3.3% (1/30) in the control group. The difference between the 2 groups was statistically significant (P = 0.013). A tendency to a higher cow cure rate was also observed in the treated group when a cow had only one quarter infected (36.8%) compared to if more than 1 quarter was infected (8.3%) (P = 0.1). The quarter cure rates at 10 d post-treatment were 77.5% (38/49) and 17.6% (9/51) in the treated and control groups, respectively (P < 0.0001). The overall quarter cure rates were 39.4% (13/33) and 4.5% (2/44) in the treated and the control groups, respectively (P < 0.01). Antibiotic residues were tested on 13 treated cows. Only 1 cow had cephapirin residues above the accepted limit 4 d (96h) after the end of the treatment. This cow was under the accepted MRL limit 4.5 d (108h) after the end of the treatment period. Based on these results, the suggested milk withdrawal time for this study was set at 5 d (120h). Conclusion: The cow and quarter cure rates were significantly higher in the treatment group than in the control group. The treatment tends to be more effective when only 1 quarter was infected. A milk withdrawal time of 5 d (120h) was sufficient in all tested cows. Acknowledgements: The study was supported by Fort Dodge Animal Health.

milking machine (OR=2,45), inadequacy of teat cups (OR=1,48), infrequent and ill-defined scheme for cleaning the pulsators (1,41), milkers not trained for operating the milking equipments (2,51), no information about the mastitis pathogens present in the herd (1,84), immersion of teat cups in disinfectant solution between milkings (OR=2,19) and total insertion of cannula in teats during antibiotic treatment (OR=2,64). The high variation of SCC values (608.000967.000 cells/ml) suggests that others factor such number of infected mammary quarters and pathogens enrolled can influenced the results. The methodology used did not allow to identify all risk factors that increasing SCC. Therefore, the results can also to be used to improve the mastitis control programs currently used by these herds. Study supported by Fundaçao de Amparoa Pesquisa do Estado de Minas Gerais (FAPEMIG). Grant number EDT 401/05 Key words: bovine, mastitis, risk factors, somatic cell count 551 Some Epidemiological Studies on Subclinical Mastitis in Dairy Cows in Assiut Governorate A. Abdel-Rady , M. Sayed 1 Faculty of Veterinary Medicine, Assiut University, Animal Medicine, Assiut, Egypt 2 Faculty of Veterinary Medicine, Assiut University, Department of Food Hygiene, Assiut, Egypt In this investigation, some epidemiological studies were run on subclinical mastitis for totally 350 dairy cows of different breeds, ages and distributed in different villages in Assiut governorate, Assiut, Egypt, along a whole year (during the period from June 2006 till July 2007) through field screening surveys by using of the California mastitis test (CMT) for each quarter milk sample followed by bacteriological examination.The dairy cows were differed from the breed point of view as 230 Holstein Friesian breed and 120 native breed. Also, they were differed from the age point of view as a group of 95 cows aged from 2 to 4 years old and another group of 255 cow aged from 5 to 8 years old. The obtained results revealed that 67 cows (19.14%) had 80 infected quarters (5.71%). Prevalence 52.5, 31.25 and 16.25%, respectively. With studying the breed factor, it was found Friesian breed was sensitive towards infection (20.43% at the cow level and 6.09% at the quarter level) than of native breed (16.67% at the cow level and 5% at the quarter level). It was also noticed that the prevalence of subclinical mastitis in hot weather as during summer (9.14% at the cow level and 2.64% at the quarter level) and during spring (4.86% at the cow level and 1.36% at the quarter level) was higher than in cold weather as during winter (2% at the cow level and 0.64% at the quarter level) and during autumn (3.14% at the cow level and 1.07% at the quarter level). In relation to age susceptibility, 5-8 years old cows (15.43% at the cow level and 4.36% at the quarter level) were susceptible than those of 2-4 years (3.71% at the cow level and 1.36% at the quarter level). The degree of quarter attack according to positive CMT was varied from 35 quarters (2.50%) showed degree (+++), to 45 ones (3.22%) showed degree (++), to 120 ones (8.57%) showed degree (+) and the rest (85.71%) showed degree (-). The obtained results threw the light on the epidemiology of subclinical mastitis in Assiut villages and provided an importance of the CMT for diagnosis of subclinical mastitis due to it is a reliable, easy, rapid and cheap tool helping in diagnosis and controlling the disease. Programs for control of subclinical mastitis may be planned around the routine examination of all lactating cows, and consequently early treatment can be applied towards positive cases rapidly for preventing their conversion towards clinical form among dairy cows and for protecting the herd health, milk hygiene and consequently the consumer health. Key words: epidemiology, subclinical mastitis, California Mastitis Test (CMT), intramammary infection (IMI), dairy cows 552 Field Trial in Nothern Italy to Evaluate the Efficacy of Two Different Therapies against Acute Clinical Mastitis in Dairy Cows V. Cuteri , R. Froyman , L. Nisoli , A. Romero Tejeda , 1 1 S. Preziuso , A. Attili 1 University of Camerino, Veterinary Science, Matelica (MC), Italy 2 Bayer Health Care, Animal Health Division, Monheim, Germany 3 Bayer Health Care, Animal Health Division, Milan, Italy This study compared the efficacy of two different treatment regimens against acute clinical mastitis cows by enrolling 155 clinically ill dairy cows into a multicentric, controlled and randomised trial. One group (76

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POSTER ABSTRACTS

549 Study on Frequency, Etiology and Somatic Cell Counts of Subclinical Ovine Mastitis in Tabriz, Iran R. Beheshti Govij , J. Shaigh , J. Dulgari , M. Mousapour 1 Islamic Azad University-Shabestar Branch, Veterinary, Shabestar, Iran 2 East Azarbaijan Burau, Department of Veterinary, Tabriz Veterinary Network, Tabriz, Iran To investigate the period prevalence, etiology and some epidemiological features of subclinical mastitis in ewes from the Tabriz region, milk samples from 193 lactating ewes were aseptically collected for bacterial and somatic cell counts (SCC) tests. Milk samples were collected from April to May of 2007. An association was observed between the occurrence of subclinical mastitis and the age of ewe. The periodic prevalence rate of SCM was 13.5%. Staphylococci were the most prevalent bacteria, representing 88.4% of the isolates. Coagulase-negative staphylococci (69.2%), was the most prevalent species followed by staphylococcus aureus (19.2%). E.coli and C.bovis was the second bacterial group in importance according with the distribution among flocks representing 7.7% and 3.8% of the isolates. Subclinical mastitis seems to be, as deduced from the high prevalence observed in this study, an important health problem for milking sheeps in the Tabriz region. Key words: subclinical mastitis, sheep, etiology, prevalence 550 Risk Factors for Subclinical Mastitis in Dairy Cows G. Souza , C. Coentrao , J. Brito , M. Brito , W. Lilenbaum 1 National Center for Resarch on Dairy Cattle, Milk Quality Laboratoy, Juiz de Fora - MG - Brasil, Brazil 2 Universidade Federal Fluminense, Faculdade de Veterinária, Niterói - RJ - Brasil, Brazil 3 National Center for Research on Dairy Cattle, Milk Microbiology Laboratory, Juiz de Fora - MG - Brasil, Brazil 4 Universidade Federal Fluminense, Faculdade de Veterinária, Juiz de Fora - MG - Brasil, Brazil This study was conducted to identify risk factors for subclinical mastitis (SCC > 200,000 cells/ml). A total of 2,657 lactating cows from 24 herds in the State of Minas Gerais, Brazil, were included in the study. Each of the farms was visited three times in an 8-month period from November 2005 to June 2006. At each visit, all milking cows were examined for clinical mastitis by a single observer. A total of 3,987 milk samples were examined for somatic cell counts (SCC). Statistic analyses were conducted using descriptives statistics and logistic regression models to estimate the relative risk (odds ratio = OR). Considering all milk samples, the mean, median and standard deviation values for SCC were respectively 608,000, 219,000 and 967,000 cells/ml. Risk factors for subclinical mastitis were: udder positioned at the same height or below the hock (OR=1,73), presence of cracks or fissures in the rubber parts of the

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Mastitis / Udder Health and Milk Quality 53

animals) was treated with 5 mg enrofloxacin/kg BW (Baytril® 10% inj. solution) intravenously for 3 consecutive days while the other group (79 animals) was treated with 1 intramammary syringe of Cobactan® LC (75 mg cefquinome)/mastitic quarter three times every 12 hours and concurrently with one daily intramuscular injection of 1 vial Mamyzin® (equivalent to 10 grams of penethamate/head) for 3 consecutive days. The animals were followed for 14 days and received a daily physical examination of the udder on days 1 (enrolment day and first day of treatment), 2, 3, 5, 7 and 14. The rectal temperature, the udder secretion, and general symptoms were evaluated. Sterile samples of milk were collected on days 1, 5, 7 and 14 and were cultured to reveal the presence of pathogens. Further, the effect of treatment on the incidence of clinical mastitis, on bacteriological cure and on somatic cell counts was examined. The most frequently isolated microorganisms were Str. uberis (31/155, 20%) and E. coli (20/155, 13.0%). Both therapies resulted in a significant reduction (p <0.0001) of microbiologically infected quarters, i.e. from 100% to 7.69% (4/52) for Baytril® and to 7.14% (4/56) for Mamyzin®/Cobactan® LC. There was a full microbiological cure for all E. coli cases in both groups. There was an important decrease of somatic cells number in both treatment groups: from 106,75 to 106,16 for Baytril® and from 106,74 to 106,34 for Mamyzin®/Cobactan® LC. Cows with acute mastitis and treated with Baytril® recovered sooner from fever (p= 0.003) and generalised disease symptoms (p= 0.02) than those treated with Mamyzin®/Cobactan® LC. Post-treatment recovery of milk yield was likely better (p=0.06) for Baytril® treated cows as well. As to the other parameters, recovery was similar for both treatment regimens. Irrespective of the causative organism, intravenous Baytril® treatment cures acute mastitis more rapidly (fever, morbidity, milk yield) than the Mamyzin®/Cobactan® LC combination, resulting also in a reduction of both labour fee and milk withdrawal period (4 days vs 6 days). 554 Evolution of the SCCT during 2 Periods of 1 Year in Nothern Costa Rica L. CorderoUmana, J. Quiros, C. Boschini Universidad Nacional, Costa Rica, Clinica de Especies Productivas, Heredia, Costa Rica A project was laborated for a two year period to evaluate the rate of the somatic cells counts in tank (SCCT) in San Carlos a region which is the largest milk producing area of our country. The investigation included 1205 farms. During the first year (October 2001 to September 2002) 576 dairy farms were sampled (41.7 % of the total dairy farms in this region), and for the second period (October 2003-September 2004) 629 dairy farms were tested (45.48%). Every farms were visited once a week. Milk was collected from the tank and transported on ice to the laboratory where processed under a Fossomatic 420®. The data was analyzed with the program "SAS system". The data were reported by district, based on the political division of the country. The average of the SCCT for the first year was of 541.150 cells/ml with a standard deviation (SD) of 336.110, and for the second period the average was of 485.110 cells/ml with a SD of 263.400. The absolute annual difference average was 56.040 cells/ml. The dairy farms were grouped under seven categories, according to SCCT. The samples with less than 200.000 cells/ml were in the first category and we found 40 (6.96%) dairies during the first years and 57 (9.1%) dairies during our second year of study.For the second category (200.000400.000 cells/ml) we detected 172 (29.88%) dairies during the first year and 214 (34.09%) for the second year. For the third category and during the first year of study we detected 158 (27.56%) dairies with an increase during the second year to 190 (30.21%). A fourth category was detected in 97 (16.82%) dairies and 100 (15.9%) dairies, respectively during the first and second year of the study. For the fifth category (800.0001.000.000 cells/ml) 52 farms (9.09%) during the first year and 41 (6.49%) for the second year were observed. A total of 28 (4.78%) dairies during the first year and 16 (2.5%) for the second year have the sixth category (1.000.000- 1.200.000 cells/ml), and our final category (seventh) included farms with >1.200.000 cells/ml with 29 (4.92%) dairies for the first year and 11 (1.71%), for the second year. 555 A Study to Measure the Effect of Using a Sanitizer to Clean Cows' Teats before Milking, on the Bacterial Count of Teat Skin K. Fehlings , R. Huber-Schlenstedt , J. Deneke , A. Gangl 1 Bavarian Animal Health Service, Udder Health Service, Poing, Germany 2 Bavarian Animal Health Service, Bacteriology, Poing, Germany

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The impeccable hygienic quality of milk basically is defined by the content in saprophytic, human pathogenic and animal pathogenic bacteria. To ensure this bacteria have to be as low as possible. In three Bavarian dairy farms the sanitizing and cleaning of teats (60 randomly chosen cows) before milking with a 0.5% (750 ppm active chlorine) Tosylchloramide sodium solution (Desinficin® CL; DeLaval GmbH, Germany) applied to one-way paper tissues was carried out. A significant reduction of the total bacterial count (min. 98.8%, max. 99.3%, average 99.0%) on the teat skin as well as an effective reduction of the share of specific mastitis pathogens was shown. For use the residues of the disinfectant solution were squeezed out and the wet udder paper tissue used. Two control groups cleaned with one-way paper tissues showed a reduction of 87.5% (min. 82.5%, max. 92.7%). For demonstration of the bacteria reducing effect the average values of the logarithms were formed and the relative bacteria reduction by the treatment was calculated. The 3 bacteria per 2 cm2 before treatment were between 7.3 x 10 cfu (log 3.87) 5 3 und 5.2 x 10 cfu (log 5.72) and after treatment between 1.4 x 10 (log 2 3.15) cfu and 7.5 x 10 cfu (log 2.88). In the control group the bacteria 4 5 were between 3.0 x 10 cfu (log 4.48) und 8.7 x 10 cfu (log 5.94) and 3 4 between 1.6 x 10 (log 3.20) cfu and 9.3 x 10 cfu (log 4.97). A teat hygiene score (score 1 no growth/< 0.5 cfu, score 2 < = 10 - 30 cfu, score 3 < = 30 - 50 cfu, score 4 < 50 cfu) was used to show the reduction of contaminated teats (CNS, streptococci). The share of teats contaminated with mastitis pathogens decreased from score 2-4 corresponding to score 1 (70.4% resp. 72.5% remaining bacteria). The aim of this study was to check the positive effect of Tosylchloramide sodium solution on the reduction of bacteria on the teat skin under practical conditions ("sanitizing cleaning") in order to evaluate this approach with respect to the reduction of an infection risk of the mammary gland with special consideration of mastitis pathogens. Desinficin® CL is registered for sanitizing and cleaning of teats before milking and as drug for teat disinfection after milking. Probably it is a method eliminating dirt and faeces on the surface of the teats, but of appropriately reducing undesired bacteria from the udder/teat skin as far as possible. Key words: milk quality, udder cleaning, sanitizing cleaning, Tosylchloramide sodium solution, teat disinfection 556 Aflatoxin Contaminations in Milk and Feed in Kermanshah, Iran N. Jamali , M. Moeini , M. Ghasempour 1 Razi University, Animal Science, Kermanshah, Iran 2 Faculty of Veterinary, Animal Medicin, Kermanshah, Iran Alfalfa and corn silage and milk samples of 9 industrial and semiindustrials of dairy farms in Kermanshah were collected during 3 months experiment. Feed samples extracted with methanol: water (70:30 V/V) and analyzed by Enzyme linked Immune Sorbent Assay (Elisa) method. Milk samples collected from dairy herd and aflatoxin M1 (AFm1) measured by Elisa method. Blood samples were collected by tail venipuncture from %10 cows in each farm for measuring biochemical and hematological parameters.The result showed that the contamination AF in feed samples in small, medium and big dairy farms determined were: 12.47, 10.32 and 11.24 µg/kg respectively. The average of AF in January and end of February were: 9.392, 13.29ng/Kg DM. There was a significant different between aflatoxin in small and big size farm (p<0.05) and between aflatoxin in January and end of February (p<0.01). The AFm1 average of row milk of small, medium and big dairy farms were 33.9, 33.3 and 38.7 ng/kg respectively. The mean protein, albumin, WBC, RBC, lymphocyte and hematocrit did not differ significantly amongst the dairy cows.In this study the aflatoxin contaminations (B+G) found in feed samples was below the level of 50 µg/kg and the amount of AFM1 in % 77.8 of milk samples were lower than the maximum safe limit accepted by European Union. Key words: alfalfa, corn silage, aflatoxin, milk, cow 557 Studies on Some Aspects of Mastitis in Lactating One Humped Camels in U.A.E. A. Yas, N. Kamat Municipalities and Agricultural Sector, Animal Wealth, Abu Dhabi, United Arab Emirates A total of 630 milk samples collected from 162 lactating camels in U.A.E. were used. The over all incidence of mastitis was 18.52 % (7.94 % on quarter basis), The incidence of clinical and sub clinical mastitis was found to be 24.70% and 11.67% on animal basis; it being 9.70 %

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and 5.86 % on quarter basis respectively. The hind quarters were more frequently affected than the fore quarters Bacteriological examination of milk samples revealed that staphylococci were the chief etiological agents both in clinical and sub clinical mastitis (41.67%) in camels, followed by Streptococcus spp. (21.67%), Enterobacter spp. (15.00%), C. pyogenes (10.00%), Micrococcus spp. (5.00%) , Pasteurells spp. (5.00%) and Pseudomonas aeruginosa (1.66%). Most of the Staphylococcus spp., Streptococcus spp. and C. pyogenes strains were sensitive to carbenicillin, gentamycin , kanamycin, and erythromycin, but resistant to colistin and sulphamethoxazole. The other mastitis pathogens like Enterobacter, Micrococcus, Pasteurella spp. and Ps. aeuroginosa isolates were showed variable pattern of sensitivity to the antimicrobial agents. 558 Comparison of Pathogens and Antimicrobial Susceptibility of Isolates from Conventional and Robotic Milking Herds in Korea D. Kim, S.Y. Jeoung, S.J. Ahn Kangwon National University, School of Veterinary Medicine, Chuncheon, Kangwon, South Korea, In Korea, robotic milking system is introduced about 2 years before. However, mastitis control program on the robot milking dairy herds are not fully understood. An observational study was conducted to compare the pattern of pathogens and the antimicrobial susceptibility of Staphylococcus spp. from 8 conventional (pipeline of parlor milking) and 8 robotic milking dairy herds. All herds composed with Holstein cows and the robotic milking herds are using robotic milking system for at least 1 year before the start of this study. For each robotic milking farm, a neighboring conventional dairy farm served as a control. After bulk tank milk samples were collected, samples cultured on blood-esculin agar and Baird-Parker agar. The cultured staphylococci colonies were identified as Staphylococcus aureus and coagulase negative Staphylococcus (CNS) by tube coagulase test. Regardless of the milking system, Staphylococcus aureus was mainly cultured from the dairy farms in which teat dipping is not completely conducted and Streptococcus uberis was cultured from the farms in which udder washing was not completely conducted. The total of 119 Staphylococcus spp isolates was tested for minimum inhibition concentration (MIC) against 14 antimicrobial agents by microdilution method. The concentration that inhibits 90% (MIC90) of the analysed strains in micrograms per mililiter, for S. aureus were 0.5, 256, 16, 4, 1, 0.25, 1, 32, 8, 0.5, 64, 512, 1 and 0.25 to ampicillin, bacitracin, cephalothin, chloramphenicol, norfloxacin, erythromycin, gentamicin, kanamycin, oxacillin, penicillin, streptomycin, sulphamethoxazole, tetracycline, and vancomycin, respectively. And the MIC90 values for CNS were 1, 64, 0.5, 4, 1, 0.25, 0.5, 2, 0.5, 0.5, 4, 1024, 16 and 0.5 to ampicillin, bacitracin, cephalothin, chloramphenicol, norfloxacin, erythromycin, gentamicin, kanamycin, oxacillin, penicillin, streptomycin, sulphamethoxazole, tetracycline, and vancomycin, respectively. Resistant S. aureus isolates were found only to to ampicillin, bacitracin, oxacillin, streptomycin, sulphamethoxazole, and penicillin on the basis of NCCLS interpretive standards. Significant differences in antimicrobial susceptibility between robotic and conventional milking dairies were founded only to tetracycline. Key words: robotic milking, Staphylococcus spp, antimicrobial susceptibility, microdilution method, Korea 559 Use of Quarter Milk or Composite Samples for Diagnosis of Subclinical Bovine Mastitis in Dairy Farms C. Ribeiro , D. Castro , J. Galí , P. Teixeira J. Simoes 1 Centro Veterinário de Aveiro, Aveiro, Portugal 2 Grup Tecnic Veterinari S. L., Gerona, Spain 3 University of Trás-os-Montes e Alto Douro, Veterinary Science Department, Vila Real, Portugal Characterisation of pathogenic flora from composite milk (CM) samples of subclinical intramammary infected cows is frequently performed, instead of quarter milk (QM), in part due to economic reasons. One QM or CM samples are also used in cows with only one QM to positive California Mastitis Test (CMT). The aim of present work was to compare pathogenic flora between QM and CM samples from cows with one positive CMT quarter. Twenty Holstein-Friesian cows positive CMT in one quarter were randomized from 3 dairy farms. Both QM (positive CMT) and CM samples were collected in each cow. Somatic cell count (fossmatic) and microbiologic analysis (Plate Count Agar) were

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performed. Staphylococcus aureus were isolated in 35% (7/20), Streptococcus uberis in 20% (4/20) and Streptococcus agalactiae in 10% (2/20) of cows. Remained cows were affected by others contagious or environmental microorganisms, including fungi. Pathogenic flora was observed in QM but not in CM in 30% (6/20), in CM but not in QM in 5% (1/20) of cows, similar in 15% (3/20) and different in 50% (10/20; P<0.01) of both samples. Only 20% (2/10) of coupled samples with different flora had, at least, one similar microorganism. The number of somatic cells per ml was 3 3 3 3 6.400x10 ± 1.342x10 in QM (n=20) and 245x10 ± 58x10 in CM (±S.E.M., n=20, P<0.001). The linear score was also higher in QM (7.8 ± 0.4, n=20) than in CM (3.7 ± 0.4, n=20, P<0.001). In the present study, neither one QM nor CM samples were sufficient to caracterise the pathogenic flora of the udder from cows with one positive CMT quarter. This suggests the use of all QM in separate samples for more accurately epidemiological studies. Additionally, these results may justify some treatments failure and reinforce the importance of treatments others than the specific antibiotherapy in affected mastitic cows. Key words: mastitis, somatic cells, epidemiology, dairy cattle 560 Characterization of the Milk Lipid Globules in VarzeseOttonese Cow by Laser Scatter Granulometry M. Faustini , A. Riccardi , M.L. Torre , C. Colombani , 1 3 4 5 1 S. Scocca , M. Lazzati , M. Marone , M.L. Longeri , D. Vigo 1 University of Milan, Veterinary Sciences and Technologies for Food Safety, Milan, Italy 2 University of Pavia, Department of Pharmaceutical Chemistry, Pavia, Italy 3 Associazione Provinciale Allevatori, Pavia, Italy 4 ASL, Pavia, Italy 5 University of Milan, Animal Science, Milan, Italy Fat globule dimensional distribution influences micro- and macrostructural characteristics of milk and have significant impact on functional and sensory properties of dairy products (Michalski et al., 2007, Food Res. Int., 40, 191). Aim of this work was to verify the hypothesis that milk fat size presents breed-to-breed differences and it can be exploited as quality index for the safeguard of niche dairy products.Size distribution of milk fat globules for 19 Varzese-Ottonese cows, a Northern Italy autoctone endangered breed, has been evaluated; 18 Holstein Friesian milk samples have been considered as a control group. All cows were kept in the same farm (province of Pavia, Po Valley, Italy) and fed with the same feeding regimen. The analyses were performed by a laser light scattering granulometer (Beckman Coulter LS230) with refractive index set at 1.460 for milk and 1.330 for water. Milk samples were diluted in 120 mL of water in the measurement cell and treated with 1.25 mL of NaEDTA 35 mM at ++ pH 7 to chelate Ca and promote casein micelles dissociation. Size th th th distribution curves were processed to obtain linear 10 , 50 , 90 percentile diameters (d0.1, d0.5 and d0.9 respectively), volume-weighted diameter (d4,3), specific surface area (SSA) and span calculated as (d0.9d0.1)/d0.5. The differences for parameters between groups, resumed with mean and standard deviation, have been analysed with ANOVA. While d0.1 and d0.5 did not differ between breeds, d0.9 was significantly higher in Friesian cows with respect to the Varzese breed (5.15±0.63 Ïm and 4.65±0.70 µm, respectively, p<0.05), and so the d4,3 (3.31±0.61 µm and 2.99±0.26 µm, for Friesian and Varzese, respectively, p<0.05). 2 The SSA was lower in Friesian than in Varzese (23550±2576 cm /mL 2 and 24537±2252 cm /mL, respectively), but no differences have been statistically evidenced. The span of the diameter was found to be at the limit of significance (1.21±0.11 µm in Friesian and 1.09±0.26 µm in Varzese, p=0.08).Results evidence that several fat globule dimensional parameters differ between the breed: this feature can have a significant impact on fat cropping out during milk process, milk derivatives sensory characteristics as well as on digestibility. These indexes can be employed in quality control for the protection of niche foodstuffs. Grant by: FSE, Regione Lombardia, Min. Lavoro, INGENIO, PRIN, FAR. Key words: milk quality, milk fat globule, granulometry, laser scattering 561 Importance of the Udder Health Condition before Dry-off for the Subsequent Early Lactation N. Grabowski, J. Hamann, G. Klein University of Veterinary Medicine Hanover, Foundation, Institute of Food Quality and Food Safety, Hannover, Germany

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In dairying, good management practice should not only focus on calving and early lactation but also consider the foregoing late lactation, seeking to have a maximum of quarters secreting normally (i. e. culture-negative and < 100,000 somatic cells/ml foremilk) before dry-off. In order to evaluate the impact of the udder health status before dry-off has on the subsequent early lactation, quarter foremilk samples from 31 HF cows (n nd rd th = 126 quarters) in Northern Germany (2 , 3 , 4 and 5th lactation) were drawn weekly, three times before dry-off and eight times after subsequent calving, and analysed cyto-bacteriologically according to IDF recommendations. The udder health status as observed before dry-off was maintained during the early lactation in approx. 67 %. Of those, the ratio of normally-secreting quarters to quarters with secretion impairments was almost 1:1. While approx. 12 % of quarters secreting normally before dryoff became diseased during early lactation, curing was observed in another 21 % of quarters. Although varying in degree between lactations, the overall odds ratio analysis showed these factors to be significant (p < 0,001). Of the quarters with culturally-positive result before dry-off (n = 30 quarters), bacteriological cure during the dry period occurred in 67 % of cases. The same pathogen species re-emerged during early lactation in 13 % of cases while in 20 %, another pathogen species was diagnosed in that period. Regarding odds ratio analysis of the microbiological findings, colonisation itself also represents a weak factor of influence, while the pathogen type did not play a significant role. Results suggest that the udder health status before dry-off is of crucial importance for the development of the subsequent early lactation and, with that, the total lactational performance of a dairy cow (irrespective of her lactation number). The reaction potential of the body seems to account more than the species or group of pathogens involved, so that late lactation management should not only rely on mere hygiene but also promote metabolic homoeostasis (feeding, dry season). Key words: dry-off, udder health, early lactation, periparturient period 562 Efficacy of Extended Intramammary Therapy to Treat Clinical Mastitis in Lactating Dairy Cows under Field Conditions V. Kroemker, C. Zinke, J.H. Paduch, D. Klocke, J. Friedrich University of Applied Sciences and Arts, Microbiology, Hannover, Germany Streptococcus uberis and Staphylococcus aureus are important causes of mastitis in dairy cows in the entire world. Recent publications showed that an extended therapy can be more efficient in combating such intramammary infections. The objective of this study was to evaluate the effect of an extended intramammary therapy to treat moderate and severe mastitis cases in lactating dairy cows under field conditions in northern Germany. From December 2005 to August 2007, a total of 236 clinical mastitis cases (242 quarters) on 10 farms in northern Germany were enrolled in the study and randomly assigned (blocked by parity and body temperature) to one of three treatment groups (intrammammary lincomycin/neomycin 1.5-d (ALK-Control) or 5-d (ALL-Treatment 1), and 75 mg cefquinom 5 d (COL-Treatment 2)). Clinical and microbiological cure rates were evaluated. Treatments were initiated before culture results. Cows were observed and evaluated on d 1 to 6, 19 and 26. Nine cases in 236 (3.8 %) resulted in a lost cow (died or culled). Overall, treatments were not significantly different to controls regarding culled cows and clinical cure rate. However, when the bacteriological cure rate was evaluated, differences were observed. ALL appeared to be the most efficient treatment (43/44 [97 %] P < 0.01, ALK 24/32 [75 %] and COL 35/50 [70 %]). This was due to the fact that ALL yielded increased cure rates towards mastitis caused by Streptococcus uberis which was the most frequent micro organism. We conclude that in farms with Streptococcus uberis mastitis, the 5-d extended lincomycin/neomycin treatment regimen was significantly more efficient than the standard 1.5 d treatment. Key words: mastitis, extended therapy, Streptococcus uberis 563 Reasons for Culturally Negative Results of Quarter Foremilk Samples from Clinical Bovine Mastitis V. Kroemker, C. Zinke, JH. Paduch, D. Klocke University of Applied Sciences and Arts, Microbiology, Hannover, Germany It is well-known that approx. one third of all secretion samples from clinical mastitis cases do not yield any cultural bacteriological results. The

present study was aimed towards analysing the reasons for this unsatisfactory situation and eventually proposing some simple modifications of the microbiological analysis of milk samples. For this, 261 quarter foremilk samples from cows with clinical mastitis were reevaluated microbiologically after the routine analysis recommended by the German Veterinary Association (practically, IDF procedures) yielded no results (mastitis pathogen). After repeating this standard diagnosis procedure with an inoculum of 0.01 ml, culture-negative samples were analysed further. First, inoculum quantity was raised to 0.1 ml. In order to free intracellular bacteria, sediment was frozen. Inhibitory substances were detected using agar diffusion test with Paenibacillus stearothermophilus var. calidolactis. Analysis for Escherichia coli and other coliforms was carried out on 3MTM PetrifilmTM E. coli/Coliform Count Plates and 3MTM PetrifilmTM High-Sensitivity Coliform Count Plates, resp. The presence of endotoxins of gram-negative bacteria was verified using the limulus test while growth of Mycoplasma spp. was done on specific culture media. The sediment of samples was analysed both microbiologically on aesculin agar and microscopically. Of those 193 samples remaining culture-negative after the second standard analysis, mastitis pathogens could be detected in a total of 178 samples when the additional procedures were applied. Findings related to E. coli and other coliforms (24.4 %), coagulasenegative staphylococci (27.5 %), inhibitory substances (17.1 %), streptococci (6.7 %), Stapyhlococcus aureus (1.6 %), and mixed infections with coagulase-negative staphylococci being involved (7.8 %). Merely 7.3 % of samples remained bacteriological negative. Increasing the amount of inoculum leads to bacteriologically positive findings in 50 % of the samples that had been culture-negative before. Key words: mastitis, culturally negative, milk samples 564 Bacteriological Quality of Believed Milk (Case of the Farms of the Area of Mitidja) D. Guetarni, D. Baazize-ammi, E. Lebres, A. Dechicha, S. Kebbal University of Blida, Biology, Algeria, Algeria In Algeria, milk is paid neither on bacteriological quality, nor on the rate of the somatic cells and even less on the presence of the residues of inhibitors on the level of the dairies. The stockbreeders perceive premiums on the butyric and proteinic rates. The present study related to 146 samples of milk of tank taken on the level of the breedings of the wilayas of Algiers and Blida (circuit of collection). The evaluation of bacteriological quality, on the basis of FAMT, indicating germs of hygiene and certain pathogenic germs, i.e. the criteria retained in the Algerian legislation, shows that milk is strongly contaminated. The results show that 91,78% of the samples present an aerobic flora mésophile total > 105 UFC/ml and 80,13% reveal the presence of the total coliformes. These flora reflect the conditions of hygiene on the level of the exploitations. The high rates of contamination obtained are probably the result of a bad hygiene of the draft or a faulty operation of the tank of refrigeration. The contamination of milks by Staphylococcus aureus accounts for 80,13%. This important contamination can be due to a lack of hygiene or the consequence of the infections mammaires because the health of worse in our breedings represents a real problem. Escherichia coli was highlighted in 17,80% of the analyzed taking away. This germ signs a fecal contamination and represents a real danger to the health of the consumer. Key words: bacteriological quality, believed milk, total aerobic flora, Coliformes, Escherichia coli, Staphylococcus aureus 565 Bacteriological Quality of Believed Milk (Case of the Points of Sale of the Area of Mitidja) D. Guetarni, D. Baazize-ammi, A. Lebres, A. Dechicha, S. Kebbal University of Blida, Biology, Algeria This study aimed one hundred raw milk samples from milk shop (saling points) of Mitidja region. The valuation of the bacteriological quality shows that the milk is highly contaminated. Results showed that 81% of samples present a total aerobic mesophilic counts > 105 CFU/ml and total coliforms are present in 86% of milk. These counts reflect the hygienic level, the conditions of storage and transport. The obviousness of pathogenic agent, Escherichia coli, Staphylococcus aureus in 30%, 58% of milks respectively present a real danger for the consumer health. Enterococcus are placed in prominent position in 95% of milks and 13% of milks contained brucella antibodies. Key words: bacteriogical quality, raw milk, total mesophilic counts, Escherichia coli, Staphylococcus aureus, Enterococcus, Brucella

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566 Somatic Cells Count of Milk in Algeria S. Kebbal, I. Gharbi, M. Amara, S. Guemra, D. Guetarni University of Blida, Veterinary Medicine, Algeria For a long time used for the following sanitary and management engineering of the milk's cattle, count cells is been the official criteria of the milk's payment and the control milk. In the present work, we give a method of somatic cells' count to mean of particular counter "COULTER COUNTER" which is a new application in Algeria. After calibration of the device, an evaluation of its performances has been realised by the determination of reliability and repeatability of analysis on samples of quarter's milk classed before microscopic count by the method of Prescott & Breed. The results obtained revealed a good correlation (CC = 0, 98) with regard to the reference method. the repeatability has showed that the coefficient of variation don't exceed the 5% (3,26%, 2,39% and 3,57%) 3 for the cellular concentrations lower than 250 x 10 included between 250 3 3 3 x 10 and 750 x 10 and superior to 750 x 10 Cells/ml respectively. the reproducibility showed a coefficient of variation o 0,55%. The performances of "COULTER COUNTER" are good and this method of measure of cells concentration can serve for the somatic cells count of milk with a good precision. Key words: Mastitis, concentration, somatic cells, raw milk 567 Prevalence of Mastitis and Raw Milk Safety in Hamdallaye Dairy Cattle (Niger) A. Harouna , C. Locatelli , L. Scaccabarozzi , P. Cremonesi , 1 1 2 1 1 V. Bronzo , G. Adamou , P. Boettcher , G. Ruffo , P. Moroni 1 University of Milan, Milan, Italy 2 Institute of Agricultural Biology and Biotechnology (IBBA), Milan, Italy A study was performed in 2007 in Hamdallaye dairy herds (Niger) to establish the prevalence of subclinical mastitis and possible associated risks for public health. The studied sites are organized in the cooperative "UPROLAIT", created by Project of Safety of Niamey Periurban Farming and Agriculture. Seven herds participated in the study and 572 quarter milk samples were collected from 143 lactating cows from four local breeds: Azawak, Bororo, Djelli and Goudali. Each sample underwent bacteriological examination and automated counting of somatic cells (SCC). Selected strains (N = 15) of Staphyloccocus aureus isolated from the samples were subject to antimicrobial susceptibility tests and multiplex PCR to evaluate the potential production of enterotoxins A, C, D, G, H, I, J and L. Descriptive statistics of general mastitis prevalence, herd prevalence and relationships between bacteria and SCC were estimated using SPSS version 15.0. According to surveys of dairy farmers, 33% of cows had open sores on their udders and teats. Statistically significant differences in prevalence and SCC were observed among pathogen species (p < 0,05). General mastitis prevalence in Hamdallaye dairy cattle was 28,15% with: S. aureus (10,45%); Coagulase Negative Staphylococci (CNS) (15,55%) and environmental microrganisms (2,15%) being the most common pathogens. Herd prevalence ranged from 2% (Toka binekani) to 18% (Gagare) and from 4% (Bartchawal) to 39% (Toka binekani) respectively for S. aureus and CNS. The SCC ranged from 103 to 107 cell/ml of milk. Infections were assigned to four classes, according to the major pathogen, and the respective mean SCC (<0,05) were: non3 3 infected (262,9950 x 10 cells/ml); S. aureus (775,5606 x 10 cells/ml); 3 CNS (447,0555 x 10 cells/ml) and environmental micro organisms 3 (407,3125 x 10 cells/ml). Most of the tested strains of S. aureus were sensitive to the 13 antibiotics tested but only the combination of Neomycin-Bacitracin-Tetracycline was fit for treatment in all of the studied herds. In three herds all strains were resistant to Cefquinome and others to Ceftiofur and Rifaximina Danofloxacinand Marbofloxacina.Finally, all the 15 tested strains were negative with respect to the PCR tests for enterotoxins production. Nevertheless, we recommend that farmers continue to submit milk to warm treatment and sustain additional research of enterotoxigenic strains. Key words: dairy cattle, Niger, mastitis, toxins 568 Therapeutic Effects of Silver Ion on Clinical Mastitis in Holstein Cows SJ. Kang, TY. Hur, YH. Jung, YJ. Do, HS. Kim National Institue of Animal Science, Dairy Science Division, Cheonan, South Korea

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Silver has been widely used during this century to treat clinical diseases. Silver therapy is of significant clinical benefit to control bacterial infections, various new products containing the silver have been developed over the past decades. However, the scientific literature is totally devoid off in explaining any therapeutic response of silver ions against clinical mastitis in Holstein cows. The objectives of this study were to evaluate the effects of intra-mammary infusion of silver ion against clinical mastitis in Holstein cows. Cows; Ten quarters of six Holstein cows with clinical mastitis were selected on the basis of milk SCC (MSCC, > 2,000x1000cells/ml) and clinical signs. Silver ion was infused through intramammary route. Ten quarters of 6 cows were infused with 20 ml silver ion solution for consecutive 3 days. Blood samples at 0, 1, 2, 3, 5 and 7 days after intra-mammary infusion of silver ion were collected for hematological analyses from coccyegeal vein using K3 EDTA tube. Milk samples were aseptically collected from the infused quarters of each cow at 0, 12, 24, 48, 72, 120 and 168 hrs after intra-mammary infusion of silver ion. The clinical signs of mastitis in each cow responded differently to intra-mammary infusion of silver ion. However, significant reduction in swelling (80%), firmness of mammary gland (100%) and clots in milk (86%) was observed in silver ion treated quarters. Mean SCS in milk from quarters having clinical mastitis was reduced (P<0.05) from 9.2±0.89 before silver ion intramammary infusion to 6.7±0.52 at day 7 after silver ion intra-mammary infusion. Total bacterial count in milk from quarters having clinical mastitis was reduced (P<0.05) from 5.7±0.86 at day before to 3.6±1.33 CFU/ml at day 7 of silver ion intra-mammary infusion Mean BSA in milk from quarters having clinical mastitis was reduced (P<0.05) from 3.32±1.04 at day before treatment to 0.55±0.56 mg/ml at day 6 of silver ion intra-mammary infusion. Mean Lf concentration in milk from quarters having clinical mastitis was reduced (P<0.05) from 350.4±180.42 at day before treatment to 142.2±137.25 at day 6 of silver ion intra-mammary infusion. Daily milk yield and Circulating blood cell counts in cows with clinical Mastitis was not affected (P<0.05) by the intra-mammary infusion of silver ions. Key words: silver ion, mastitis 569 Effect of Silver Ion and Staphylococcus aureus-derived Alpha-toxin on Normal Bovine Mammary Gland Epithelial Cell SJ. Kang, TY. Hur, YH. Jung, YJ. Do, HS. Kim National Institute of Animal Science, dairy science division, Cheonan, South Korea Bovine mastitis still represents the biggest hazard faced by the dairy industry worldwide. The economic loss due to bovine mastitis led to the development of various therapeutic strategies. Silver has been well known as a disinfectant and widely used to treat clinical diseases. We acquired mammary tissue from lactating cows and then separated mammary gland epithelial cells using enzyme probe. In this study we have investigated the effects of silver ion and/or S. aureus-derived alpha-toxin, an important virulence factor of bovine mastitis, on primary cultured bovine mammary gland epithelial cells. Silver ion treatment induced mammary gland epithelial cell death at the doses more than 4 ppm which was accompanied by DNA fragmentation and reactive oxygen species (ROS) generation. However, doses less than 2 ppm did not show any effect on DNA fragmentation and ROS generation and thereby did not induce cell death. Further, S. aureuesderived alpha-toxin did not change cell morphology and viability but induced DNA fragmentation, ROS generation and dissipation of mitochondrial transmembrane potential (MTP) at indicated doses. It also showed cytochrome c release without bax activation. In addition, silver ion below 2 ppm inhibited S. aureues-derived alpha-toxin effect on cell death, DNA fragmentation and ROS generation. However at above 4 ppm doses it did not. Taken together, here we demonstrate that silver ion below 2 ppm can inhibit the S. aureus-derived alpha-toxin effect in bovine mammary epithelial cells and thus might be a useful treatment against the bovine mastitis, especially induced by S. aureus. Key words: silver ion, ROS, Alpha toxin, MTP 570 Morphology and Viablility Assessment of Leukocytes in Low Somatic Cell Count Milk D. Mehne , S. Meyer , HJ. Schuberth , H. Zerbe , W. Petzl 1 University of Munic, Clinic for ruminants, Munich, Germany 1 University of Veterinary Medicine Hanover, Foundation, Immunology, Hannover, Germany

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Mastitis / Udder Health and Milk Quality 57

Somatic cell count (SCC) is one of the most common parameters for detection of subclinical mastitis. Udder quarters with SCC <100.000 are generally considered healthy. Differential leukocyte counts of milk cells give insight into the current immune status of an individual udder quarter. Since assays using monoclonal antibodies are time consuming and costly a rapid two stain flow cytometric method was established using acridin orange and propidium iodide for morphological differentiation of viable milk cells. Three groups of 29 healthy heifers (group A: SCC 1.000-19.000; group B: 20.000-49.000; group C: 50.000-100.000) were used. In total 355 milk samples were examined bacteriologically, by Fossomatic® and by flow cytometry (FACScan®). Briefly, three major populations were differentiated morphologically (forward scatter/side scatter): polymorphonuclear granulocytes (PMN), lymphoid cells and MPh/epithelial cells. As a control 210 samples were counterstained with monoclonal antibodies against epitopes on bovine PMN, lymphocyte subsets and MPh/epithelial cells. The percentage of PMN differed between and increased across the three different low somatic cell count groups: group A 38%, group B 47% and group C 62%. Percentage of lymphoid cells decreased with increasing SCC: group A 56%, group B 48% and group C 32%. Differences between the groups were statistically significant (p<0.001). The population of macrophages did not differ and was similar across the three groups (56%).This study shows that the percentage of PMN and lymphoid cells in milk cells are a highly sensitive parameter of early changes in individual udder quarters. Although milk with SCC 100.000 is classified as healthy, increasing amounts of inflammatory cells can be seen already in low SCC milk (<50.000/ml). 573 An Evaluation of Therapeutic Efficacy of Rifaximin in Mastitis Control Program at Drying-off and at Calving, in Two Dairy Herds under Field Condition S. Ávila-Téllez , R. Rivera-Alvarez , J. Ángeles-Manjarrez , 2 3 1 R. Rosiles-Martínez , J. Navarro-Hernández , A. Olguin y bernal 1 Universidad Nacional Autónoma de México, Facultad de Medicina Veterinaria y Zootecnia, Producción Animal:Rumiantes, MEXICO, D.F., Mexico 2 Universidad Nacional Autónoma de México, Facultad de Medicina Veterinaria y Zootecnia, Nutrición Animal, MEXICO, D.F., Mexico 3 Universidad Nacional Autónoma de México, Facultad de Medicina Veterinaria y Zootecnia, Genética y Bioestadística, MEXICO, D.F., Mexico Objective: To determine the effectiveness of intramammary treatment of cows. We analyzed the frequency of sub clinical and clinical mastitis and microorganism occurrence in milk samples at drying-off and calving. Experimental design: From 800 cows in tow dairy herds, 35 cows were selected monthly for a total sample of 105 animals. Cows were th chosen by good general health and being in their 7 month of pregnancy. Four days before drying-off, a milk sample was obtained from each teat, prior to any antibiotic treatment. Samples were cultured for bacteriology and pathogen sensitivity patterns with especial attention to rifaximin. Additionally CMT, total count from the udder (CTU) and MCSC were performed. At drying-off, a gland with CMT: N, T, 1, 2, was treated with rifaximin (100 mg), CMT 3 and blind quarter with 200 mg. Milk samples form the first 5 days after calving were examined for rifaximin by HPLC method. Analysis of results was done by hypothesis contrast to obtain differences between two proportions of positive cases and the value we suggest for clinical mastitis (pcl = 0.1) and sub clinical (pcl= 0.2) with significance a = 0.05 and test power 1 b = 0.95, respectively. Results: At drying-off for sub clinical mastitis, MCSC was 54.29% negatives cows (< 200,000 cells/ml) and 45.71% positive; in CMT 64.76% negatives glands, 34.05% positives and CTU was 47.62% negatives cows and 52.38% positives. At this time we found 1.43% blind quarter. Twelve cows were positive to bacteriology; to Staphylococcus spp (b hemolytic) 8.33%, Streptococcus spp 58.33% and E. coli 33.3%, pathogens susceptible to rifaximin. At calving the MCSC was 77.14% negative cows to sub clinical mastitis and, 20% were positive. Per CMT, 90.47% glands resulted negative and 7.86% positives. For CTU 82.86% negatives cows and 13.33% positive. The analysis of results by hypothesis contrast for the two proportions resulted lower than the hypothetical value; which was 10% for clinical and 20% for sub clinical mastitis; showing statistical significance p< 0.05. From the bacteriological cultures in 8/420 glands, two cases of Staphylococcus spp were isolated, five Streptococcus spp and one E. coli. From de six blind quarters three had colostrums at calving. At calving milk samples examined by HPLC for rifaximin were negative.

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Conclusion: Sub clinical mastitis control by Rifaximin resulted effective at 90%, and milk samples examined by HPLC for rifaximin were negative. 574 Evaluation of Four Fluoroquinolone Treatment Regimens for Acute Bovine Mastitis R. Froyman , S. Filios , B. Brunner , N. Brunner , A. Torres , 2 K. Hellmann 1 Bayer HealthCare, Animal Health Division, 51368 Leverkusen, Germany 2 Klifovet, 80689 Munich, Germany In a multicenter GCP trial, comparing 4 intravenous (IV) fluoroquinolone (FQ) treatments, 173 cows with acute mastitis (temperature [T] >39.6 °C), were enrolled at day (d) 0 (first treatment) until ~100 bacteriologically positive cases were obtained. Clinical criteria were assessed at d0, d0+8h and then daily until study end (d5), with clinical cure defined as T <39.0 °C. Udder inspection was done at d0-5 as well as a milk sample taken for culture. Cows of group (G) 1 (n= 37) and 2 (n= 48) received one single and two daily injections, resp., of 5 mg enrofloxacin (ENR)/kg (Baytril 10%). Cows in G3 (n= 43) were given one injection of 6 mg danofloxacin (DAN)/kg (Advocid 180) whereas those in G4 (n= 45) received 3 daily injections of 2 mg marbofloxacin (MAR)/kg (Marbocyl 10%). In all groups, except G1, the products were used per label. If poor improvement was observed with a single injection in G1 (ENR) and G3 (DAN), a second dose was given at 24 and 48 h after first dosing, resp. In cows with T >39.5 °C at d1, IV meloxicam treatment was given (0.5 mg /kg). Statistical analysis comprised comparisons between ENR and other FQ treatments. From 173 cows treated, 108 were bacteriologically positive, thereof 55% for E. coli and 23% for S. uberis. Across treatments bacteriological cure rate was 79% for coliform bacteria and 54% for S. uberis. Redosing in G1 and G3 was required for 22% and 37% of cows, resp. Fever decreased rapidly in all animals within 8 h. General illness score dropped from 6.4 to <1.0 at d2 in G2, at d3 in G1 and G4, and at d5 in G3. Mean mastitis score (MMS), high at inclusion (d7), fell below 3 at d3 for G2, at d4 for G1 and G4, and at d5 for G3. At d0+8h clinical cure rate (CCR) was better (p= 0.0025) in ENR treated cows (61%) than in MAR treated cows (33%). At 24 h CCR was 58% for MAR, 75% for ENR and 71% for DAN treated cows. At 48 h CCR was better for ENR (90%) (p= 0.006) than for DAN (68%). At d5 CCRs were 91%, 93%, 84%, and 95% in G1, G2, G3 and G4, resp.; overall mastitis cure rates (clinically cured and MMS <1) were 54%, 69%, 51% and 60%, resp. Mean milk yield at d5 compared to d0 was increased by 79% in ENR, 60% in DAN and 43% in MAR treated cows. A dedicated study would be needed to substantiate whether a single ENR dose with optional redosing 24 h later is as efficacious as 2 daily doses. All in all ENR showed most rapid clinical cure among FQ treatments and is a preferred therapy for acute mastitis with generalised disease. 575 Virulotyping and Genetic Comparison of Strains of Staphylococcus aureus Associated with Bovine Mastitis B. Taminiau, I. Dizier, J. Mainil University of Liege, Vetrinary Faculty, Liege, Belgium Bacterial infection of the udder (=mastitis) is the most frequent and prejudicial cause of the reduction of milk production and quality in the bovine dairy industry. Clinical and sub-clinical mastitis, resulting in macroscopic or microscopic alteration of the milk respectively, can be caused by many different Gram positive bacterial species (streptococci, staphylococci, corynebacteria). Up to date, there exist no real efficacious vaccines against mastitis, partly as a consequence of a lack of knowledge about the properties of the bacterial pathogens. The species Staphylococcus aureus is one of the major bacterial pathogens of the udder and is responsible for both clinical and sub-clinical mastitis. In order to select putative immunogens entering into the composition of a vaccine, almost 300 strains of S. aureus isolated from cases of bovine mastitis have been collected in Belgium during the last few years. They have been assayed for the presence of 25 virulence factors by PCR targeting genes coding for the following virulenceassociated factors: adherence factors (ClfA, ClfB, Can, IcaA, FnbA, SdrC and EbpS), toxins (Enterotoxines, TssT-1, Panton-Valentine leukotoxin, exfoliating toxins and leukocidin), exoenzymes (serine proteases, aureolysin, haemolysins) and surface antigens (protein A, capsular antigens). Summary tables with the frequency of the different

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58 XXV. Jubilee World Buiatrics Congress 2008

virulence factors and virulotypes will be presented. More than 50 virulotypes were observed when the results were concatenated. Strains belonging to the most two frequent virulotypes (9 and 4%) were positive for the type 8 capsular antigen and for ClfA/B, FnbA, EbpS, SdrC, Spa, HlA/B/D/G, SspA and lukD/E/M, but differed by the collagen adhesin and the intercellular adherence factor. Twenty-five strains were further compared by the multi-locus sequence analysis (MLST) based upon 7 housekeeping genes: 12 strains belonged to the clonal complexes 97 and 133, but the other strains belonged to various sequence types. Though the clonal complex 133 was related to one the most frequent virulotype, the clonal complex 97 comprised strains belonging to different virulotypes. Based on this limited number of strains tested, the clonal relatedness of S. aureus strains based on MLST analysis can not be transposed to the classification into virulotypes. Results of similar studies in other countries will help to choose the immunogens to incorporate into the vaccines. Key words: Mastitis, cattle, S. aureus, virulotyping, Multilocus Sequence Analysis 576 The Effect of Fermentated-herb Extract on Subclinical Mastitis in Dairy Cows TY. Hur , SJ. Kang , YH. Jung , YJ. Do , CY. Choi , IS. Ryu , 1 1 1 2 HS. Kim , YS. Lee , SJ. Park , GH. Suh 1 National Institue of Animal Science, Animal Resources Development, Cheonan, South Korea 2 Chonnam National University, College of Veterinary Medicine, Gwangju, South Korea Therapeutical potential of fermentated-herb extract (FHE) in bovine subclinical mastitis was investigated. The fermentated-herb extract was made by the process of aqueous extract of twelve herbs inoculated with yeast and probiotic to provide feed additive. Somatic cell count (SCC), milk yields and total bacterial count (TBC) were evaluated after fifty grams of the FHE fed each cows per day for 4 weeks. The results revealed that the FHE reduced the TBC and SCC, and increased milk yields. The results suggest that the components of FHE possesses some biologically activity in bovine subclinical mastitis. As such, the present work substantiates the therapeutic use of fermentated-herb extract and also emphasizes on the potential of the commonly available non-toxic substance to enhance the cow health. However, more experiments are necessary for confirmation of these results. Key words: mastitis, fermentated herb, somatic cell count 577 Prognosticable Fatal Outcomes of Naturally Occurring Escherichia coli Mastitis in Dairy Cows ­ Consumption Coagulopathy, Hemoconcentration and Hepatobiliary Damage K. Mori , S. Hagiwara , H. Okada , H. Nagahata 1 Engaruchiku NOSAI, Clinic of Farm Animals, Engaru, Japan 2 Hokkaido NOSAI, Livestock Insurance, Sapporo, Japan 3 National Institute of Animal Health, Research Team for Prion Diseases, Tsukuba, Japan 4 Rakuno Gakuen University, Department of Animal Health, Ebetsu, Japan Objectives: To determine diagnostic characteristics capable of prognosticating the fatal outcomes of naturally occurring Escherichia coli mastitis in dairy cows. Materials and Methods: Vital signs (temperature, pulse, respiration) were measured, and blood tests, including white blood cell (WBC) count, antithrombin (AT) activity, platelet (Plt) count, hematocrit (Ht) value,-glutamyl transpeptidase (GGT) activity and non-esterified fatty acid (NEFA) levels, were conducted for all the cows. These values were compared between 7 cows that died due to E. coli mastitis and 17 cows that survived. Five blood test parameters (AT, Plt, Ht, GGT and NEFA) that showed significant differences between the dead and surviving cows were used to determine the cut-off values; these parameters would also be used as criteria to differentiate between the outcomes. The diagnostic criterion to prognosticate the fatal outcome was decided using the cut-off values. The practicability of the diagnostic criterion was verified by trial use on 51 other clinical cases (12 dead and 39 surviving cows). Results: All the values measured within 12 hours of onset (day 1) did not vary between the dead cows and those that survived. However, significant differences in the five parameters (AT, Plt, Ht, GGT and

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NEFA) were observed from the second day (day 2). The diagnostic criterion for fatal prognosis was determined using the cut-off values of the above-mentioned parameters. The fatal outcomes could be predicted if at least two of these parameters were within the ranges 4 defined by their cut-off values (AT < 120%, Plt < 15 x 10 /ml, Ht 32%, GGT 45 IU/l and NEFA > 0.6 mEq/l). The sensitivity, specificity, predictive value and likelihood ratio of the diagnostic criterion on day 2 were 100%, 76%, 64% and 4.25, respectively. Verification of the practicability of the diagnostic criterion in the 51 other clinical cases examined on day 2 showed the sensitivity, specificity, predictive value and likelihood ratio to be 67%, 69%, 40% and 2.16, respectively. Conclusions: Consumption coagulopathy (low AT and Plt), hemoconcentration (high Ht) and hepatobiliary damage (high GGT and NEFA) were determined as the diagnostic characteristics in the fatal cases. AT, Plt, Ht, GGT and NEFA values can be used as criteria for the prognosis of fatal E. coli mastitis. 578 Pseudomonas Mastitis: Elimination from Contaminated Wash-water Systems by Slightly Acidic Electrolyzed Water K. Kawai , M. Kitou , T. Mitamura , I. Uchida , K. Hirose , 5 H. Nagahata 1 Seibu Veterinary Clinical Center, Tokachi NOSAI, Shimizu, Japan 2 Ecolofrontier Co., Ltd., Sapporo, Japan 3 Hokkaido Research Station, National Institute of Animal Health, Sapporo, Japan 4 Clinical Research Center, MeiJi Seika Kaisha Ltd., Yokohama, Japan 5 School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Japan Objectives of study: Pseudomonas aeruginosa (PA) is usually responsible for less than 1% of the mastitis infections in dairy cows. The source of infection for PA mastitis is contaminated water used to wash the udder or when antibiotics used for intramammary infusion have become contaminated. PA mastitis infrequently causes clinical mastitis and their spread of infection inflicts damage on the farms. In this study, we present effective mastitis control in dairy herd with increased incidence of PA mastitis. Materials and Methods: 387 dairy cows were milked in the parlor. The preventing measures of PA mastitis were started since Oct. 2005. Milk samples of clinical cases were obtained for bacteriologic culture, and isolated bacteria were identified using standard techniques. Liners, jeters, drop hoses and wall of cistern in the parlor were wiped with cotton swabs and cultured. Milking procedure was checked and changed to the recommended procedure. The chlorine concentration of water was increased from 0.05 ppm to 0.5 ppm. Milking equipment and floor of parlor were sterilized by 200 ppm sodium hypochlorite. The cows infected with PA were isolated and milked at the end. The milk samples of all cows were cultured after calving and before dry period. Slightly acidic electrolyzed water system (Purester MP240E: Morinaga Milk Industry Co., Ltd., Tokyo) was introduced to the parlor and was used at milking time since Jan. 2006. Results: The occurrences of PA clinical mastitis were 35cows in 2003 and 45cows in 2004. After implementation of the preventing measures, the occurrence of PA clinical cases was 8.3% on all clinical mastitis in 2005. P. aeruginasa, P. fluorescences and P. putida were isolated from environment (eg, liners, jeters, drop hoses, wall of cistern in the parlor and reservoir in free stall), and same pathogens were isolated from clinical cases and 23 of nonclinical cases. However, after the use of slightly acidic electrolyzed water (hypochlorous acid: available chlorine, 12 ppm) in 2006, Pseudomonas sp. in the environment, the occurrence of PA clinical mastitis and the reservoir of Pseudomonas sp. in the nonclinical cases were disappeared. Conclusions: The use of slightly acidic electrolyzed water at milking time is beneficial in reducing exposure of the cows to the organism and it was proved to be effective on prevention of PA mastitis. Key words: mastitis, P. aeruginosa, slightly acidic electrolyzed water 579 Acute Clinically Mastitic Animals in Some Assiut Governorate Villages: Diagnosis and Treatment M. Sayed , A. Abd El-Rady 1 Faculty of Veterinary Medicine, Assiut University, Food Hygiene, Assiut, Egypt 2 Faculty of Veterinary Medicine, Assiut University, Animal Medicine, Assiut, Egypt

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This investigation was run in some villages in Assiut governorate during summer 2007 to diagnose acute clinical mastitis in different animal species and to apply different lines of treatment for evaluating which treatment line of choice giving cure, aiming to another goal by preventing the conversion of acute mastitis towards the chronic one which is difficult to be treated and the dairy animal will be excluded. Therefore, 2150 animals were clinically examined in 5 villages located north to Assiut city, Egypt, including 400 cows, 950 ewes and 800 she goats, and the incidence of acute clinical mastitis was 22.50, 2.63 and 4.63%, respectively. Milk samples were collected from all clinically mastitic cows for bacteriological examination to identify the causative agents of the intra-mammary infection (IMI). It was found the major causative agents isolated were Staphylococcus aureus, Streptococcus agalactiae, Escherichia coli and Corynebacterium pyogenes, causing either single or mixed type of infection. When applying different lines of treatment, all diseased animals were classified into st 3 groups: 1 group received local treatment by intra-mammary nd infusion antibiotic. 2 group received systematic treatment by intra-muscular (I/M) injection of both antibiotic and antiinflammatory drugs. While, 3rd group received combination of both local and systematic treatment lines together. Cure% was achieved rd as 50% for 1st group, 90% for 2nd group, while 3 group gave complete cure by 100%. It was noticed that the incidence of acute clinical mastitis among examined cows was worrisome and can be considered as indicator of the epidemiology of the disease. While, spreading of the disease among ewes and she goats was somewhat low in comparison with that of cows. In conclusion, combination of both local and systematic treatment lines together should be advised in treatment of acute clinical mastitis to ensure complete cure. The obtained results highlighted the focus towards the spreading of acute clinical mastitis among cows lived in some Assiut governorate villages. 580 Efficacy of Extended Therapy of Staphylococcus aureus with Intramammary Cefuroxime J. Spatz Shelgren, K. Parker, S. McDougall Animal Health Centre, Research Division, Hamilton, New Zealand Bacteriological cure following antibiotic therapy of Staphylococcus aureus during lactation results in cure rates averaging between 2535%, but with a range of between 4 and 92%. Increasing the duration of therapy increases cure proportion using beta-lactams and lincosamides. However, there no data on the efficacy of extended therapy with a cephalosporin is available.Cows (n=61) with a SCC of >200,000 were examined with a California Mastitis Test (CMT) and those glands with a CMT >1 were aseptically sampled for bacteriological culture. Cows (n=34) from which 1 or more glands were culture positive for S. aureus (total n=55 glands) were blocked by number of glands within cow with S. aureus then ranked by SCC before being assigned randomly within blocks of 3 to no, 3 or 6 intramammary infusions at 12 h intervals with 250 mg cefuroxime sodium (Spectrazol Milking Cow, Schering-Plough Animal Health, Upper Hutt, New Zealand). Glands were re-sampled at 41 and 48 days after the first treatment. Cure was defined as having occurred where S. aureus was isolated from neither of the both post treatment samples. The probability of cure was modeled with main effect being treatment (0, 3 or 6 tubes) on cure at quarter level (yes/no). To account for the non-independence of gland within cow, the confidence intervals were adjusted by the variance inflation factor (pscale; Proc GENMOD, SAS 9.1). A log link function was used as exponentiation of the coefficients produces relative risks. The cow-level log10 SCC and milk yield 43 days post treatment were analysed using ANOVA with treatment as the main effect.The least square mean (95% confidence intervals) cure proportions were 0.13 (0.03-0.53), 0.24 (0.10-0.56) and 0.53 (0.320.87) for glands treated with 0, 3 or 6 tubes respectively. The cure proportion tended to be higher following treatment with 6 tubes compared to 0 tubes (RR = 4.2 (0.9-19.7); p=0.07) and for 6 tubes compared to 3 tubes (RR = 2.2 (0.8-6.0); p=0.11). There was no difference in log10 SCC or milk production at the herd test 43 days post-treatment among the treatment groups (both p>0.3).It is concluded that increasing the duration of therapy with the second generation cephalosporin cefuroxime from 0 to 3 and 6 tubes resulted in a tendency for a higher bacteriological cure rate in glands infected with S. aureus.

581 Relationship between Somatic Cell Count (SCC) and Bacterial Infection of Goat Milk: 1st Report of Caprine Subclinical Mastitis from Iran H. Ghasemzadeh-Nava , S. Bokaee , P. Tajik 1 Faculty of Veterinary Medicine, University of Tehran, Department of Clinical Sciences, Tehran, Iran 2 Faculty of Veterinary Medicine, University of Tehran, Epidemiology, Tehran, Iran The SCC is a good indicator for milk quality and can be implicated as a quality character of an animal. An increased SCC is either the consequence of an inflammatory process due to the presence of an intramammary infection (IMI) or under non-pathological conditions. For this reason the SCC of milk represents a sensitive marker of the health of the udder and is considered a useful parameter to evaluate the relationship between IMI and changes in milk characteristics.The aims of our study were (a) to determine the occurrence rate and etiology of subclinical mastitis in native meat-type breed goats in Garmsar (a city in Semnan province in Iran) and (b) to evaluate threshold of SCC for bacteriologically negative and positive goat milk. Goats were restrained in a sitting position and the teat end of each half udder was scrubbed thoroughly using cotton wool soaked in 70% ethyl alcohol. The three first streams were discarded, and 10ml milk samples were taken in a sterile tube held horizontally. The tube was capped and labeled, then placed on ice and sent to the laboratory for bacteriology and SCC within 8 h. Of the 300 milk samples collected, 24 (8%) udder halves were bacteriologically positive. The most frequent isolate from subclinical cases was cougulase negative staphylococci (90.2% of isolates). The other isolate was Pseudomonas aer. In bacteriologically positive samples, the lowest and the highest of goat milk SCC were 6 6 1.1x10 and 0.5x10 cells/ml respectively in which the mean of it was 6 0.75x10 cells/ml. In bacteriologically negative samples, the lowest 6 6 and the highest of goat milk SCC were 0.7x10 and 0.025x10 cells/ml 6 respectively in which the mean of it was 0.025x10 cells/ml. The 6 threshold of SCC for negative and positive goat milk was 0.5x10 cells/ml in our study. Isolation of bacteria was associated with an increased SCC of goat milk in our study. There is not any report pertaining to caprine subclinical mastitis in different breeds from Iran. Cougulase negative staphylococci have been considered to be the major cause of non-clinical intramammary infections in a number of previous investigations. Due to different pathological and nonpathological factors contribution to change SCC in milk of goats, a complete study to take into consideration of different factors is needed. Key words: mastitis, goat, SCC, bacteriology 582 Comparing the Efficacy of Copan and Delvotest to Detect of Antibiotic Residues in Bulk Tank Milk Samples of Garmsar Dairy Farms M. MOhammadsadegh , S. Bokaii , T. Mosavi , E. Abbaszadeh 1 Islamic Azad University, Garmsar branch, Large Animal Clinic for Theriogenology and Ambulatory Services, Garmsar, Iran 2 University of Tehran, Epidemiology, Tehran, Iran 3 Islamic Azad University, Garmsar branch, Food Hygiene, Garmsar, Iran 4 Islamic Azad University, Garmsar branch, Large Animal Clinic for Theriogenology and Ambulatory Services, Garmsar, Iran Antibiotic residues in milk are from mastitis treatment with intramammary infusions of MC ointments an /or with systemic therapy in many diseases. Milk antibiotic residues have a hazardous effects on milk industry and milk consumer welfare. In order to compare Delvo test with Copan kit in field of evaluating antibiotic residue in milk, 100 bulk tank samples of collected milk were prepared in the milk collecting center of Garmsar. Antibiotic residues were analyzed by Copan and Delvo test, and total bacterial counts were determined in all samples. Results showed that antibiotic residue were present in 19% of samples. Nine present of positive samples were found by Delvo test and 10% by the two Delvo test and Copan, consequently. Total bacterial count in only Delvo test positive samples were 24944/44 ­ 1710/35, and in Delvo test and Copan positive were 13160 ­ 2061/6. Statistical analysis showed that substantial agreement of the two test was acceptable (kappa = 0.643). It is concluded that Delvo test is a significantly susceptible test, at least in present condition of this study. Key words: Delvo test. Copan test, antibiotic residues in milk, Garmsar

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583 An Integrated Approach to Production Process and Education in Dairy Cattle Farming: the American Farm School E. Souglis , G. Arsenos , G. Valergakis , A. Giamoustaris 1 American Farm School, Thessaloniki, Greece 2 Aristotle University of Thessaloniki, Animal production, Thessaloniki, Greece

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University of Helsinki, Faculty of Vet. Medicine, Department of Production Animal Medicine, Helsinki, Finland 2 Finnish Food Safety Authority Evira, Helsinki, Finland Objectives of the study: Bacteriological diagnosis of bovine mastitis determines treatment decisions and is important for herd health programmes. In Finland, farmers can send milk samples to laboratories or in cases of acute mastitis, the practicing veterinarian often cultures the milk sample. The aim of this study was to evaluate the accuracy of bacteriological diagnostics carried out by practicing veterinarians. Materials and methods: Thirty veterinarians volunteered to participate in the study. Autoclaved milk was inoculated with dilutions of mastitis causing bacteria. Samples were sent to the participating veterinarians, who were asked to culture the samples according to their normal routines. A questionnaire on diagnostic methods and quality control systems used by the veterinarians was included. Bacteriological diagnoses of the milk samples considered to be correct were A. K. oxytoca (Klebsiella sp.); B. Str. uberis and A. pyogenes; C. S. simulans (coagulase-negative staphylococci, CNS); and D. E. faecalis (Enterococcus sp). The diagnoses were scored as correct, partly correct and incorrect. The diagnoses were scored as partly correct if only bacterial genus was identified. Results: For sample A, the diagnoses were correct in 22 out of 28 cases, partly correct in 2 and incorrect in 4 cases. For sample B, the diagnoses were partly correct in 18 out of 28 cases, and incorrect in 10 cases. Different Streptococcus species were mentioned and A. pyogenes was not recognised. For sample C, diagnoses were correct in 26 out of 28 cases and partly correct in 2 cases, and for sample D, correct in 24 out of 28 cases and incorrect in 4 cases. The two most commonly used agars were diagnostic triplate Mastix®-agar (used by 64% of the veterinarians) and blood-esculin-agar (21%). Additional tests included Gram-staining, rapid coagulase test, tube coagulase-test, and various tests for antibiotic sensitivity. Quality control procedures were seldom used, but included monitoring the temperature in the incubating closet and noting the expiry date of agars. Conclusions: Samples with CNS were most frequently diagnosed correctly. Sample B, comprising Str. uberis and A. pyogenes, caused most problems. Slow growth and small colonies of A. pyogenes remained undetected and the Mastix®-agar was not suitable for identifying the various Streptococcus species. Small field laboratories run by the veterinarians would need guidelines for implementing quality control. 586 Mycoplasma bovis Outbreak in a Dairy Herd: Consequences on Production and Reproduction L. Pinho , I. Maia , N. Canada , C. Azevedo , M. Machado , 1 1 1,4 E. Silva , G. Thompson , J. Carvalheira 1 Universidade do Porto - ICBAS, Department of Clinical Sciences, Porto, Portugal 2 Medicina da Produçio Leiteira Veterinária, Tocha, Portugal 3 Laboratório Nacional de Investigaçio Veterinária, Bacteriology, Vila do Conde, Portugal 4 Universidade de Porto, Centro de Investigaçio em Biodiversidade e Recursos Genéticos, Vila do Conde, Portugal The objective of this report was to document an outbreak of M. bovis in dairy cattle. Mastitis was the major pathology, but arthritis and pneumonia were also observed, including a decrease in reproductive efficiency. The outbreak occurred in a dairy farm with 160 lactating cows in northwest Portugal. To determine the cause of the mastitis outbreak, individual and bulk tank milk samples were obtained between September 2006 and April 2007. Milk samples were inoculated in modified Hayflick Agar and enrichment solution. Standard cultures for other bacteria detection were also performed. Mycoplasma species were identified according to biochemical characteristics and PCR. M. bovis was isolated from 13 of the 48 individual milk samples collected. At the peak of the outbreak, the rate of clinical mastitis exceeded 10 cases per month, with the majority of cows presenting elevated SCC and decreased milk yield that persisted even in clinically recovered animals. Multiple quarters were affected with little or no response to antibiotic therapy. Atrophy of affected quarters or complete destruction of the gland, was also observed. Prophylactic measures included segregation and culling of the positive animals, introduction of iodine based pre and post-dipping and the use of single towels. Treatment was changed to single disposable intramammary tubes instead of systemic antibiotics given by intramammary route. The

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The quality of dairy products has become an issue for consumers over the last decades due to various outbreaks of food-borne zoonoses. Moreover, consumer concern about production methodologies and the way that dairy cows are being kept is increasing. Hence, quality assurance programmes have been implemented in dairy farms to counteract the latter concerns. The objective of this study was to assess the quality assurance program implemented by the American Farm School of Thessaloniki (AFS). The AFS was chosen because it is a small dairy enterprise that first introduced pasteurised milk to Greece in 1935. Since then, it has been the industry leader in productivity and quality of milk serving also as an education centre in cow herd management and milk processing for farmers and students. The AFS dairy herd comprised of 124 Holstein cows reared in a freestall barn. All the cows were included in this study. Data regarding milk quality parameters, disease occurrence and management in relation to health and productivity were collected over the last three years. More particularly, Somatic cell counts (SCC), Total Bacterial Counts (TBC), fat and protein content of milk were measured in individual cows on a monthly basis. Milk yield and reasons for culling were assessed every lactation, whereas the cost of particular diseases was estimated. The results revealed relatively low SCC (mean: 253,628, S.E:19,022), and TBC (51,214, S.E: 13,032). Protein content of milk was similar (3.26, 3.28, 3.21, s.e.d. 0.044) whereas there was a significant increase in fat content (3.96, 4.26, 4.09, s.e.d 0.082) over the three lactations, respectively. The average replacement rate was 24% and apart from age, culling was associated with feet and reproduction problems. It is concluded that based on developments within the dairy sector in Greece and neighbour countries such practices on dairy farms will be a common practice in the near future and the AFS could disseminate such knowledge though its educational program. The later is likely to help dairy farmers identifying and managing the quality hazards and risks occurring in the production process, and hence providing the consumer with more certainty about the welfare of dairy cows and the quality their milk at retail. 584 Apoptosis and Autophagy in Involuting Bovine Mammary Gland ­ the Role of TGF-beta1 and Somatotropic Pathway Proteins J. Zarzynska , B. Gajkowska , U. Wojewódzka , T. Motyl 1 Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Department of Food Hygiene and Public Health, Warsaw, Poland 2 Polish Academy of Science, Medical Research Center, Warsaw, Poland 3 Faculty of Veterinary Medicine, Warsaw University of Life Sciences - SGGW, Department of Physiological Sciences, Warsaw, Poland The mechanism of secretory tissue involution in bovine mammary gland is still poorly understood. We can find many factors, which could regulate this process. It has been shown that expression of TGF2-a1 and its receptors in mammary epithelial cells (MEC) increases during mammary gland involution in mouse, goat and sow. It has been also shown that TGF2-a1 is an antiproliferative and apoptotic factor in bovine MEC. Our recent study revealed that expression of TGF2-a1 in bovine MEC undergoes complex endocrine and auto/paracrine regulation by hormones of the somatotropic axis. Similarly to apoptosis, the intensity of autophagy was the highest in the dry period, as shown by increased expression of Beclin1 and morphological features. Autophagy observed in the involuting mammary tissue could be the natural cell defense against transient undernourishment and action of apoptogenic peptides (e.g. TGF2-a1, IGFBPs), thus maintaining cellular homeostasis in the dry period. Key words: TGF2-a1, mammary gland, apoptosis, autophagy 585 A Pilot Study on the Bacteriological Diagnostics of Bovine Mastitis Carried Out by Finnish Veterinarians H. Simojoki , S. Erkkilä , A. Pitkälä , S. Pyörälä

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reproductive data showed that cows found positive to mycoplasma had lower reproductive efficiency. The percentage of cows with 3 or more AI per conception was higher (82%) for infected cows. The same pattern was observed on other parameters such as days-open and metritis incidence. The rate of pneumonic infections was highest after the onset of the outbreak. Standard treatments were inefficient. Arthritis was also diagnosed in some cows showing enlarged joints and lameness. There was an increase of SCC between September and November in positive cows. During those 3 months 67%, 83% and 100% of the infected cows had SCC above 200,000 cells/ml with an average higher than 1,200,000 cells/ml. The production level of the herd decreased from 26.5 kg in August to 22.6 kg of milk/day/cow in November. This report confirmed for the first time the association of M. bovis with clinical and subclinical mastitis in dairy cattle in Portugal. These results suggest that M. bovis should be considered in the differential diagnosis of mastitis in Portuguese dairy farms. Key words: mycoplasma bovis, dairy cattle, mastitis, arthritis, pneumonia 587 Bulk Milk Tank Staphylococcus aureus UCF Count as Predictive Datum for Bovine Herd's Infection Level G. Varisco , L. Bertocchi , N. Contessi , M. Tranquillo , G. Bolzoni 1 Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna Brescia, Brescia, Italy 2 Centro Miglioramento Latte, Brescia, Italy

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evenly over the bacterial suspension and allowed to remain on the bacterial suspension for 30 minutes. The bedding treatment applied 2 (1g/100cm ) was consistent with the prescribed amount indicated on the product label directions. Then the bedding treatment was swept off of the cow mat and the amount of bacteria in the bedding treatment was determined. A control experiment (0.5 g of a log 10.11 CFU/ml suspension allowed to sit at 220C on a cow mat for 30 minutes) was conducted to see if the suspension could be swept off of the mat in the absence of bedding treatment. Numbers of bacteria were determined by the Standard Plate Count Method. Results: The control experiment showed that attempting to sweep a bacterial suspension off of a cow mat in the absence of bedding treatment did not remove any of the bacterial suspension from the cow mat. Sweeping off the bedding treatment from the cow mat removed more than 99.99999% (7 log) of the bacteria from the cow mat after both 30 seconds and 30 minutes contact time of the suspension with the cow mat. After 30 seconds of contact time, 99.9% (10 log) of the bacteria were removed by the bedding treatment. After 30 minutes of contact time, 99.9999945% (> 7 log) were removed by the bedding treatment. Conclusions: The results indicate that applying DryMaxxTM bedding treatment to the bacterial suspension facilitated the removal of the bacterial suspension from the cow mat. We suggest applying DeLaval bedding treatment to DeLaval cow mats, letting the bedding treatment remain on the cow mat for at least 30 minutes, and then sweeping off the bedding treatment and removing it from potential future contact with the cow. 589 Evaluating the Teat Condition Performances of a New Teat Spray in an Automatic Milking Environment X. Goossens , L. Bommelé , W. Ingalls , T. Hemling 1 DeLaval, PP Milk Quality & Animal Health, Gent, Belgium 2 Ghent University, Biocenter Agri-Vet, Melle, Belgium 3 DeLaval, PP Milk Quality & Animal Health, Kansas City, United States Automatic milking is a growing practice in dairy farming. In an automatic milking system the cows are milked more frequently and the variation in the interval between milkings is greater than with conventional milking. As the automatic milking creates a new and different management and environment with different challenges for the teats, new teat dips with the objective to maintain good teat condition when sprayed in an automatic milking system should be tested in the system itself. Results abstracted from conventional milking parlours cannot be the only basis to evaluate teat dips in this new way of milking. Teat condition studies done in conventional milking parlours, use a split udder or split herd design. In the conventional milking parlours this dipping or spraying is done by human interaction and can be done selectively with the right product on the correct teats. In automatic milking systems, teat spraying is done automatically. Technically it is not possible to spray the teats of different cows with different products. And because of the spraying it is not possible to spray only two teats of each cow without hitting the two control teats. This makes it impossible to do a side by side control trial in a robotic system. The objective of this study was to evaluate a new teat spray for an automatic milking system in a commercial dairy farm environment with a positive control. To evaluate teat condition performance of a new peroxide teat spray (DeLaval Prima) in an automatic milking system we made use of a switch back design (Rasmussen et al. 2001). Control product was Delaval Proactive Plus, a teat dip with well know teat condition performance. Before the start of the trial, the teats had already been sprayed for a long period with the control product. During the trial period of 18 weeks, the experimental product and the control product were switched every 6 weeks. At the start and every two weeks during the trial period, teat skin condition, teat end condition and orifice thickness were scored by the same trained person using an ordinal scale. Every two weeks individual SCCs were recorded. Weather data were gathered throughout the trial. The switch back design should help to evaluate the teat condition performances of a new teat spray in an automatic milking environment with one robot. 590 Prevalence of Intramammary Infection in Pregnant Heifers M. Vasil, J. Bires University of Veterinary Medicine, Animal Husbandry and Veterinary Science, Kosice, Slovakia Examination carried out in 3 farms on 110 heifers. Clinical examination of the udder and bacteriological tests of quarter secretion

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Aim of this work is to verify the predictivity of bulk milk tank (BMT) analysis as simple tool for epidemiological surveillance system on herd's Staphylococcus aureus infection level. The comparison between BMT analysis results and single cow milk (SCM) analysis results was investigated to verify the relationship between BMT Staph. aureus UFC value and dairy herds infection rate. During 2005, 47.208 cow milk samples were analysed (7.166 positive, 38.530 negative and 1.512 to much polluted)) of 31 dairy herds (approximately 4.000 cows were checked). Staph. aureus count in BMT (UFC) was found to decrease during time, due to dairy Staph. aureus infection rate reduction at cow level. Statistical analysis shows that within herds the prevalence is 1.28 grater (exp 0.25) every one UFC log unit (p<0.001). UFC variable did not show normal distribution therefore it was been transformed in log. the model allow to calculate odds log for an infected herd. a) log (odds) = 2,829 + 0,25 * (UFC) To obtain prevalence value the following formula is to be used: b) Prevalence= Odds/1+Odds where odds value is obtained using natural value formula a) result Ex: BMT resutl is 3UFC (Staph. aureus) Log(odds)= -2.829+0.25*(3)= -2.079 natural vale: exp.079)= 0.1250 prevalence= 0.1250/(1+0.1250)= 0.107 (10.7%) In conclusion: ­ results obtained confirm the significative statistical relationship between UFC, in log, and cows' infection rate; ­ BMT analysis is a valid tool to evaluate dairy herds infection prevalence; this model is unable to describe all the prevalence variability. ­ Although this high statistical relationship between UFC log and prevalence, the model predictivity performance is to improve and to use Staph. aureus UFC count in BMT in routine UFC as prediction of cow's infection level within herds further study have to be run. 588 Investigation of Bedding Treatment Ability to Remove Escherichia coli Suspensions from Rubber Cow Mats R. Reinbold , T. Hemling , X. Goossens 1 DeLaval, PP Milk Quality & Animal Health, Kansas City, United States 2 DeLaval, PP Milk Quality & Animal Health, Gent, Belgium Objective: Escherichia coli are incriminated in mastitic and systemic infection in dairy cattle and therefore the exclusion of this organism from contact with the cow can be of great economic importance. We conducted studies to determine if an absorptive DeLaval bedding treatment would facilitate removal of Escherichia coli suspensions from DeLaval rubber cow mats. Materials and Methods: Rubber cow mats (DeLaval Pen Mat PM30P) were cut into 10 x 10 cm squares. Escherichia coli suspensions (0.5 g of a log 10.46 CFU/ml suspension) were spread evenly on the mats. The bacterial suspension was allowed to remain on the mat at 220C for 30 seconds or 30 minutes and then 1 g of the absorptive bedding treatment (DryMaxxTM, DeLaval), was distributed

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62 XXV. Jubilee World Buiatrics Congress 2008

were performed between the 4 and 2 week before parturition. The sings of clinical mastitis were found in udder of 22 heifers (20%). Intramammary infections caused by 15 species of microorganisms were found in 140 quarters (32.1%) of 78 pregnant heifers (70.9%). Arcanobacterium pyogenes, Streptococcus agalactiae, Streptococcus uberis, Streptococcus dysgalactiae, Escherichia coli, Staphylococcus aureus and coagulase-negative staphylococci (S. hyicus, S.. epidermidis, S. simulans, S. cohnii, S. xylosus, S. intermedius, S. haemolyticus) were most frequently isolated. Key words: mastitis, heifers, intramammary infection 591 Concentration of Ascorbic Acid in Blood of Cows Affected with Clinical Mastitis before and after Treatment Using Cobactan M. Kleczkowski, W. Klucinski, T. Jakubowski, K. Dembele Warsaw University of Life Sciences, Faculty of Veterinary Medicine, Clinical Sciences, Warsaw, Poland The aim of the investigations was evaluation of AA concentration in the cows serum with clinical mastitis caused by Staphylococcus aureus before and after treatment using Cobactan. The experiment was conducted on 30 cows, Black-White breed, at the age of 3-8 years. The animals were divided into two groups (A-experimental and B-control). All cows were subjected clinical examination, evaluation of macroscopic changes of milk, and evaluation of somatic cell count. Milk samples, before treatment, were taken simultaneously for identifying pathogenic micro-organisms. All cows from group A were given Cobactan LC during three days, at 24-hour intervals. All cows from group A were also treated intramusculary with Cobactan once daily, for 3 days. Blood samples from all cows (group A and B) were st collected from the external jugular vein 9 times: 1 -before giving a nd rd drug, 2 -6 hours (h) from the first dose of drug, 3 - 6 h from the th th th th th second dose of drug, 4 -6 h, 5 - 24 h, 6 -48 h, 7 -72 h, 8 -96 h and th 9 -120 hours from the third dose of drug. AA concentration was measured by the hydrazine method. Somatic cell count in the milk cows from group A also was higher (1.81 million) than in milk of cows from group B (0.56 million).During the bacteriological examination of milk samples Staphylococcus aureus was isolated. Results of experiment show, that average serum AA concentration in group 3 experimental A was 48.99 µmol/dm , but in group control B 63 3.The results of clinical examination of the mammary gland, µmol/dm organoleptic evaluation of mammary gland secretion, number of somatic cells, and bacteriological examinations of milk indicated a clinical form of mastitis caused by Staphylococcus aureus in cows from group A. It confirm of decrease antioxidant protection potential in blood of cows with clinical inflammation of mammary gland. Key words: cow, mastitis, ascorbic acid 592 Glutathione Peroxidase Activity in Blood of Cows Affected with Subclinical Mastitis before and after Treatment Using Cepravin TM Dry Cow T. Jakubowski, W. Klucinski, M. Kleczkowski, K. Dembele Warsaw University of Life Sciences, Clinical Sciences, Warsaw, Poland The aim of the investigations was evaluation of GSH-PX activity in the whole blood of cows with subclinical mastitis caused by Streptococcus agalactiae before and after treatment using Cepravin TM Dry Cow.The experiment was conducted on 24 cows, Black-White breed, at the age of 4-7 years. The animals were divided into two groups (Aexperimental and B- control, 12 animals in each). All cows were subjected to an initial clinical examination, including the mammary gland, evaluation of macroscopic changes, and evaluation of somatic cell count in milk. Milk samples, before treatment, were taken simultaneously for identifying pathogenic micro-organisms. However, before treatment of 12 cows the somatic cell count was estimated with the microscopic method. All cows from group A were given Cepravin TM Dry Cow. The drug was given once intramammary to each quarter st separately during 1 day of dry period. Blood samples from all cows (group A and B) were collected from the external jugular vein 9 times: st nd rd th 1 -before giving a drug, 2 -6 hours (h) from treatment, 3 - 24, 4 -48, th th th th th 5 -72, 6 -96, 7 -120, 8 -144 and 9 -168 h from treatment. GSH-PX activity in the whole blood of cows was measured by the GSH-PXRandox test.The examinations indicate on subclinical mastitis only in cows from group A. Somatic cell count in the mammary gland

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secretion 12 cows from group A before treatment was higher (660 000/ml) than in the milk of 12 cows from group B (107 000/ml).During the bacteriological examination of milk Streptococcus agalactiae was isolated. Results of experiment show, that average GSH-PX activity in the whole blood in group experimental A was 42.1 U/g Hb, but in group control B 33.1 U/g Hb. The higher one was in the blood of cows from group A-before treatment (51.1 U/g Hb). It confirms an increase of enzymatic antioxidant protection in blood of cows with subclinical inflammation of mammary gland Key words: cows, glutathione peroxidase, antioxidants, mastitis 593 Etiology of Bovine Mastitis in the Municipality of Patzcuaro, Michoacan, Mexico C. Bedolla , A. Mejia , S. Renteria , J. Conejo , H. Castaneda , 3 4 W. Wolter , M. Kivaria 1 Universidad de Michoacan, Department for Infectious Diseases Epidemiology; Faculty of Veterinary Medicine, Morelia, Mexico 2 Universidad de Guadalajara, Department for Infectious Diseases Epidemiology; Faculty of Veterinary Medicine, Guadalajara, Mexico 3 Landesbetrieb Hessiches Landeslabor, Department of Veterinary Medicine, Giessen, Germany 4 Central Veterinary Laboratory, Department of Microbiology, Dar es Salaam, Tanzania The objective of this study was to determine the bovine mastitis etiology in the municipality of Patzcuaro, Michoacan, Mexico. The research was made from September to November 2007 in Patzcuaro, Michoacan. Eight milk cattle barns of Holstein Friesian cows were sampled, which count on 10 cows in average and which they are raised in small scale cattle barns production system. A total of 347 milk samples were collected aseptically in sterile tubes of single udder quarters of the mammary gland in 88 lactating cows and transported to the laboratory to low temperature (4° C). The samples were inoculated in both, blood agar with esculin and McConkey agar, incubated at 37 oC, and observed 24 h and 48 h after inoculation to assess colony growth. Staphylococci isolations were reinoculated in agar with sheep blood, isolated, in order to observe colonial morphology, hemolysis as well as Gram stain, catalase tests, coagulase, manitol and gelatinase. The Gram negative pathogens were identified accord to colonial morphology, and biochemical tests. From the results of the bacteriological examination of the milk samples, 64 (18.44%) corresponded to Staphylococcus aureus, 85 (24.49%) to coagulase negative staphylococci, 24 (6.91%) to esculin positive streptococci, 2 (0.57%) to Escherichia coli, whereas 122 (35.46%) corresponds to other pathogenic bacteria. We concluded that coagulase negative staphylococci were the main pathogen agents found of the bovine mastitis, by self the Staphylococcus aureus, was the main causal agent of the contagious bovine mastitis in this study. Key words: bovine mastitis, small scale barns, S. aureus, clinical mastitis 594 Origin of Clinical and Subclinical Mastitis in Dairy Herd's of the Federal State Jalisco, Mexico. H. Castaneda , S. Jäger , W. Wolter , M. Zschöck , 1 1 3 M. Castaneda , G. Perez , J. Serratos 1 Universidad de Guadalajara, Department of Veterinary Medicine, Guadalajara, Mexico 2 Landesbetrieb Hessiches Landeslabor, Department of Veterinary Medicine, Giessen, Germany 3 Instituto Tecnologico de Tlajomulco, Depto de Biotecnologia, Tlajomulco, Mexico Aim of the present work was to prove the occurrence of subclinic and clinic disturbances of udder health in 33 herds of dairy cattle in Jalisco, Mexico. 1996 (66.9%) out of 2937 udder quarters examined by means of California Mastitis Test, showed a positive reaction, 1087 (37%) out of these reactions were from clearly up to significantly positive reactions. Compared to the bacteriological examinations the prevalence for subclinical mastitis came up to 43.7%. On the other hand clinical mastitis could be proved in 2.5%. In 53.8% of the examined quarter milk samples there was no bacteriological pathogen content. From the rest of the samples we could isolate CNS (coagulase negative staphylococci) (15.4%), Corynebacterium spp (13.9%), Streptococcus agalactiae (6.6%), Staphylococcus aureus (5.8%), coliform pathogens

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(3.6%) and others (Bacillus spp., Nocardia spp, Candida spp.) (1.7%). These results demonstrate a significant share of minor pathogens beside contagious mastitis pathogens as S. aureus and S. agalactiae in mastitis incidents in Jalisco, Mexico. Key words: clinical and subclinical mastitis, bacterial pathogens, dairy cattle, Staphylococcus aureus, Streptococcus agalactiae 595 Evaluating Post-Milking Teat Dip Efficacy Using Somatic Cell Count Data A. Bradley , J. Breen , P. Janowicz , M. McKinzie , T. Hemling 1 University of Bristol, School of Veterinary Science, Bristol, United Kingdom 2 Quality Milk Management Services Ltd., Wells, United Kingdom 3 DeLaval, Kansas City, United States

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Objectives: Post milking teat disinfectants are traditionally assessed by identifying new intramammary infections (IMI) using regular individual quarter bacteriology. Somatic cell counts (SCC) are recognised as a proxy for IMI; quarters can be classified as "infected" or "uninfected" and new infections defined by movements across defined thresholds. The aim of this study was to compare the efficacy of two post milking teat disinfectants using both bacteriology and somatic cell counts, and to assess the utility of these two approaches. Materials and Methods: Two teat dip formulations (DeLaval Prima (0.5% hydrogen peroxide) and Hamra Blue (0.4% chlorhexidine)) were compared on three farms in the UK. Four hundred and fourteen cows were randomly allocated (ratio 1:1) to be dipped post milking with one of the two formulations. Cows were recruited over a 20 week period; 377 cows were eligible for inclusion in the subsequent analyses. Quarter milk samples were collected for bacteriology and SCC from all cows at enrolment and on completion of the study. During the study, quarters were sampled for SCC every 14 days. Bacteriology to confirm new IMIs was performed when quarters experienced a rise in SCC from below to above 100,000 cells/ml. All cases of clinical mastitis were sampled during the study in order to determine the cause of IMI. New IMIs were defined using various criteria; efficacy was evaluated using statistical protocols recommended by the National Mastitis Council (NMC). Results: Efficacy of the two products was compared; new IMIs were defined by movements across the 100,000 cells/ml threshold alone, and in combination with supporting bacteriology. Efficacy was also evaluated by comparing clinical mastitis incidence and the change in prevalence of IMI between entry and exit of quarters from the study. With the exception of the change in prevalence of infection between entry and exit from the study (due to a difference in minor pathogen IMI), none of the measures demonstrated a significant difference between the products. Analysis by various statistical protocols recommended by the NMC showed the experimental product (DeLaval Prima) was equivalent to the control product (Hamra Blue). Conclusion: The use of SCCs, with/without the judicious use of bacteriology offers a viable, cost effective alternative to the extensive use of bacteriology alone in assessing post-milking teat dip efficacy. 596 The Effectiveness of Cefalosporine+Kanamicine and Ampiciline Gentamicine + Dexametazone in the Treatment of Clinical Mastitis in the Livestock Bovine Milk in Mexico M. Posadas, L. Matus, O. Blanco, B. Pena Facultad de Medicina Veterinaria y Zootecnia-UNAM, Animal production, México DF, Mexico The present study was carried in Tizayuca Hidalgo, Mexico during July to September in order to determining the effectiveness of the treatment with the combination cefalexine monohydrate, kanamicine sulfate and ampiciline - gentamicine + dexametazone. We administered by the intrammary road for the solution of problems of clinical mastitis in bovine producing of milk. The sample was took in a population of 50 animals diagnosed with clinical mastitis and that they presented alterations in the milk besides that they obtained a grade 2 in test of California (CMT), which were integrated in two groups of 25 glands distributed randomly.The control group (A) it was conformed by 25 glands affected which was administered cefalexine monohydrate + kanamicine sulfate during 2 days for a intramamary road.The witness group (B) was conformed by 21 affected glands, to which was administered: ampiciline - gentamicine + dexametazone for a intramamary road with it forms to their medical prescription. They took samples of milk of the glands affected to carry out the bacteriological

exam and to know the etiology.The isolated agents of the sample of milk of the two groups "A" and "B" was the following ones: Streptococcus agalactiae, Staphtlococcus aureus, Corynebacterium pseudotuberculosis, Pasterella spp., Moraxella spp., Eschericha coli, Staphylococcus epidermidis, Staphylococcus, saprophyticus, Serratia spp., Streptococcus uberis, Citrobacter spp., coagulase negative Staphylococci, Actinobacillus salpingitidis, Staphylococcus intermedius and Aeromona hydrophila. The results demonstrated that the combination cefalexine monohydrate and kanamicine sulfate, it had an effectiveness of 88% to control the presentation of clinical mastitis of the group "A" in comparison with the group "B" that was of 66%. In the statistical analysis they were not significant differences (P=.065>0.05). As for the presentation of alterations in the milk, the animals of the group "A" they recovered in 86% while the group "B" was of 76%. With regard to the test of California for the group "A" 82% of the glands didn't present reaction some after the treatment while in the group "B" was of 68%, what allows to deduce that the cefalexine monohydrate and kanamicine sulfate is a highly advisable product for the treatment of clinical mastitis. 597 Effect of Anhydrous Cephalonium-Based Drug Indicated for Bovine Mastitis Treatment during the Dry Period on Phagocytosis Rates C. Batista, M. Blagitz, M. Azedo, C. Stricagnolo, M.C. Sucupira, A.M. Della Libera Faculdade de Medicina Veterinaria da Universidade de Sao Paulo, Clinica Medica, Sao Paulo, Brazil The dry period is considered a very vulnerable phase of lactation to mastitis and the phagocytosis mechanisms are fundamental for this period. The antibiotic therapy used for the treatment of dry cows acts on the elimination of existing intramammary infections, avoiding new infections and decreasing its incidence on the next lactation. An efficient drug must contain an adequate antimicrobial component in harmony with other eventual components and must be harmless to the organism's defense mechanisms. Twenty-one paired samples, negative to bacteriological tests, were used to evaluate the influence of Cepravin®, an anhydrous cephalonium-based drug indicated for the treatment of mastitis during the dry period, on the phagocytic function of milk leukocytes. In order to evaluate the cellular activity, leukocytes were isolated in adequate quality (viability) and quantity (concentration) and submitted to in vitro phagocytosis test with Zymosan (Saccaromyces cerevisae cell wall), after exposition to a solution containing the drug. The average phagocytosis rate for cells submitted to the treatment (64.43% ± 15.10) did not differ in relation to the average phagocytosis rate for the control group (58,33% ± 12,00). In addition, the coefficient of variation of phagocytosis rate obtained from cells submitted to the treatment (23.44) was not different from the coeficient found in control samples (20.57). The results suggest that the treatment, besides the antibiotic activity, does not interfere in the phagocytic function of milk leukocytes, which is fundamental for the defense of mammary gland, mainly during the dry period. Key words: mastitis, treatment, phagocytosis, dry period 598 Microbiological Evaluation, Somatic Cell and Differential Leukocytes Count in Relation to California Mastitis Test in Sante In's Ewes M. Blagitz , F. Souza , V. Gomez , C. Batista , M. Azedo , 1 2 2 1 C. Strincagnolo , N. Benites , P. Melville , M.C. Sucupira , 1 A. M. Della Libera 1 Faculty of Veterinary Medicine and Animal Husbandry FMVZ-USP, Clinica Medica, Sao Paulo, Brazil 2 Faculdade de Medicina Veterinaria da Universidade de Sao Paulo, Preventive Medicine and Public Health, Sao ppaulo, Brazil To evaluate the variation in microbiological profile, total somatic cell (SCC) and differential leukocytes counts (DSCC) in milk of Santa Ins ewes according to CMT scores, 170 milk samples were examined for microscopic somatic cell count (SCC), differential leukocytes count (DSCC), microbiological examination and their relationship with California mastitis test (CMT). On the 24.21% bacteriologically positive samples, 86.96% were coagulase negative staphylococci (CNS), and 13.04% were Streptococcus dysgalactiae. The mean (± Standard deviation) logarithm SCC of uninfected sheep's gland was 5.01 (±0.87) and 5.93 (±0.94) (p<0.0001) for infected glands. Therefore, the percentages of positive bacteriology test were 4.62 %,

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64 XXV. Jubilee World Buiatrics Congress 2008

30.00%, 58.97% and 58.33% in negative, 1, 2, and 3 CMT scores respectively. The proportion of glands considered positive for CMT 1, 2 and 3 was 29.41%, 23.53% and 7.65% correspondingly. The results showed a higher somatic cell count on higher CMT scores, but no significant difference on 2 and 3 scores, thus the means (± Standard deviation) logarithm SCC according to CMT scores were 4.44 (+0.57), 5.21 (±0.46), 6.35 (±0.49), 6.52 (±0.73). Besides this, a positive and significant correlation (r = 0.822, p<0.0001) was encountered between SCC and CMT. Besides this, the correlation between DSCC (Neutrophils (PMN), Macrophages (MA), Lymphocytes (LY), Eosinophils (EOS)) and CMT were r = 0.356 (p<0.0001), r = -0.406 (0.0001), r = -0.057 (p>0.05) and r = 0.19 (p<0.013) respectively. Furthermore, the correlation between PMN and MA were r = -0.951 (p< 0.0001). The means (± Standard deviation) of the percentages of DSCC (PMN, MA, EOS, LY) according to CMT reactivity were 48.61 (±35.84), 43.29 (±34.31), 7.58 (±6.89) and 0.52 (±1.56) for negative CMT; 77.17 (±20.90), 15.30 (±18.94), 5.51 (±4.53), 2.01 (±6.41) for CMT 1; 80.37 (±21.32), 11.01 (±19.85), 6.19 (±6.19), 2.44 (±6.02) for CMT 2; 75.73 (±28.06), 16.63 (±23.86), 7.56 (±9.00) and 0.63 (±1.55); for CMT 3. In face of, CMT presented as a feasible tool for detect mastitis in Santa In's ewes. Financial support was provided by FAPESP. Key words: mastitis, ewes, somatic cell count, California Mastitis Test, leucocytes 599 The Effects of Pasture Management on Udder and Teat Hygiene Scores and Somatic Cell Counts on UK Dairy Farms K. Parker , A. Bradley , S. McDougall 1 Animal Health Centre, R&D, Hamilton, New Zealand 1 University of Bristol, Department of Veterinary Clinical Sciences, Bristol, United Kingdom Pasture management may affect the incidence of diseases such as mastitis and lameness. Dairy cattle may spend periods of time outside of barns, especially over the summer months in some production systems. Pasture may or may not constitute a large portion of the diet during these times. Where the pasture contributes little to the diet, the cattle may be confined to a small area or for a long period of time. In this case, a high density of faecal contamination of the pasture or laneways may result in exposure of cows to faecal-borne pathogens. Poor pasture management and subsequent pasture contamination may lead to a decrease in hygiene scores of the udder and teats, and hence an increased exposure to environmental mastitis pathogens and consequently increased somatic cell counts and increased number of cases of clinical mastitis. Twenty herds from the dairying areas of Wales and England were surveyed. Herds were classified as having a high (n = 8) or low (n = 12) herd incidence of clinical mastitis. Each farm was visited once during May; a number of herd and cow level measurements (on a random sample of 30 cows within each herd) were collected that related to pasture management, farm and cow hygiene and incidence of mastitis. Univariate data analysis was used to define the associations between pasture contamination scores and herd level hygiene scores with clinical mastitis incidence, individual cow SCC and bulk tank SCC. Herds classed as having a high mastitis incidence had a higher udder and teat hygiene score (i.e. were dirtier; p < 0.1). There were positive correlations between average cow hygiene score within a herd and faecal contamination of the pasture area grazed (R2 = 0.28; p = 0.02), and between average cow hygiene score within a herd and average bulk tank somatic cell count (R2 = 0.2; p = 0.09). Higher stocking rate (cows/ha) was positively associated with faecal contamination of the pasture area grazed (R2 = 0.3; p = 0.01). It is concluded that higher stocking rates of cattle at pasture are associated with greater faecal contamination of the pasture which in turn is associated with poorer cow hygiene, and higher incidence of clinical mastitis and bulk tank SCC. This suggests that interventions to improve pasture management may result in cleaner cows, reduced bulk tank SCC and a lower incidence of clinical mastitis. 600 Effect of Lactation Stage on Milk Composition in Healthy Sante Ines Ewes A. M. Della Libera , M. Blagitz , C. Batista , N. Benites , 1 1 1 M. Azedo , F. Souza , V. Gomes 1 Faculty of Veterinary Medicine and Zootechny of the University of Sao Paulo, Clinica Medica, Sao Paulo, Brazil

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Faculdade de Medicina Veterinaria da Universidade de Sao Paulo, Preventive Medicine and Public Health, Sao Paulo, Brazil

The aim of this study was to evaluate milk composition of healthy Santa Ines ewes during lactation. Santa In's is a breed well adapted to tropical condition and are raised for meat production, although it has a considerable milk production. Thus milk samples and udder halves from 11 ewes were examined once a month. Therefore, the maximum lactation period was no more than 4 months. Halves udder was regarded as healthy when the bacteriologically culture was negative. Firstly, the physical examination was performed. To evaluate physical examination, scores (0-3) were used to describe the alterations. Then, milk samples were aseptically collected for bacteriologically examination, pH determination and fat, protein, lactose and total solids content in automatic measurements. The percentage of positive cultures during the lactation was 13.64%, 9.52%, 9.52% and 0.0%. The results showed no significant difference on physical examination scores (1.35 ± 1.09; 1.33 ± 0.86; 1.10 ± 0.97; 1.83 ± 0.94) (p>0.24). Although, higher fat content was observed in the last month (9.78 ± 2.62%) compared to the second month (6.63 ± 1.40%) (p<0.02). The protein content was higher in the third month (7.58 ± 2.81%) in relation to first (5.38 ± 1.32%) and second (5.19 ± 0.69%) (p<0.0001) months. The lactose and total solid content were higher in the first (5.14 ± 1.62% ; 18.72 ± 4.72%) month than the second (4.15 ± 0.78%; 15.30 ± 2.17%) and third (3.35 ± 1.25% ; 14.46 ± 2.83%) months (p<0.0001), the same were seen in the last (3.38 ± 0.82%; 16.38 ± 2.15%) month but no significant difference were observed. No significant difference were encountered in pH (p>0.35) (6.73 ± 0.25; 6.79 ± 0.16; 6.74 ± 0.29; 6.63 ± 0.15). Thus, the lactation period can be responsible for milk quality changes, but it cannot be applied for pH. Financial support was provided by FAPESP Key words: milk composition, ewes, lactation. 601 The Quality of the First Colostrum and the Prophylaxy of the Neonatal Diarrhoea in Calves R. Morar, D. Pusta, I. Pasca, R. Sobolu USAMV Cluj-Napoca, Animal Health Department, Cluj-Napoca, Romania The purpose of the study was to find a correlation between the colostrum's quality and calves diarrhoea.The researches were made on 32 cows, belonging to Romanian Spotted breed, during the first 3 days post-partum and on their obtained 32 calves. There were determined the main biochemical characteristics of the colostrum from each quarter (128 quarters) and the diarrhoea incidence in calves.The pH determinations were made in stable, and the biochemical ones in laboratory. The biochemical determinations were: total acidity (Torner method), chlorides (titrimetric method with potassium sulfocyanide), lactose (method with potassium ferrocyanide) and chlorine - lactose index.There were tested all the 128 quarters. The pH dynamic varies from 5.8 to 8.6. The pH values ranged between 6.5 and 8.6 were considered as positive.In the first day there were found 42 positive quarters, respectively 32.81%, belonging to 20 cows (62.5%). In the second day there were 26 positive quarters (20.31%), belonging to 13 cows (40.62) and in the third day 17 positive quarters, respectively 13.28%, belonging to 7 cows (21.8%). The chlorides vary from 0.17 to 2.92, and when they are over 1 the calves got sick. The acidity is high in normal colostrum (between 16-18.5 T) and decreased in the positive one (under 16 T). The lactose varies between 2.84 ­ 4.00 g/100 ml in the positive colostrum and between 4.00 ­ 7.66 g/100 ml in the normal colostrum. From the 32 obtained calves, after the first day, the neonatal diarrhoea appears in 21, meaning 65%, representing all the calves which have suckling the positive colostrum. It can be noticed that the main determined parameters are decreasing from beginning with the first day to the followings, reaching after 5-6 days to the values of the normal milk. We consider that the determination of the colostrum's pH is an important indicator of its sanity. It is recommended not to be administered nor the colostrum or milk to the calves from the positive quarters, found in the first day. If the cow has all the quarters positive it is administered to the calf the colostrum from another cow. It is recommended that just after birth to wash the cow's udder and to completely milking, after that it has to be controlled the pH of each quarter, then the colostrum should be administered in small quantities. Key words: colostrum, calves, neonatal diarrhoea

Mastitis / Udder Health and Milk Quality 65

602 Coagulase Gene Polymorphism of Staphylococcus aureus Isolated from Subclinical Bovine Mastitis O. Aslantas , C. Demir , H. Turutoglu , Z. Cantekin , Y. Ergun , 3 G. Dogruer 1 Mustafa Kemal University, Microbiology, Antakya, Turkey 2 Mehmet Akif Ersoy University, Microbiology, Burdur, Turkey 3 Mustafa Kemal University, Reproducción, Antakya, Turkey This study was conducted to investigate the coagulase gene polymorphism of Staphylococcus aureus isolated from subclinical bovine mastitic milk samples from 3 different Turkish cities (Hatay, Gaziantep, and Burdur). Amplification of the coagulase gene from 80 S. aureus isolates produced 5 different polymerase chain reaction (PCR) products; 79 isolates showed only 1 amplicon, and 1 isolate showed 2 amplicons. The isolates were grouped into 9 genotypes by analysing the restriction fragment length polymorphism (RFLP) of the gene; the 2 most common genotypes accounted for 73.8% of the isolates. The results indicated that many coagulase gene genotypes are present in the studied regions and only 1 or 2 genotypes predominated. Key words: Staphylococcus aureus, RFLP-PCR, mastitis 603 Variation of the Cow Milk Quarter-level Somatic Cell Count and Lactose Concentration in Relation to Blood Urea, Complement and Hematological Parameters I. Lusis , V. Antane 1 Faculty of Veterinary Medicine, Latvia University of Agriculture, Institute of Food and Animal Hygiene, Jelgava, Latvia 2 Faculty of Veterinary Medicine, Latvia University of Agriculture, Clinical institute, Jelgava, Latvia Introduction: Immune system status in the cows body is essential to keep udder healthy and free from mastitis pathogens. Activation of the immune response is characterized by increase in cell activity, release of mediators and increased number of cells in tissues, body fluids, secretions. It is necessary to differentiate between adequate or effective somatic cell count (SCC) increase in milk and such as long lasting, excessive or pathological one. Subclinical mastitis is the main cause for SCC increase. The aim: Evaluate correlation among quarter-level SCC, lactose content, bacteria in milk and cow blood urea nitrogen, complement C3, C4 concentrations as well as hematological parameters. Material and methods: Group of 20 cows was selected in a 400-dairy cow herd of teaching and research farm Vecauce.Fat, protein, lactose concentration and SCC were estimated in the quarter milk samples by Milkoscan and Somacount analysers. Milk from each quarter was tested bacteriologically with direct culture on the blood agar. Mammary infections were categorized in three groups without infection (score 0), coagulase negative staphylococci (score 1), major pathogens (Staphylococcus aureus, Streptococcus agalactiae, Streptococcus uberis) (score 5). Transformation of the quarter level data to cow level ones was performed. Arithmetic mean for LgSCC, lactose, milk protein, fat concentrations and sum for bacterial infection scores were used. Concentration of the comlement factors C3 and C4 in citrated venous blood samples were detected by immuneturbidimetry. Correlation coefficients among all parameters were calculated by software STATA 9.0 and showed in results only significant ones (p<0.05). Results: LgSCC correlate negatively with lactose concentration (r=0.77), blood urea concentration (r=-0.27), but positively with blood leukocyte count (r=0.25) and complement factor C3 (r=0.56). Lactose concentration is associated with blood urea nitrogen (r=0.36) and negative impact on the complement factor C3 (r=-0.64). Blood urea nitrogen concentration correlate negatively also with complement factor C4 (r=-0.31). Complement factor C3 is positively correlated to blood lymphocyte and monocyte concentrations (r=0.32 and r=0.31, respectively). Mammary infection occurrence in milk samples was negatively correlated with lymphocyte percent in leucogram (r=-0.26) and positively with granulocytes percent (r=0.31). Key words: lactose, milk cell, mastitis, complement 604 Evaluation of Intramuscular Administration of Penicillin G and Oxytetracycline on Milk Yield in Holstein Cows during Milking Period M. Pourjafar , B. Hojati , M. Shakhse-Niaie 1 School of Veterinary Medicine, Shahrekord University, Clinical Sciences, Internal Medicine, Shahrekord, Iran

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Islamic Azad University, Faculty of Veterinary Medicine, Faculty of Veterinary Medicine, Shiraz, Iran 3 School of Veterinary Medicine, Shahrekord University, Faculty of Veterinary Medicine, shahrekord, Iran In this study, 20 Holstein cows with pregnancy age up to most 5.5 months, from a 3200 head sized dairy herd were selected in a randomized manner to test this hypothesis that intramuscular antibiotic administration during lactation affects milk yield. One-week milk-yield was recorded and then the milking cows were divided evenly into four groups (1-4). Groups 1 to 3 received daily intramuscular injection of penicillin G (10,000 IU/Kg), oxytetracycline (5mg/Kg) and normal saline (25 ml), respectively during 1week and in group 4(control) no treatment was done. Recording of milk yield were continued during injection week (IW) and one week after injection. Then milk-yield mean in each group, before IW, IW and after IW were statistically analyzed by paired T-test. Penicillin G injection, during IW, results in low and temporary reduction in milk-yield (P<0.05), that returned to the level of normal after IW. Oxytetracycline injection results are more milk-yield reduction, compared to Penicillin G injection group (P<0.05), and this reduction last few days after IW. So it could be concluded that administration of aforementioned antibiotics during milking seasons in milking cows did not result in milk reduction compared to control and administration of these antibiotics are allowed. Key words: Holstein cows, milk, oxytetracycline, penicillin 605 The Usefulness of Portable DeLaval Cell Counter to Measurement of SCC in Milk from Healthy and Mastitic Cows E. Malinowski, S. Smulski, A. Arczynska National Veterinary Research Institute, Department of Pathophysiology of Reproduction and Mammary Gland, Bydgoszcz, Poland The aim of the examination was to evaluate the effect of the preservative and the temperature and time of samples storing on the SCC by DeLaval cell counter (DCC), Fossomatic 90 (FS) and microscopic method (MM) in cows` milk. Somatic cell count was measured in 25 milk samples from healthy quarters of clinically healthy cows, in 25 milk samples from quarters with subclinical mastitis of other clinically healthy cows, in 25 composite milk samples of clinically healthy cows, and in 25 samples of bulk tank milk. Cells numbers determined by DCC immediately after sampling ranged from 1,000 to 156,000/ml in premilk from healthy quarters, from 142,000 to 1 672,000/ml in cisternal premilk from subclinically inflamed quarters, from 5,000 to 1 442,000/ml in composite cow milk and from 46,000 to 603,000/ml in bulk tank milk. It was stated that DCC shows the same numbers of cells in milk immediately after sampling, and in the same samples cooled and stored for 1h, 24 h, and 72 hours or frozen and stored for 7 days, independently on the preservation with Bronopol. Somatic cell counts in non-preserved milk determined by DCC were very close to numbers of cells stated by Fossomatic and microscopic method. The correlation between data by DCC, FS and MM was statistically significant (p<0.01). In conclusion, DCC is highly useful apparatus for adequate measurements of somatic cell count in milk from healthy and sick cows. DCC is especially useful to evaluation of somatic cells count in bulk tank milk and in quarter milk from subclinically inflamed udders. DCC can measure the somatic cell counts both in fresh non preserved and preserved milk samples. Measurements of SCC with the use of DCC can be performed also in milk samples stored in temperature of 4 °C for 24 and 72 hours as well as in milk samples frozen to minus 20° C and stored for 7 days. Key words: cow milk, SCC, preservative, storage, DCC, Fossomatic, microscopic method 606 Bactericidal Action of the Combination between Cefalexin and Ciprofloxacin against Bovine Mastitis Pathogens E. Bousquet , L. Denault , JP. Ganiere 1 VIRBAC, Carros, France 2 ENV Nantes, Unité de Pathologie Infectieuse, Nantes, France Objective: Injectable and intramammary antibiotic treatments can be combined to improve bovine mastitis recovery. This in vitro study was performed to check for possible synergy between cefalexin (available as an intramammary suspension) and ciprofloxacin (main active metabolite found in milk after enrofloxacin intravenous injection in dairy cows) against major mastitis pathogens.

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66 XXV. Jubilee World Buiatrics Congress 2008

Materials and methods: Strains tested were French isolates of Escherichia coli, Staphylococcus aureus and Streptococcus uberis. Time-kill curves were determined either in Mueller-Hinton broth or in UHT sterilized milk over 24 hours for both antimicrobials alone and in combination. Results: A synergistic bactericidal effect of cefalexin and ciprofloxacin was observed on Escherichia coli and Staphylococcus aureus. Bactericidal action of the combination against Streptococcus uberis reflected mainly effect of cefalexin (combination considered as indifferent against this pathogen). Conclusion: In vitro interaction observed here may be of clinical relevance to improve mastitis recovery in cattle. Key words: cefalexin, ciprofloxacin, enrofloxacin, mastitis, synergy 607 Keratin Plug Presence in Heifers 60 Days before Calving and its Relation with Intramammary Infections L. Jimienez , R. Timon , J. Marco 1 Servet Talavera, Dairy Milk Quality Centre, Talavera de la Reina, Spain 2 Public Health Laboratory, Health Department, País Vasco, Spain Objectives: To study the prevalence of intramammary infections (IMI), their nature and their relation with the presence of the keratin plug, determining the: 1. Prevalence of IMI; 2. Presence of the keratin plug (KP); 3. Microbiology of the samples. Materials and Methods: The study included 158 Holstein heifers from two intensive farms located in central Spain. Milk samples were taken from each quarter 60 days before calving, according to the National Mastitis Council (NMC). At the moment of sample collection the presence of the keratin plug was assessed. Microbiological testing was performed according to NMC procedures. Results: The prevalence of isolated micro-organisms showed a 36,7% of IMM with coagulase negative staphylococci (CNS) found in 75% of the isolates, Arcanobacterium spp in 6%, other Gram positive in 10%, other Gram negatives in 7%, 6% of yeast and 1% of mixed culture. The distribution of CNS isolated from milk samples from heifers 60 days before calving was the following: Staph. choromogenes the most prevalent 66,4%), followed by Staph. sciuri (8,8%) and, Staph. lentus (5,2%). Other isolated species were: Staph. Auricularis, Staph. capitis, Staph. caprae, Staph. cohnii cohnii, Staph. epidermidis, Staph. equorum, Staph. gallinarum, Staph. haemolyticus, Staph. hominis, %), Staph. simulans, Staph. warneri, Staph. xylosus. The distribution of CNS species isolated (173 strains) from milk samples obtained from heifers 60 days before calving. The criterion of novobiocin resistancesensitivity to determine pathogenicity was used, with 90% of the strains being sensitive to novobiocin (CNNSS), and 10% of the strains resistant to novobiocin (CNNRS). The presence of the KP was assessed in a total number of 628 teats, and it was only found in 306 teats (48,7%). The stratified IMI study depending on the presence or not of the KP showed that heifers without KP had 59 % quarters with IMI, while quarters with KP only had 13 % of IMI (P<0,0001). When evaluating the prevalence of the different micro-organisms, 86% of the CNS were isolated from quarters without KP, as well as 100 % Archanobacterium spp, 57% of other Gram +, 76% of other Gram - and 85 % of the mixed growths. Conclusions: The presence or absence of the KP is a very important risk factor for IMI in heifers.It was absent in more than half of the dairy heifers included in the study 60 days before calving.These findings makes preventive treatment (Cepravin® V.S.) advisable to ensure udder health of heifers before their first calving. 608 Preliminary Results of Researches on Correlation between Some Blood Acid-Base Balance Indices and Yield as well as Composition of Cows' Milk S. Kinal , R. Bodarski , J. Pres , J. Twardon , S. Krzywiecki , 1 M. Slupczynska 1 Wroc_aw University of Environmental and Life Sciences, Department of Animal Nutrition and Feed Science, Wroclaw, Poland 2 Wroclaw University of Environmental and Life Sciences, Department and Clinic of Obsterics, Ruminant Diseases and Animal Health, Wroclaw, Poland Objectives of study: The high yield dairy cows are exposed on many metabolic disturbances, from among which the fluctuations of acid1 1 1 2 1 1 1 2

base balance (ABB) are the major. These fluctuations can involved changes in mammary gland metabolism, and thus changes in milk yield and composition. The aim of presented, preliminary study was calculation of correlation coefficients between some blood acid-base balance indices, yield and composition of milk, as well. Materials and Methods: The study was conducted on the 64 dairy cows, which coming from six herds from South-East Poland region. In blood samples taken from cows some indices of acid-base balance were determined.In first trimester of lactation, the yield and milk composition (fat, protein, lactose, dry matter and urea) were controlled. On the basis of obtained results correlations coefficients between some blood acid-base balance indices and yield and composition of milk were calculated with use Statistica software. Results: The correlation coefficients were calculated for ABB indices, such as pH, BE and HCO3 and milk yield and components, such as fat, protein, lactose, dry matter and urea contents. All of calculated correlation coefficients were statistically insignificant (P=0,97-0,14) and had low levels. From among milk components the dry matter and lactose content were the worst correlated with pH, BE and HCO3 of blood. The negative correlations were obtained for pH and protein content, pH and urea concentration, HCO3 and protein content and BE and protein content. Conclusions: On the basis of obtained data it could be stated that there is slightly correlation between ABB indices and yield and milk composition. Those data should be treat as preliminary and its demand to extend and revision with take into consideration greater number of animals. Key words: acid-base balance, milk yeld, milk coposition, correlation 609 Multiplex PCR Detection of Enterotoxin Genes of Staphylococcus aureus Strains Isolated from Bovine Mastitis B. Sareyyupoglu , Z. Cantekin , M. Akan , D. Essiz 1 Faculty of Veterinary Medicine, Ankara University, Department of Microbiology, Ankara, Turkey 2 Faculty of Veterinary Medicine, Mustafa Kemal University, Department of Microbiology, Hatay, Turkey 3 Faculty of Veterinary Medicine, Kafkas University, Department of Pharmacology and Toxicology, Kars, Turkey The staphylococcal enterotoxins (SE) are distinguished for their role in the pathogenesis of human and animal illness. These toxins are responsible for food poisoning outbreaks and toxigenic syndrome in humans, and may contribute to the persistence of Staphylococcus aureus in bovine mammary glands. The objective of this study was to determine the phenotype of enterotoxins and to detect the genes encoding these in S. aureus strains those were derived from mastitic milk specimens from cows in Ankara region. Staphylococcal enterotoxins (SEA, SEB, SEC, and SED), except SEE, were investigated by reverse passive latex agglutination (SET-RPLA) tests in 72 isolates of S. aureus from mastitic milk. Multiplex-PCR (mPCR) assay was used to determine the distribution of enterotoxin genes (sea, seb, sec, sed, and see) in these S. aureus strains isolated from mastitic cows. MPCR assay combined the primers specific for enterotoxin genes with the primers amplifying femA gene specific to S. aureus in a set. PCR amplification of femA gene was used as an internal positive control in mPCR assays. All isolates were found to harbor femA so were they were confirmed to be S. aureus. The prevalence of enteroxin genes was determined as 1 (1,4%) for seb, 35 (48,6%) for sec, 9 (12,5%) for both sed and see, respectively. Following SET-RPLA tests, SEB was detected in only 1 (1,4%) isolate, SEC in 25 (34,7%), and, SED in 4 (5,6%) isolates. SEA was not detected in either mPCR assay or SET-RPLA tests. As a conclusion, S. aureus isolated from bovine mastitis has been found to have a high enterotoxigenic potential in the study. This might be significant for food hygiene especially in cases of subclinical mastitis due to S. aureus and might also contribute to an increased udder pathogenicity of the organisms and, being a fast, reliable and sensitive test multiplex-PCR was found to be a useful assay for the detection of staphylococcal enterotoxin genes. Key words: bovine mastitis, enterotoxin genes, multiplex PCR, Staphylococcus aureus 610 Prevalence of Mastitis Pathogens in Quarter Milk Samples of Dairy Cows in Switzerland M. Brechbuehl , S. Moret-Stalder , S. Albini , T. Kaufmann 1 Clinic for Ruminants, Vetsuisse Faculty, University of Bern, Berne, Switzerland

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2 Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Bern, Berne, Switzerland Objectives of study: The survey was conducted to estimate the prevalence of mastitis pathogens in Swiss dairy cows. The correlation between Somatic Cell Count (SCC) and the different pathogens was investigated. Materials and Methods: The selection procedure was a random onestep-cluster-sampling with stratification according to herd size. Quarter foremilk samples (QFM) were taken from each cow in lactation of the 47 herds for microbiological analysis and SCC determination. QFM samples were considered as positive if > 1 colony was isolated from 10µl milk. A cow was considered positive when at least one quarter yielded a positive result. All isolated streptococcus spp. were further differentiated. Results: The cow prevalence of the most important pathogens of subclinical mastitis ranged from 0.5% to 62.7% (Tab.1). Of the isolated streptococcus spp. 21.5% were S. uberis, 21.7% S. dysgalactiae, 1.3% enterococcus and 48% were diagnosed as probably apathogenic streptoococci (lactococcus or rare types of enterococcus). The quarters infected by S. aureus, streptococcus spp., coagulase-negative staphylococci (CNS) and C. bovis had a significantly higher SCC than QFM with a negative result (Wilcoxon rank sum test, p<0.001). Table 1. Prevalence of mastitis pathogens found in quarter milk samples calculated on the levels of quarters (n=2538), cows (n=638) and herds (n=47) Prevalence % (confidence intervall) Quarter Cow Herd S. agalactiae Streptococcus spp. S. aureus CNS C. bovis Enterobacteriaceae Bacillae 0.2 (-0.2­0.5) 6.0 (4.6­7.4) 1.6 (0.5­2.6) 27.5 (23.1­32.0) 18.9 (13.7­24.0) 0.2 (0.00.4) 0.9 (0.4­1.5) 0.5 (-0.5­1.4) 18.2 (14.6­21.7) 4.9 (1.7­8.0) 62.7 (56.1­69.3) 42.9 (34.2­51.7) 0.8 (0.0­1.6) 3.8 (1.7­5.9) 2.1 (0.0­7.3) 83.0 (71.2­94.8) 38.3 (23.3­53.3) 100.0 93.6 (85.6­100.0) 8.5 (0.0­17.6) 29.8 (15.8­43.9)

23.9% in 2004 and 2005 to 16.4% and 17.4% in 2006 and 2007, respectively. A total of 11,359 strains were isolated from 13,925 infected quarter milk samples out of 164,341 tested in 2007. The most frequently isolated pathogens were Staphylococci including S. aureus (42.6%; 4,841/11,359), E. coli (15.8%; 1,796/11,359), Streptococci (13.7%; 1,558/11,359), Enterobacteriacae (7.9%; 903/11,359), and Gram negative bacilli (4.3%; 486/11,359). Conclusion: This study shows relatively high incidence of bovine mastitis in Korea, however, it also shows a steadily declining trend in annual incidence. The most common isolates from milk samples with SCC greater than 200,000 were Staphylococci including S. aureus, which accounted for over 40% of isolates from infected udder. Key words: bovine mastitis, incidence, Korea 612 Pharmacokinetics and PK/PD Integration of Azithromycin in Lactating Dairy Cows with Subclinical Mastitis Caused by Staphylococcus aureus M. Lucas, L. Marchetti, A. Lambertini, J. Errecalde, N. Mestorino Faculty of Veterinary, UNLP, Pharmacology, La Plata, Argentina Azithromycin (AZT), time-dependent antimicrobial with long persistence. The main AZT characteristics suggest that it could be useful for treating bovine mastitis caused by S. aureus. The objective was to investigate AZT pharmacokinetic (PK) profile in lactating dairy cows with subclinical S. aureus mastitis, and calculate the corresponding pharmacokinetic/pharmacodynamic (PK/PD) parameters. Six Holstein lactating cows with subclinical mastitis with at least one affected mammary quarter (mq) were selected. AZT (10% experimental aqueous solution) was administered IM in 2 doses of 10 mg/kg in a 48h interval. Blood and quarter milk samples were collected at different times until 96h post administration. Milk and plasma AZT concentrations were measured by microbiological assay.The MIC90 was determined in 51 S. aureus strains to calculate PK/PD parameters. PK parameters were determined by non-compartmental analysis. Each mq was considered independently (n=24), since our purpose was to compare the findings in mastitic quarters (n=8) against healthy ones (n=16) and in quarters of high-producing cows (n=12) against lowproducing cows (n=12).The PK/PD parameters calculated were AUC024h/MIC90 (area under the concentration vs. time until 24 h/MIC for 90% of the isolates), AUC00-24h/MIC90 (AUC until infinity/MIC90) and T>MIC90 (time during which the concentrations exceed the MIC90).Milk maximal concentration (Cmax) was 7.6 µg/mL (16.7h rst nd post-1 adm) and 7.7 µg/mL (14h post-2 adm). Milk elimination nd half-lives 50 were 128.1h and 157.9h after 1st and 2 administrations. The mean milk AUC0-24h was 154.2 µg.h/mL. Plasma concentrations were lower. The production level had a major effect on the time during which antimicrobial concentrations in milk exceeded the MIC90 (T>MIC90), being longer in quarters of low-producing cows. The AUC0-24h in milk was higher in the low production group, although the ratio AUC0-24h in milk/AUC0-24h in plasma (Fmilk) may indicate that antimicrobial passage is higher in quarters of high-producing cows. Infection presence in the quarters had a significant effect (p<0,05) on AUC0-24h and clearance from the mammary gland (Clmam). Also, Fmilk, T50, AUC0-24h and mean residence time (MRT) in milk, resulted higher in mastitic quarters. The production level and the mammary health status influenced PK and PK/PD parameters of AZT treatments in bovine mastitis. Key words: azithromycin, pharmacokinetic/pharmacodynamic, Staphylococcus aureus, mastitis, cows 613 Clinical Course of Staphylococcal Bovine Acute Mastitis during Postpartum after Antibiotic Therapy J. Fagliari , L. Matins Filho , A. Vidal-Martins 1 Faculdade de Ciencias Agrarias e Veterinarias/UNESP, Departamento de Clinica e Cirurgia Veterinaria, Jaboticabal-SP, Brazil 2 Centtro Universitario de Rio Preto-UNIRP, Departamento de Medicina Veterinaria, Sao Jose do Rio Preto-SP, Brazil The goal of this study was to evaluate the clinical course of fourteen cases, in thirteen dairy cows, with acute mastitis staphylococcal which early clinical signs had started three to eight days after parturition and were treated immediately. Milk culture was perfomed in 5% blood agar for 24-48 hours and the organisms were isolated and identified by Gram´s staining method, coagulase activity, pigment production,

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Conclusions: The cow prevalence of the two major mastitis pathogens S. aureus and S. uberis was low in Switzerland. These results may be due to the farmers awareness of mastitis and to the high standards set by the milk quality legislation in Switzerland. SCC is a useful monitoring tool to detect cows with intramammary infections. 611 The Incidence of Bovine Mastitis in Korean Dairy Cattle from 2004 to 2007 HM. Nam , JM. Kim , KC. Jang , JM. Kim , HM. Kang , JS. Moon , 2 1 1 HR. Han , SC. Jung , YS. Cho 1 National Institute of Veterinary Research, Bacteriology and Parasitology Division, Anyang-city, Korea, South 2 College of Veterinary Medicine, Veterinary internal medicine, Seoul, Korea, South Objectives of study: The object of this study was to report the nationwide incidence of bovine mastitis and the distribution of pathogens associated with mastitis in Korea during 2004-2007. Materials and Methods: Quarter foremilk samples collected from each cow in selected dairy herds were tested somatic cell counts (SCC), and samples showing SCC greater than 200,000/ml were subjected to bacteriological examination and antimicrobial susceptibility test. Data were collected and analyzed to estimate the incidence of bovine mastitis over a period of four years, from 2004 to 2007. Results: In total, 685,397 quarter milk samples collected from 209,543 heads in 12,686 herds were examined by each local veterinary diagnostic laboratory in 15 provinces throughout the country from 2004 to 2007. Among those tested, 44,391 heads (21.2%) and 71,449 quarters (10.4%) were found to be infected with mastitis, respectively. It shows a steadily declining trend in annual incidence from 26.4% and

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hemolysis, catalase test, glucose and manitol fermentation in aerobiosis and anaerobiosis and acetoin production by Voges-Proskauer test, antibiotic sensitivity test. Organisms were identified as Staphylococcus aureus with in vivo sensitivity for gentamicin. All 14 mastitis cases treated with 300 mg q 12h of gentamicin, after complete milking from the affected udder quarter, five minutes after intramuscular injection of 20UI ocytoxin, for five days. It was administred 3 mg/kg q12h of gentamicin, for 5 days. At the end of the terapy five treated udder quarters had developed both, clinical remission of symptons and bacteriological cure at 8, 12, 16, 17 and 21 days, respectively. In six udder quarters had no response to gentamicin therapy and, after another therapeutic tentative was decided by chemical cauterization with 10% iodine solution. Three udder quarters developed necrosis; overt ischemia was observed 3-5 days after the onset of therapy. It was observed complete sloughing of the affected mammary tissue. Additionally to antibiotic therapy, were provided support fluid therapy for these cows; one cow died. The results showed that the treatment of acute mastitis induced by Sthaphylococcus aureus in early postpartum period had no good efficacy, probably due to biological factors of S. aureus, in particular in lactant mammary gland and, cow physiological conditions at postpartum. Therefore, it is important to provide special hygiene environmental conditions for periparturient cows. Financial Support: FAPESP Key words: cow, acute mastitis, Staphylococcus aureus 614 Comparison between Delaval Cell Counter DCC and Fossomatic 5000 for the Determination of Somatic Cell Count of Bovine Milk JP. Roy, L. DesCôteaux, D. DuTremblay Université de Montréal, Department of clinical science, St-Hyacinthe, Canada Objective: To compare somatic cell count (SCC) measured by the DeLaval cell counter (DCC) and the Fossomatic 5000 (Foss 5000) on individual quarter of Holstein cows. Materials and methods: This study was part of another study to evaluate extended lactational therapy for cows chronically infected by S. aureus. Quarter milk samples were collected from 100 cows coming from 14 dairy herds in St-Hyacinthe (Québec, Canada) at 3 sampling periods: 14 days before the beginning of the treatment, 10 and 31 days after the end of the treatment. At sampling, Bromopol was added to milk samples as conservative agent. Milk samples were analyzed with the DCC before being submitted within 2 days to VALACTA (provincial DHI system) for an evaluation of the SCC by the Foss 5000. Coefficients of determination (R2) were calculated between DCC and Foss 5000 using real and logarithmic fits of the SCC. Results: A total of 827 quarters were sampled and analyzed by both 3 3 methods. The mean values were 2 204 X10 c/mL (1 ­ 29 633 X10 3 3 c/mL) and 1 177 X10 c/mL (1 - 7 632 X10 c/mL) for Foss 5000 and 3 DCC, respectively. The median values were 395 X10 c/mL and 337 3 X10 c/mL for Foss 5000 and DCC, respectively. A total of 315 3 3 samples were below 200 X10 c/mL and 257 were over 1 000 X10 3 c/mL using Foss 5000. A total of 331 samples were below 200 X10 3 c/mL and 230 were over 1 000 X10 c/mL for DCC. When all results were included in the analysis, coefficients of determination (R2) were 0.62 and 0.99 for real and logarithmic fits of the SCC, respectively (P < 0.001). Difference in R2 between both fits is largely explained by the fact that the maximum value reported by the 3 DCC is limited to 4 000 X10 c/mL. For somatic cell counts below 1 3 000 X10 c/mL, the regression coefficient was 0.92 without 3 logarithmic conversion. Below 1 000 X10 c/mL the DCC gives results 10 % lower than the Foss 5000 reader (parameter estimate: 1.103 ± 0.014) and the standard deviation of the residuals is 108 3 X10 c/mL. Conclusion: DCC is an accurate tool to measure the somatic cell counts in quarter milk samples. However, there is limitation for very high values and a 10 % correction factor should be applied to the value obtained by the DCC. The study was supported by DeLaval and Fort Dodge Animal Health 615 Effect of Systemic Antibacterial Administration during Prepartum Period on Coagulase Negative Staphylococcal Intramammary Infection in Holstein Heifers O. Ataie , P. Hovareshti , M. Bolourchi , A. Barin , A. Gerami , 2 A. Niasari-Naslaji

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1

Faculty of Veterinary Medicine, Islamic Azad University, Karaj Branch., Clinical Sciences, Karaj, Iran 2 Faculty of Veterinary Medicine, University of Tehran, Clinical Sciences, Tehran, Iran 3 School of Mathematics, University of Tehran, Statistics and Computer Science, Tehran, Iran Pre- and peripartum intramammary infections may impair mammary development and affect milk production after calving in dairy heifers. In the present study, The efficacy and cure rate of prepartum intramuscular injection of Tylosin and Cefquinome on coagulase negative staphylococcal intramammary infection and reducing somatic cell counts was evaluated during the early lactation period. A total of 183 coagulase negative staphylococcal infected quarters from 229 pregnant heifers were randomly allotted into 3 experimental groups: infected quarters that did not receive any treatment (positive control group; n=60); infected quarters that received Tylosin (Tylosin group; n=61; 10 mg/kg of 20% solution; sid x 3; IM; Tyloject®; Razak, Iran) or Cefquinome (Cefquinome group; n=62; 1 mg/kg of 2.5% suspention; sid x 2; IM; Cobactan®; Intervet, The Netherlands). Uninfected quarters that did not receive any treatment were considered as negative control group (n=64). Treated heifers received Tylosin or Cefquinome 10 to 14 days before expected calving date. Antibiotics residues were monitored on Day 3 after calving using a commercial test (Copan test; Hansen Company, Denmark). Cure rates, based on sampling obtained 3 days after calving, were 85.3% and 69.4% in Tylosin and Cefquinome treated groups, respectively. At the same time, self limited rate in positive control group was 53.3%. Tylosin treated heifers had significantly lower somatic cell count scores (4.2±0.2) compared to positive control group (6.0±0.21; P<0.05). In conclusion, prepartum systemic treatment with Tylosin, 10-14 days prior to calving, may be effective in reducing somatic cell count scores and the prevalence of IMI of CNSs during early lactation period of dairy heifers without producing any detectable antibiotic residues. Key words: heifer mastitis; coagulase negative staphylococci; systemic antibacterial administration 616 Bovine Mastitis Pathogens Isolated from Subclinical and Clinical Mastitis in France O. Bidaud , P. Houffschmitt , J. Swinkels 1 Intervet-Schering Plough, Farm and Production Animals Business Unit, Angers, France 2 Intervet Schering-Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: The objective of this study was to determine the national incidence of mastitis causing pathogens in France. Materials and methods: 90 vet practices from 38 different representative French areas were asked to sample quarters with mastitis. 618 milk samples were taken from November 2005 to July 2007 from subclinical or clinical mastitis cases and sent to an official lab (LDA 35, using CA-SFM standards) for bacteriological culture. Subclinical mastitis was identified by a positive California Mastitis Test. Clinical mastitis was identified by visible changes in the milk aspect. Sampling was mostly performed by the farmers. Results: Bacteria were cultured from 464 of the 618 samples. 5 samples were contaminated (at least 3 different pathogens). 48 samples showed a two different-pathogens growth. 24 different bacterial species were identified. 70% of the pathogens belonged to only 4 different species.The most frequently isolated pathogens for both clinical and subclinical cases were Strep. uberis (25%), E. coli (18%), CNS (14%), Staph. aureus (13%), Klebsiella spp. (5%) and other Enterobacteriaceae (4%). Neither Mycoplasma spp. nor Strep. agalactiae were isolated. Mastitis type (clinical or subclinical) was known for 282 samples. For subclinical mastitis cases (n=119, pathogens in 146 samples), the most frequently isolated pathogens were: Strep. uberis (31%), followed by Staph. aureus (19%) and E. coli (18%), CNS (9%), Klebsiella spp. (6%) and Corynebacterium spp. (6%). For subclinical mastitis, 31 samples showed no growth and 5 were contaminated. For clinical mastitis cases (n=105 pathogens in 136 samples), the most frequently isolated pathogens were Strep. uberis (25%) and E. coli (25%), followed by Enterococcus spp. (8%), Staph. aureus (7%), Strep. dysgalactiae (7%), CNS (6%) and Klebsiella spp. (6%). For clinical mastitis cases, 43 samples showed no growth and no contaminated samples were found. Conclusions: This study shows that in France, Strep. uberis and E. coli are responsible for the majority of clinical mastitis cases and represent

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an important part of the pathogens involved in subclinical mastitis. A significant number of Klebsiella spp. was also found in samples from clinical as well as subclinical infections. 617 Antibiotic Susceptibility of Staphylococcus aureus Strains Isolated from Bovine Mastitis Milk in France O. Bidaud , J. Swinkels , M. Bonnier 1 Intervet Schering-Plough, Farm and Production Animals Business Unit, Angers, France 2 Intervet Schering-Plough, Farm and Production Animals Business Unit, Boxmeer, the Netherlands 3 Laboratoire D'Analyses d'Ille-et-Vilaine, Microbiology, Rennes, France Objectives: Despite the current dominance of environmental pathogens in clinical mastitis in France, contagious pathogens such as Staphylococcus aureus remain important pathogens in both clinical and subclinical mastitis. Furthermore, Staphylococcus aureus infections are often chronic and difficult to cure. This study determines susceptibility of Staphylococcus aureus to antibiotics commonly used for mastitis treatment in France. Materials and Methods: A total of 94 Staphylococcus aureus isolates from samples collected from naturally occurring subclinical and clinical mastitis cases in France were cultured and tested for Minimal Inhibitory Concentrations (MIC) at Laboratoire D'Analyses d'Ille-etVilaine in Rennes (LDA 35), from January 2006 to November 2007. Antibiotic susceptibility was assessed in vitro for 10 antibiotics commonly used in French veterinary practice, including 8 ,-lactams (penicillin G, amoxicillin/clavulanic acid, cloxacillin, nafcillin, cephalexin, cefazolin, cephapirin, cefquinome, gentamicin and enrofloxacin). The agar dilution method was used to determine MIC values. Reference strains (ATCC 29213 and ATCC 25923) were tested and the results were compared to CLSI and CA-SFM standards. Results: The MIC of all antibiotics tested ranged from 0.0312 to 16 µg/mL and MIC90 ranged from 0.125 to 4 µg/mL. In this study, cephapirin and enrofloxacin had the lowest MIC90 (0.125 µg/mL) and therefore were the most potent antibiotics against Staphylococcus aureus. However, enrofloxacin was also the only antibiotic that demonstrated a bimodal distribution indicating the existence of resistant strains. For penicillin G, amoxicillin/clavulanic acid, cephalexin, cefazolin and enrofloxacin, some strains showed MIC > 2 µg/mL. Conclusions: This study shows recent data on susceptibility of Staphylococcus aureus to antibiotics frequently used in European veterinary practice against mastitis. It confirms the potential efficacy of several cephalosporins like cephapirin or cefquinome, and of other molecules like nafcillin or gentamicin against Staphylococcus aureus. However, some Staphylococcus aureus strains with a high MIC for enrofloxacin were found. 618 Antibiotic Susceptibility of Coagulase Negative Staphylococci (CNS) Isolated from Bovine Mastitis Milk in France O. Bidaud , J. Swinkels , M. Bonnier 1 Intervet Schering-Plough, Farm and Production Animals Business Unit, Angers, France 2 Intervet Schering-Plough, Int. Marketing, Boxmeer, The Netherlands 3 Laboratoire D'Analyses d'Ille-et-Vilaine, Microbiology, Rennes, France Objectives: CNS have been reported as emerging mastitis pathogens. The objective was to determine the prevalence of the different CNS strains in France and their susceptibility to antibiotics commonly used for mastitis treatment in France. Materials and Methods: A total of 94 CNS, isolated from milk samples taken from mastitis cases in France, were cultured at LDA 35, from November 2005 to November 2007. If a CNS was isolated, the exact strain was identified using an API ID 32 Staph test. In vitro susceptibility was established for 10 antibiotics: penicillin G, amoxicillin/clavulanic acid, cloxacillin, nafcillin, cephalexin, cefazolin, cephapirin, cefquinome, gentamicin and enrofloxacin. The agar dilution method was used to determine MIC. Reference strains were tested and the results were compared to CLSI and CA-SFM standards. Results: The most prevalent isolated CNS strains were S. chromogenes (n=25), S. warneri (n=13), S. xylosus (n=11), S. epidermidis (n=9), S. simulans (n=8) and some 29 others from 10

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different species.Overall susceptibility of the isolated strains to ,lactams was good, although cephalexin had higher MIC than the other cephalosporins tested. Regarding the MIC distribution, gentamicin and cephapirin were the most potent antibiotics against CNS (MIC90=0.25 µg/ml). Penicillin G and cloxacillin displayed a bimodal distribution indicating the existence of some resistant strains. Regarding the 39 highest MIC values (representing the strains with highest MIC for each antibiotic), S. epidermidis (that represented only 9 strains among the 94 strains tested), was responsible for 23 (i.e. 59%) of these highest MIC values. Conclusions: This study provides original new French data on the antibiotic susceptibility of CNS for antibiotics commonly used in European veterinarian practice for the treatment of mastitis. It confirms the potential efficacy of several cephalosporins like cephapirin, cefquinome and cefazolin, as well as nafcillin, enrofloxacin and gentamicin against CNS. This study is also in line with previous studies and confirms that MIC of most antibiotics for S. epidermidis is higher when compared to the MIC for other CNS species. 619 Uterine Bacteriology Data in Cows with Post Partum Acute Metritis Treated with Cobactan® IU B. Huyghe , C. Sa, de , J. Szabo , J. Foldi , P. Cox , J. Swinkels 1 Intervet Schering-Plough, R&D, Angers, France 2 Faculty of Medical Science, Department of Microbiology, Debrecen, Hungary 3 Intervet Schering-Plough, Farm and Production Animals Business Unit, Budapest, Hungary 4 Intervet Schering-Plough, Int. Marketing, Boxmeer, the Netherlands Objectives: A field trial was conducted in 2006 in Hungary evaluating the efficacy of Cobactan® IU in the treatment of acute metritis in cows. Results of bacteriological identification of uterine pathogens isolated during this trial and their semi-quantitative evaluation are presented here. Materials and Methods: Cows included in the study were diagnosed at 4-14d p.p. with acute metritis, defined as presence of red-brown watery foul smelling uterine discharge. At inclusion (D0) 25% of cows were randomly selected for intra-uterine swabbing for bacteriological investigation (n=33). Thereafter, cows were treated with intra-uterine application of 900mg cefquinome (Cobactan® IU, Intervet). On D21 post treatment a second swabbing was performed (n=25). The swabs were within 24h transported to the diagnostic lab in Portagerm tubes (Biomerieux, France) and cultured for identification and semiquantitative evaluation. Results: At D0, 100% of samples were found to contain various bacteria species. On average each swab contained 3.4+0.80 different 3 5 bacteria species, 10 -10 CFU per swab. E. coli, Prevotella spp. and A. pyogenes were found with 75.8%, 81.8% and 33.3% prevalence respectively. Fusobacterium spp. was detected in 12.1% of swabs. Streptococcus spp. was present in 81.8% of samples, but considered an opportunistic rather than metritis pathogen. On D21, the average number of different species per swab was reduced to 1.2 +1.20. E. coli was no longer detected in 66.7%, A. pyogenes in 100% and Prevotella spp. in 53.6% of cases. If bacteria species were still detected, in most cases the bacterial load was reduced (at least 3 log CFU decrease per swab vs. D0). These results corresponded well with the clinical cure of 78% observed. Conclusions: On D21 after treatment with Cobactan® IU, bacteriological cure or at least 3 log reduction of the bacterial load was reached in 63.2%, 88.9% and 100% for Prevotella spp., E. coli and A. pyogenes, respectively. 620 Effects of Subclinical Mastitis during Early Lactation on Reproductive Parameters in a Dairy Herd in Iran R. Asadpour, M. Tooloei Faculty of Veterinary Medicine, University of Tabriz, Clinical Sciences, Tabriz, Iran Bovine subclinical mastitis is an inflammation of the mammary caused by bacteria which occur frequently during early lactation. The objective of this study was to determine the effect of subclinical mastitis in early lactation on reproductive parameters of dairy Holstein cattle. Composite foremilk samples were collected from 60 th th Holstein cattle during early lactation between 5 to 14 postpartum

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days in a dairy farm in Iran from May to December 2007. The animals were apparently normal and their milks had no alteration in color or other morphologic features. Reproductive parameters of the studied animals including services per conception, days open, and days to first service were recorded from their hygienic files. The milk samples were examined by microbiological analysis. Briefly, foremilk sample (10µl) of each cattle plated onto blood agar culture medium supplemented with 5% defibrinated sheep blood. Plates were incubated at 37 0C and bacterial growth was recorded at 24-h intervals for 2d. Bacteria on primary culture medium were identified tentatively according to colony morphologic features, hemolytic characteristics, Gram stain reaction and catalase test. Gram negative isolates were plated on MacConkeys agar. Presence of any pathogens in at least two consecutive samples on primary cultures was defined as subclinical mastitis. The results were statistically analyzed by the SPSS software Ver.11.5. 60% of the samples (36/60) had bacterial growth in the primary culture medium. The most common mastitis pathogens isolated from samples with subclinical mastitis were Staphylococcus aureus and Streptococcus agalactiae. Mean Days open of the cattle infected with subclinical mastitis (124±6.5 d) were significantly greater compared with uninfected animals (98±3 d) (0.01<p<0.05). There were any significant differences in days to first service and services per conception between the infected and uninfected cows. The occurrence of subclinical mastitis during early lactation resulted in detrimental effects on reproductive performance. Subclinical mastitis may influence reproductive response by alteration in endocrine profiles and follicular development. Another possible action of mastitis involved increasing DO may be to influence the developmental capability of the ovulatory oocyte. 621 Use of Acute Phase Proteins SAA and MAA in the Diagnosis of Mastitis in Dairy Cows G. Gerardi , V. Ferrari , S. Segato , G. Zin , S. Tenti , V. Zampieri , 1 1 2 1 L. Scenna , F. Estini , I. Andrighetto , D. Bernardini 1 University of Padua, Department of Veterinary Clinical Sciences, Legnaro (PD), Italy 2 Istituto. Zoop. Venezie, Legnaro (PD), Italy 3 DVM, Padua, Italy Objectives of study: In dairy herds, mastitis is the most frequent and expensive disease. Cows with clinical signs of mastitis are easily spotted and properly treated. However, sub-clinical infections may not be observed and remain untreated, with negative consequences. Somatic Cell Count (SCC) is an index widely used as indicator of intramammary infection but it remains still elevated for weeks after the infection. California Mastitis Test is a laborious and time consuming method if applied to a large number of animals. Thus, in this study, acute phase proteins such as Serum Amyloid A (SAA) and Milk Amyloid A (MAA) were investigated as new potential parameters in the diagnosis of mastitis. Materials and methods: Three groups of Italian Holstein Friesian cows were selected according to the clinical signs of mastitis: 5 healthy (H) (negative bacteriology; 1.15 of linear score of SCC); 10 with subclinical mastitis (S) (negative/positive bacteriology; 3.38 of linear score of SCC) and 6 with clinical mastitis (M) (positive bacteriology; 6.28 of linear score of SCC). SAA was determined both in serum and milk samples from udder quarters (Cat No: TP-802, Tridelta LTD, Ireland), meanwhile MAA only in milk samples (Cat No: TP-807). Composite data (average of udder quarter) were submitted to ANOVA and Pearson correlation test by SAS (2001). Results: As expected, MAA concentration was significantly different among groups: 0.1, 5.2 and 7.6 µg/mL for H-, S- and M-cows, respectively. A similar result was observed as regard of SAA concentration in serum samples: 17, 104 and 245 µg/mL. Linear regression analysis revealed that there was no correlation between MAA and SAA in serum samples. MAA resulted correlated with SCC (r=0.57; P<0.01), as well as SAA in serum samples and SCC (r=0.53; P<0.05), meanwhile the correlation between SAA in milk samples and SCC was higher (r=0.81; P<0.001). Conclusions: In cows suffering from sub-clinical and clinical mastitis, significantly higher SAA and MAA values has been found compared to healthy cows. According to literature, MAA concentration seems to be more correlated with severity of the disease than SAA determined in serum samples. A correlation between SAA in milk and serum samples was detected, even though this finding must be confirmed by future studies on the applicability of acute phase proteins as markers of mastitis.

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Key words: mastitis, dairy cow, serum amyloid A, milk amyloid A, acute phase proteins 622 Specificity of the PathoProof Mastitis PCR Assay Validated Using Culture Isolates J. Holopainen, L. Salmikivi, M. Koskinen Finnzymes Oy, Diagnostics, Espoo, Finland The PathoProof Mastitis PCR Assay is a novel polymerase chain reaction-based reagent kit for identification of all major mastitis-causing pathogens. In this study we analyzed the specificity of the kit using culture isolates representing the most common mastitis causing bacteria and their phylogenetically close relatives, as well as species from environmental and human flora. Bacterial targets of the PCR test include S.aureus, Staphylococcus sp. (CNS), S.agalactiae, S. uberis, S. dysgalactiae, E. faecalis, E. faecium, C. bovis, A. pyogenes, P. indolicus, E. coli, K. pneumoniae, K. oxytoca and S. marcescens, and Staphylococcal betalactamase (blaZ) gene. The PCR test was used as instructed by the manufacturer for analyzing a total of 600 bacterial strains from 80 species, families or groups including mastitis culture isolates, their phylogenetically closely related species, environmental and human pathogens. The overall specificity of the assay was >99%. Clinically significant cross-reactions were not detected suggesting that the assay has good specificity for mastitis diagnostics. In the future, the clinical specificity of the assay against bacterial culture should be evaluated. 623 Detection of Mammary Blood Flow Changes by Transrectal Colour Doppler Sonography in an E. coli Mastitis Model A. Potatpow , S. Schmauder , W. Petzl , C. Sauter-Louis , J. Friker , 2 1 C. Poulsen Nautrup , H. Zerbe 1 Clinic for Ruminants, Faculty of Veterinary Science, Munich, Germany 2 Chair of Systematic and Topographic-clinical Anatomy, Faculty of Veterinary Science, Munich, Germany Transrectal colour Doppler sonography was applied as a non-invasive method for measuring mammary blood flow during an induced E. coli infection. Five Holstein Friesian cows (first lactation, 4 to 6 months after calving, not pregnant, milk yield of 28.6 ± 1.60 kg a day on average) were examined at times of 0h, 12h and 24h post infection by scanning both the left and the right pudendoepigastric trunk as the major supplier of the mammary gland of cows. All animals were infected into the right rear udder quarter. None of them had ever shown any clinical signs of mastitis before and all examined udder quarters were clinically healthy (SCC < 50.000 / ml, bacteriologically negative). Blood flow was expressed by blood flow volume (BFV) and systolic peak velocity (SPV). Significant differences of increasing BFV between 0h and 12h (p=0.043) and decreasing BFV between 12h and 24h (p=0.043) were discovered for the pudendoepigastric trunk of the infected right side. Also SPV increased significantly between 0h and 12h (p=0.043) for the right vessel. No significant differences both of BFV and SPV were detected for the non-infected left side (p>0.05). The difference of decreasing milk yield between 12h and 24h was negatively correlated with the difference of BFV (r= 0.725) and SPV (r= 0.817) for the right pudendoepigastric trunk. Correlations between an increase of SCC and a decrease of BFV (r= 0.877) and SPV (r= 0.691) were observed between 12h and 24h. Bilateral comparisons of BFV and SPV allowed clear identification of the infected udder half. The results show that transrectal colour Doppler sonography of the pudendoepigastric trunk is an adequate technique for measuring mammary blood flow changes within certain time slots during an acute mastitis in cows. Key words: sonography, mastitis, mammary blood flow 624 Total and Differential Somatic Cell Counts in the Colostrum of Non-infected Cows V. Gomes , K. Madureira , S. Kitamura , A. Gomes , M. Blagitz , 4 4 A. Della Libera , F. Benesi 1 Faculdades Anhanguera, Medicina Veterinária, Leme, Brazil 2 Universidade Anhembi Morumbi, Medicina Veterinária, Sao Paulo, Brazil 3 Universidade Estadual do Norte do Parana, Medicina Veterinária, Bandeirantes, Brazil

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Mastitis / Udder Health and Milk Quality 71

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Universidade de Sao Paulo, Clinica Medica, Sao Paulo, Brazil

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The objective of the present study was to determine the quantity of somatic cells found in the colostrum of healthy Holstein cows. A total of 171 colostrum samples were collected before the first and second milking of the cows. Exclusion criteria were physical examination, including examination of the udders, and bacteriological examination of colostrum. Samples were collected and divided in two aliquots. The first one was used in quantitative cytology, and was obtained by adding 1mL of colostrum to 1mL of phosphate buffered saline (PBS), in glass tubes. The second aliquot, used in qualitative cytology, was obtained adding 1mL of colostrum to 49 mL of PBS in sterile Falcon tubes. Total leukocyte counts were determined by means of direct microscopy, using Rosenfeld stain, and differentiating mononuclear and polymorphonuclear leukocytes. For qualitative cytology, collostrum smears were produced by cytocentrifugation and were then stained by Rosenfeld. Results were expressed in relative values (%) for the different types of leukocytes. Medians of mononuclear and polymorphonuclear leukocytes per mL of colostrum before the first milkiing were equal to 657,064 and 178,550, and before the second milking, 828,472 and 392,810, respectively. There was a statistical difference (p<0.0001) in polymorphonuclear leulocytes counts between the first and second milking. Mean relative values of leukocytes found in colostrum before the first and second milking were respectively 69.5; 16.4; 13.3 and 0.37%; 68.0; 13.4; 18.5, and 0% for monocytes + epithelial cells, lymphocytes, neutrophils and eosinophils, respectively. There were no differences in relative values of leukocytes according to the moment of milking. Results showed the type and number of leukocytes found in bovine colostrum, which may provide important defense to neonates both systemically and locally, as intestinal mucosa immunity against diarrhea-causing agents. Key words: colostrum, cows, leukocytes 625 Imunnoglobulins in the Colostrum of Non-infected Cows V. Gomes , K. Madureira , A. Moraes , M. Blagitz , F. Benesi 1 Faculdades Anhanguera, Medicina Veterinária, Leme, Brazil 2 Universidade de Sao Paulo, Clinica Medica, Sao Paulo, Brazil The objective of the present study was to quantify immunoglobulins G, A and M found in the colostrum of bovine females. A total of 171 colostrum samples were collected before the first and second milking of the cows. Exclusion criteria were physical examination, including examination of the udders, and bacteriological examination of colostrum. Ten mL samples were collected in 15-mL Falcon tubes and sent to the laboratory under refrigeration. Tubes were centrifuged at 3,600 rpm for 1 hour, separating colostrum in 3 phases: an upper fat layer, an intermediate layer of colostrum fluid and cell sediment. The supernatant was separated and frozen at -20 °C, to determine immunoglobulin concentration by means of radial immunodiffusion using commercial kits. Three 3 µL of colostrum serum were placed in specific plates for qualitative immunoglobulin (Igs) G, A assays. Sera were diluted to fit the range of the kits. Plates were incubated for 18 to 24 hours at room temperature and Igs were quantified by the intensity of the reactions, compared with predetermined patterns for each Ig type. Values obtained were multiplied by the dilution factor of the samples and results were expressed in mg/dL of colostrum. Mean values obtained for the first and second milking were 10,078 and 6,591mg/dL of colostrum for IgG; 414 and 298mg/dL for IgA; 652 and 349 mg/dL for IgM, respectively. Significant differences (p<0.0001) were observed in IgG and IgM concentrations between the first and second milking. IgG concentration was greater than IgA and IgM (p<0.0001). Results showed that immunoglobulin found in cow colostrum provide important defense mechanism for neonates, for the bovine placenta prevents antibodies from passing to the fetus during pregnancy. Because of this, it is very important to supply first milking colostrum to the calves in the first twelve hours after birth, in order to guarantee colostrum quality and Ig absorption. Key words: colostrum, cows, imunnoglobulins 626 Direct Bactericidal Activity of Phagocytes from Bovine Colostrum against Enterotoxigenic Escherichia coli (ETEC) V. Gomes , K. Madureira , A. Moreno , J. Moura , C. Bonato , 2 3 2 M. Blagitz , G. Pontes , F. Benesi 1 Faculdades Anhanguera, Medicina Veterinária, Leme, Brazil

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Universidade de Sao Paulo, Medicina Veterinária, Sao Paulo, Brazil 3 Universidade de Sao Paulo, Instituto de Ciencias Biomedicas, Sao Paulo, Brazil The objective of the present study was to evaluate the direct bactericide activity of phagocytes found in bovine colostrum against ETEC. Duplicate samples of first milking colostrum were collected from 4 Holstein cows. Samples were collected in sterile Falcon tubes containing 20mL of phosphate buffered saline (PBS), and volume was complete to 40mL with colostrum. Phagocytes were obtained after centrifugation of the samples in a refrigerated centrifuge at 1,500 rpm and 4°C, producing cell sediment (CS), colostrum serum and a fat layer. Only CS was used, resuspended in PBS, recentrifuged to wash the cells and to obtain their concentration with trypan blue. ETEC strains were obtained from diarrheic feces of neonate calves, replicated 18 hours before each assay in tubes containing 5mL of BHI broth, and incubated at 37°C for 18 hours. After growth, bacteria were washed twice in PBS, and concentration was adjusted to 2x107 bacteria/mL, as measured in a spectrophotometer. A concentration of 2x106 phagocytes and 2x107 bacteria were mixed and volume was completed culture medium to produce 2mL. Tubes were incubated at 37°C under stirring during 30 minutes for phagocytosis and for other 30, 60 and 120 minutes for the analysis of bactericidal activity. After the incubation periods were completed, tubes were centrifuged in a refrigerated centrifuge to eliminate extracellular bacteria. The supernatant was discarded, and the sediment was treated with 0.6 mL TritonX100 at 1% for 5 minutes for the lysis of the cell membranes and release of intracellular ETECs. After that, 1.4 mL of BHI broth was added and the solution was diluted serially up to 10-6. From this last aliquot, 100µL were removed and placed in Petri dishes containing TSA. These cultures remained for 18 hours in an incubator at 37°C, and then bacterial colonies were counted. Mean counts obtained were 33, 21, 13 and 37 colonies after phagocytosis, and after 30, 60 and 120 minutes of bactericidal activity, showing the bactericidal activity of phagocytes against ETEC, and demonstrating the imporance of giving fresh colostrum containing viable cells to prevent neonatal diseases. Key words: colostrum, phagocytes, E. coli 627 Study of Microbiological Analysis and Somatic Cell Count on Milk Samples from Buffalo Females (Bubalus bubalis) in State of Sao Paulo, Brazil. J. Kapronezai, N. Benites Faculty of Veterinary Medicine and Zootechny of the University of Sao Paulo, Department of Preventive Veterinary Medicine and Animal Health, Sâo Paulo, Brazil The aim of the present work was the study of microbiological analysis and somatic cell count of milk samples from buffalo cows of different breeds and lactation periods. The milk samples were collected from 262 mammary quarters of buffalo cows to microbiological analysis and somatic cells counting. The somatic cells countings were obtained through optical microscopy reading of the milk smears, where all fields of the 1 cm2 smears area were considered, using the Prescott and Breed Method (1910), modified by Benites et al., (2001). The microbiological analysis resulted in samples with no microorganism growth (75,6%); Staphylococcus spp. (11,8%); Corynebacterium spp. (7,3%) and Streptococcus spp. (3,1%) and associated microorganisms growth (1,15%). The isolation of Corynebacterium spp. and Staphylococcus spp. was statistically superior than the others agents observed. In the samples with microorganism growth, it has been identified the following species: Staphylococcus spp. (S. hominis, S. epidermidis, S. cohnei, S. schleiferi, S. intermedius, S. hyicus), Streptococcus spp. (S. dysgalactiae, S. agalactiae), Corynebacterium spp. (C. bovis, C. xerosis). The total counting of somatic cells in the evaluated milk samples showed median of 2300 cells/mL. The total somatic cells counting/mL in the microbiological negative samples showed median of 1900 cells/mL. Considering the samples with microorganisms' isolation, the median was 8500 cells/mL for Staphylococcus spp isolation; 10350 cells/mL for Streptococcus spp and 9600 cells/mL for Corynebacterium spp. The total somatic cells counting in the microbiological negative samples were statistically smaller than the findings in the samples that showed isolation of

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Staphylococcus spp. (P<0,001); Streptococcus spp. (P<0,01) e Corynebacterium spp. (P<0,001). Comparing the samples with isolation of Staphylococcus spp., Streptococcus spp. e Corynebacterium spp. there was no statistically significant difference in the total somatic cells countings. 628 Bacteria Associated with Subclinical Mastitis in Heifers in a Dairy Herd in Iran R. Asadpour , M. Tooloei , N. Nofouzi 1 Faculty of Veterinary Medicine, University of Tabriz, Clinical Sciences, Tabriz, Iran 2 Faculty of Veterinary Medicine, University of Tabriz, Pathobiology, Tabriz, Iran Mastitis is an important culling reason in heifers. The aims of this study were to identify microorganisms that cause subclinical mastitis in Holstein heifers in a dairy herd in Iran and to estimate the relative importance of the various organisms.Composite milk samples were th th collected aseptically from 48 Holstein heifers between 5 to 14 postpartum days and were analyzed with the Fossomatic 5000 (Foss Electric, Hillerod, Denmark). The animals were apparently normal and their milks had no alteration in color or other morphologic features. To identify sub clinical mastitis in the samples, the SCC technique was used. Ten micro liters of each milk samples were spread on blood agar plates (5%defibrinated sheep blood). Plates were incubated aerobically at 37 °C and examined after 24 h. A threshold of 200,000 cells/ ml has been shown to have a high sensitivity and specificity for sub clinical mastitis. Thus the threshold was used for diagnosis of subclinical mastitis in this study. Colonies were provisionally identified on the basis of Gram stain, morphology, and hemolysis patterns. Gramnegative isolates were identified by using colony morphology, gramstaining characteristics, oxidase, and biochemical reaction on MacConkeys agar. prevalence rate of SCM in the heifers (SCC>200,000 cells/ml), was on average 34% .The organisms that were most frequently isolated from the samples were Coagulase negative Staphylococcus (CNS) (45%) and Staphylococcus aureus (11%). CNS was the predominant pathogens in the heifer's mastitis. This study gives insight in the prevalence of subclinical mastitis in Iran heifers and it shows that subclinical mastitis in heifers is an important issue in dairy practice in the Iran. Key words: subclinical mastitis, bacteria, Iran 629 Subclinical Mastitis in Moroccan Dairy Cattle: Epidemiological and Bacteriological Aspects and Resistance Profile of Isolated Bacteria Z. Zouagui, S. Alali, A. Ukili Institut Agronomique et Vétérinaire Hassan II, Morocco, Pathologie Médicale et Chirurgicale des Ruminants, Rabat, Morocco An epidemiological study of subclinical mastitis in Moroccan dairy cattle has been undertaken in 2007 in Doukkala area. The research has concerned 623 cows originated in 60 differently managed dairy herds. The udder health condition was assessed by the combination of Californian Mastitis Test (CMT) and the milk electric conductivity. A total of 492 quarters were examined. 18 were eliminated due to chronic mastitis. The results showed that 29% of the dairy cows and 9% of the quarters examined have presented, at least one time, a subclinical mastitis during the study period. Moreover, general hygiene conditions within the farms and especially the milking hygiene are of great importance in subclinical mastitis occurrence. Bacteriological analysis of 100 milk samples revealed that 16% were free of bacteria. 9% of the samples presented an association of two or more bacteria species. The frequently isolated bacteria were non haemolytic Staphylococci (40%), non haemolytic Streptococci (24%), Staphylococcus aureus (7%), haemolytic Streptococci (6%) and Escherichia coli (2%). Gram positive rods and yeast were isolated in 21% of the analysed samples. On the other hand, the antibiogram results revealed that most isolated bacteria showed out resistance to at least two antibiotics. Resistance rate to tetracycline, neomycin and cefalotine was more than 50 %, whereas marbofloxacin, trimethoprim and erythromycin showed out resistance levels of respectively 16, 41 and 50%. The least resistance rate was obtained with amoxicillin (8%). Key words: dairy cow, subclinical mastitis, epidemiology, bacteriology, Morocco

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630 Identification the Coagulase-negative Staphylococci Isolates from Bovine Mastitis and Evaluation of the Inflammatory Process E. Costa , F. Guimaraes , P. Faccioli , F. Santos , H. Langoni , 4 4 5 J. Arcaro , A. Peres , M. Cunha 1 FMVZ-Universidade de Sao Paulo, NAPGAMA, Sâo Paulo, Brazil 2 Universidade Estadual Paulista Júlio de Mesquita Filho, Veterinary Hygiene and Public Health, Botucatu-SP, Brazil 3 ICB- Universidade de Sao Paulo, Sâo Paulo, Brazil 4 Instituto de Zootecnia, Secretaria de Agricultura do Estado de Sao Paulo, Nova Odessa, Brazil 5 Instituto de Biociencias, Universidade Estadual Paulista, Botucatu, Botucatu-SP, Brazil Mastitis is an inflammation of the mammary gland caused by microbial infection. It is a major economic concern for the dairy industry worldwide. As many as 50% of all dairy cattle experience some form of mastitis at any given time. Staphylococcus spp. were the single most common mastitis agents. Staphylococcus aureus, a coagulase- positive species is the best known and considered a major mastitis pathogen, while coagulase-negative staphylococci (CNS) have been considered as a minor pathogen. Until the 1970s, reports of infections caused by CNS were rare, nowadays this group of microorganisms was assuming relevance not only in bovine mastitis as well in the etiology of many animal and human pathologies. The objective of this paper was to identify CNS species isolated from bovine milk samples and, evaluated the intensity of the inflammatory process in the infected mammary gland. Ninety-two CNS isolates from Holstein cows mammary glands milk samples, from ten Brazilian dairy herds, were identified by biochemical tests. The identification was performed according to a modified scheme of the reference method proposed by Kloos and Schleifer. The intensity of the inflammatory process was evaluated by somatic cell counts infrared SOMACOUNT IBC (Bentley®, Canada) and, clinical cases were identified by abnormalities in the milk and udder. The CNS identified species were: S. warneri (45.5%); S. hyicus (18.2%); S. chromogenes (14.3%), S. epidermidis (5.2%); S. saprophyticus (5.2%); S. haemolyticus (5.2%); S. capitis (2.6%); S. simulans (1.3%); S. xylosus (1.3%); S. hominis (1.3%). In respect to the intensity of the inflammatory process it was verified statistical difference (P<0.0001) between the SCC of the CNS subclinical 3 3 mastitis cases (mean 421.1 x 10 ) and the negatives (mean 52.1 x 10 ), as well as, between CNS subclinical and clinical cases (mean 5310.2 x 3 10 ). Among, the S. warneri and S. saprophyticus infected quarters it 3 was observed higher than 7000 x 10 . In conclusion, considering the prevalence and the intensity of the CNS mastitis cases reflected by high somatic cell counts, in the development of an effective infection control strategy for mastitis, it is also important to study the epidemiology of CNS in dairy herds. Key words: mastitis; SCC; Coagulase-negative staphylococci 631 Viability Rate of Healthy Cow's Milk Leukocytes Facing Anti-Mastitis Drugs C. Batista, M. Blagitz, M. Azedo, C. Stricagnolo, A. Della Libera College of Veterinary Medicine and Animal Science of University of Sao Paulo, Department of Medical Clinics, Sâo Paulo, Brazil The occurrence of viable cells in mammary gland secretions attests cell function. This fact, along with an applied medicine, is fundamental for the udder healing. Therefore, the objective of this work was to evaluate in vitro viability of milk leukocytes challenged with intramammary anti-mastitis drugs. Active principles of used drugs were: anhydrous cefalonio (Drug 1), gentamicin (Drug 2), procaine benzylpenicillin associated with dihydrostreptomycin nafcillin (Drug 3), and benzathine cloxacillin (Drug 4). We collected eight cow's milk samples that were negative after a bacteriological examination. Macroscopic evaluation of milk samples was made with a Tamis Test and milk cellularity was indirectly estimated with California Mastitis Test (CMT). Milk samples were centrifuged and cells were resuspended with Phosphate Buffered Saline and RPMI 1640. Cell suspensions were submitted to a pre-incubation viability evaluation through Trypan Blue exclusion test and adjusted to 2 x 6 10 cells/mL, before incubation. Anti-mastitis drug was previously diluted (1:1) in RPMI 1640. Subsequently, 1/100 of the dosage indicated to a mammary quarter, as recommended by the manufacturer, was added to adjusted cell suspensions. Cells were

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incubated for one hour at 37 oC. After this period, samples were submitted to a post-incubation viability evaluation through Trypan Blue exclusion test. Mean pre-incubation viability rate (± standard deviation) of milk cells was 50.17% (±6.50). Mean post-incubation viability rates (± standard deviation) of control group (devoid of any drug), Drug 1, Drug 2, Drug 3, and Drug 4 were, respectively, 31.36% (± 3.61); 26.23% (± 4.49); 7.82% (± 2.33); 3.10% (± 0.94); and 30.33% (± 4.14). Results show that all treatments decreased milk leukocytes viability rates (p<0.05). In addition, the uppermost impairments were found when milk cells were challenged with Drug 2 and Drug 3 (p<0.05). Although antimicrobial activity of tested drugs was not appraised, and a single aspect of cell function was tested, we conclude that treatments 2 and 3 could prejudice functionality of milk leukocytes and could provide an ineffective immunological response, which is indispensable to oppose mammary infections. Key words: bovine, milk, dry period, treatment, viability 632 Prevalence of Selected Pathogens of Mammary Gland in Cattle in the Czech Republic during Years 2003-2007 and their Sensitivities to Cephalexin J. Bzdil State Veterinary Institute, Special microbiology, Olomouc, Czech Republic Objective: The objective of our work was to update data related to prevalence of 4 contagious pathogens of mammary gland in cattle (S. aureus, S. agalactiae, S. dysgalactiae and Corynebacterium spp.) appearing in CZ during years 2003 - 2007 from 15384 milk samples examined. Concurrently with the observation of the occurrence of these pathogens, also their sensitivity to cephalexin (cephalosporin of 1st generation) has been tested. The cephalexine is being often used by Czech veterinarians due to its suitable medicinal form, easy administration, easy penetration through cell barriers and relatively low price. Materials and methods: The cultivation of all samples has been performed on blood agar, Endo agar and Edwards´ agar and after following 48 hours incubation the isolation and identification of suspect strains by biochemical tests has been performed. The sensitivitiy tests have been undertaken by common disc diffusion method while using Mueller-Hinton agar and testing antibiotic discs. Results: There were 2992 strains of S. aureus, 837 strains of S. agalactiae, 1015 strains of S. dysgalactiae and 396 strains of Corynebacterium spp. isolated in this period. Prevalence of S. aureus, except temporary decrease in 2005, stagnates at a level around 20%. For S. agalactiae the increase of prevalence has been found in 2004 and since 2005 we observe a slight decrease: the prevalence has been ranging between 2 and 8% during the observed period. For S. dysgalactiae the prevalence is stable and is ranging around a level of 6%. The prevalence for Corynebacterium spp. is relatively steady and low and is ranging between 1 and 4%. The sensitivities to cephalexin are for all above mentioned strains very high: 99% for S. aureus, S. agalactiae and S. dysgalactiae and 97 % for Corynebacterium spp. Conclusion: The observation of prevalence of the above mentioned pathogens of mammary gland in cattle will allow to veterinarians

and service companies in the Czech Republic better evaluate possible causes of mastitis, proposal of effective preventive arrangements and possible choice of appropriate treating programmes. The published data can be also used by foreign specialists for the prevalence comparision of mammary gland pathogens in cattle and it can be a source for other publications.Our work can also provide veterinarians an overview of possibilities for using cephalexin in treating mastitis in cattle. 633 The Phenotypical Correlations between Somatic Cell Count and Principals Characters of Cow Milk Production for Suceave County M. Ivancia , C. Pascal , I. Gilca 1 University of Agricultural Sciences and Veterinary Medicine, Animal Breeding, Iasi, Romania 2 University of Agricultural Sciences and Veterinary Medicine, Sheep And Goats Technology, Iasi, Romania 3 University of Agricultural Sciences and Veterinary Medicine, Animal Husbandry & Dairing, Iasi, Romania There is a well known the fact that animals with high milk productions are more sensibility to mastitis appearance. Also, there is known high positive correlation between milk somatic cell count and animals sensibility to mastitis (0.6 ­ 0.8). So, many experts have been concluding there is correlation between somatic cell count and milk production, fat percent and protein percent. The values of these correlation coefficients (between somatic cell count and the others characters) are unknown until now. Researches were making on cow milk samples from three areas in Suceava county: Dornelor, Radauti and Cîmpulung Moldovenesc areas.Studied characters were milk production, fat percent, protein percent, and somatic cell count.There were analyzed the milk samples with SOMACOUNT to Dorna Lactate and dates have been discussed with MATLAB program.The phenotypical correlation values have been varied between +0,207 ± 0,710 for fat percent and protein percent; -0,327± 0,233 for fat percent and somatic cell count, -0,345 ± 0,046 for protein percent and somatic cell count; -0,676 ± 0,292 for milk production and fat percent; -0,422 ± 0,355 for milk production and protein percent and 0,418 ± 0,500 for milk production and somatic cell count for Dornelor area. For Radauti area, the phenotypical correlation values have been varied between +0,236 ± 0,610 for fat percent and protein percent; -0,115 ± 0,166 for fat percent and somatic cell count, -0,192 ± 0,252 for protein percent and somatic cell count; -0,261 ± 0,053 for milk production and fat percent; -0,564 ± 0,005 for milk production and protein percent and -0,295 ± 0,401 for milk production and somatic cell count. Same characters were studied in Cîmpulung Moldovenesc area, too. The phenotypical correlation values have been varied between +0,234 ± 0,757 for fat percent and protein percent; -0,293 ± 0,295 for fat percent and somatic cell count, -0,441 ± 0,052 for protein percent and somatic cell count; -0,346 ± 0,228 for milk production and fat percent; -0,290 ± 0,057 for milk production and protein percent and -0,282 ± 0,484 for milk production and somatic cell count. Key words: milk cow, phenotypical correlation, fat content, protein content, somatic cell count

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13 National Milk Quality Assurance for Paratuberculosis in the Netherlands M. Weber, P. Franken GD Animal Health Service, Ruminant Health Division, Deventer, Netherlands In January 2006, a bulk milk quality assurance programme (BMQAP) for paratuberculosis in Dutch dairy herds was initiated. The aim of the BMQAP is to reduce the concentration of Mycobacterium avium subsp. paratuberculosis (Map) in milk delivered to the milk factories. The BMQAP was based on modelling studies presented at the 24th WBC. The BMQAP is run alongside the pre-existing Intensive Paratuberculosis Programme (IPP), which aims at low-risk trade of cattle between herds. To date, ~1200 herds have joined the BMQAP, along with ~500 herds participating in the IPP. The BMQAP starts with an initial assessment including a single herd examination. Test-negative herds enter a surveillance procedure consisting of biennial herd examinations ("green herds"). Test-positive herds enter a control procedure consisting of annual herd examinations and culling of test-positives ("red herds"). All herd examinations are done by ELISA. In the initial assessment and surveillance procedures, ELISA results may be confirmed by faecal culture (FC). Recently, the dairy industries have taken the lead to extend participation in the paratuberculosis programmes to all ~21,000 Dutch dairy herds. As a result, a first cohort of new participants is currently entering the BMQAP. The aim of this paper is to present the road to nation-wide participation in the

74 XXV. Jubilee World Buiatrics Congress 2008

paratuberculosis programmes, and to provide an update on the results of the BMQAP. Key words: paratuberculosis, certification, surveillance, milk quality 14 Mycobacterium Avium Paratuberculosis Invades the Human Gut Epithelium and Elicits Local Inflammatory Response Implications for the Pathogenesis of Crohn's Disease N. Shpigel , L. Golan , A. Livneh , I. Rosenshine 1 The Hebrew University of Jerusalem, Koret School of Veterinary Medicine, Rehovot, Israel 1 The Hebrew University of Jerusalem, Faculty of Medicine, Jerusalem, Israel Crohn's disease (CD) is a chronic debilitating inflammatory bowel disease (IBD) of unknown etiology whose incidence is on the rise worldwide. Mycobacterium avium paratuberculosis (MAP) is the etiology of inflammatory bowel disease of ruminants, which bears similarity to human Crohn's disease (CD). The role of MAP in CD is highly controversial, it was found in the gut and blood of CD patients, but causality was not established. To this end we tested the hypothesis that MAP can colonize the normal human gut and elicit inflammation and tissue damage. Fully developed, germ-free human small intestine and colon were established by subcutaneous transplantation of human fetal gut into SCID mice. A total of 83 intestinal xenografts originating from 9 different fetal donors were allowed to develop for 3-4 months and thereafter were infected by direct intraluminal inoculation of a bovine strain of MAP. Using immunohistochemical methods we have found that in all human donors MAP actively invaded the human gut epithelium, with specific tropism to the goblet cells of the small intestine. In 3 out of 9 human donors, invading MAP induced severe tissue damage and inflammation characterised by massive recruitment of macrophages and neutrophils into the gut mucosa producing high levels of IL1, TNF and IL6. These observations implicate that MAP can specifically colonize the normal human small intestine and elicits inflammation and severe mucosal damage. Goblet cell infection by MAP might play a key role in the pathogenesis of CD. Furthermore, this mouse model will enable to analyze MAP virulence mechanisms in the human gut and possible involvement of MAP in CD. Key words: paratuberculosis, Crohn's disease, xenografts 15 Isolation and Characterization of Verocytotoxin-producing Escherichia coli O157 from Turkish Cattle O. Aslantas , S. Erdogan , Z. Cantekin , I. Gulacti , G. Evrendilek 1 Mustafa Kemal University, Microbiology, Antakya, Turkey 2 Firat Univesity, Department of Virology, Elazig, Turkey 3 Mustafa Kemal University, Food hygiene and Quality Control Department, Antakya, Turkey

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Faculty of Veterinary Medicine, University of Tehran, Epidemiology, Tehran, Iran

Leptospirosis is an infectious disease caused by pathogenic members of the genus Leptospira. It affects both man and animal worldwide resulting in high morbidity and mortality. Infection in domestic animals especially castles can lead to economic loss and pose a potential spread to the communities. A serological study was conducted in order to investigate the present status of Leptospirosis in cattle in Bouyer Ahmad district in western Iran during June to October 2006. A total of 236 samples were collected randomly from indigenous cattle of different age and sex; and were tested for antibodies against 6 different Leptospira interrogans serovars (Gripptyphosa, Icterrohamoragia, Canicola, Hardjo, Ballum and Pomona) using a microscopic agglutination test (MAT). The results of this study showed that 30 (12.7%) of the samples had a positive reaction against one or more serovars. The most prevalent Leptospira serovar were Gripptyphosa (7.2%), Canicola (1.7%), Icterrohamoragia and Hardjo (1.3%). The less prevalent Leptospira serovar (0.4%) was Pomona. In this survey association between serum positive samples and exposure to rodents or dogs was statistically significant (P < 0.05). This prevalence study indicated there is a high prevalence of Leptospirosis in this region. Control of this disease consists of implementing biosecurity measures, use of antibiotics to clear carrier states and use of effective vaccines. Key words: seroepidemiology, leptospirosis, cattle, Bouyer Ahmad, Iran 46 Isolation of Clostridium difficile from Veal Calves L. Arroyo , A. Van Dreumel , R. Lothrop , H. Staempfli , J. Weese 1 Ontario Veterinary College, Department of Pathobiology, Guelph, Canada 2 University of Guelph, Guelph, Canada 3 Ontario Veterinary College, Clinical Studies, Guelph, Canada 4 Private, Cambridge, Canada Clostridium difficile is an important enteropathogen in humans and some domestic animals. It has been isolated and its toxin(s) detected in feces of diarrheic and non-diarrheic dairy calves, yet its role as a cause of disease in veal calves has not been investigated. This study describes the pathological findings of suspected C. difficile-associated gastroenteritis in veal calves and characterizes C. difficile isolates obtained from veal calves. Six calves were submitted for necropsy examination from a veal farm. Gastrointestinal contents were screened for pathogens associated with calf diarrhea, including enterotoxigenic E. coli, Salmonella spp. C. perfringens, viruses and parasites. Samples from 2 calves were cultured for C. difficile. Samples were tested for C. difficile toxins A/B and C. perfringens enterotoxin using an ELISA and C. difficile toxins were detected in all cases. No other pathogens were identified. Gross lesions were similar in all cases and consisted of fibrinous enteritis, colonic edema, hemorrhagic enterocolitis, dehydration, and pulmonary congestion and edema. Histologically, there were focal areas of mucosal erosion and fibrino-cellular exudates, with colonies of clostridia-like bacilli in the lumen and on the mucosal surfaces of the small intestine and abomasum. There was transmural edema and focal areas of hemorrhage in the lamina propria, with congested and thrombosed capillaries. Fecal samples were collected at the farm from 24 diarrheic calves for C. difficile culture. Three historic isolates recovered from diarrheic calves 4 years earlier from the farm were also analyzed. PCR-ribotyping and screening for genes encoding toxins A (tcdA), B (tcdB) and binary toxin (cdtB) were performed. Clostridium difficile was isolated from the 2 initial calves and 22/24 (92%) diarrheic calves. Five toxigenic ribotypes were identified from the 27 isolates. Sixteen (57%) possessed genes tcdA and tcdB, while 9 (32%) only possessed genes encoding tcdB and 2 (7.1%) possessed tcdB and cdtB genes. Overall, genes encoding tcdA, tcdB and cdtB were present in 16 (59%), 27 (100%) and 2 (7%) strains, respectively. Two ribotypes accounted for 78% of isolates. This report supports the potential capacity of C. difficile to colonize and cause disease in several animal species. Further studies of diarrheic and normal veal calves are required to elucidate the role of this pathogen as a cause of gastroenteritis and diarrhea in veal calves. 47 Severe Outbreaks of Botulism in Cattle Herds in Flanders : 4 Case Reports M. Goderis, M. Hostens, G. Opsomer Ghent University, Dep Reproduction, Obstetrics and Herd Health, Merelbeke, Belgium

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The objective of this study was to collect rectal swabs from the cattle in a slaughterhouse located in Hatay (Turkey) immediately after slaughter for the isolation and characterization of verotoxin-producing Escherichia coli 0157 in each month during a 1-year period. The rectal swab samples were analyzed for the isolation of E. coli 0157 through pre-enrichment, immunomagnetic separation and selective plating on CT-SMAC agar. E. coli 0157 was isolated from 77 (13.6%) of the samples. The presence of E. coli 0157 changed during a 1-year period, in that the occurrence of E. coli 0157 was the highest in July and November and lowest in February. A total of 66 isolates out of 77 were seroytpe 0157:H7 and 11 were serotype 0157:NM. PCR analysis of E. coli 0157 virulence genes revealed that all 0157:H7/NM were positive for rbf(O157), 74 positive for EhlyA, 72 positive for eaeA, 62 positive for vtx2, and 3 positive for both vtx1 and vtx2. It was presented by cytotoxicity tests that many of E. coli 0157 isolates showed high cytotoxicity on Vero cells. All of the isolates containing EhlyA showed enterohaemolysin production. Key words: Escherichia coli O157, Turkey, cattle, IMS, PCR 16 A Seroepidemiological Study of Bovine Leptospirosis in Bouyer Ahmad District, Western Iran E. Rahimikia , G. Abdollahpour , E. Mostafavi 1 Faculty of Agriculture, Azad University of Yasooj, Animal Science, Yasooj, Iran 2 Faculty of Veterinary Medicine, University of Tehran, Clinical Science, Tehran, Iran

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In Belgium, an increasing amount of catastrophic botulism outbreaks in cattle herds have recently been reported to the Belgian Federal Agency for the Safety of the Food Chain (FASFC). The latter confirms that at least in Belgium botulism is becoming an emerging disease in cattle resulting in large economical loss for the farmers. This presentation describes 4 major outbreaks in the Flemish part of Belgium occurring between July 2005 and February 2007. The average mortality on all farms varied from 27 to 96 %. The first case involved a mixed dairy-poultry farm that encountered a morbidity of 33 % and mortality of 27 %. Two possible causes of intoxication have been proposed. One being a severe rainfall spreading the toxins from the poultry litter to the dairy stable and maize silage, the other is an infected grass silage after fertilization with poultry litter. C. botulinum intoxication was diagnosed by identification of C. botulinum type C and D toxins in the faeces and the liver of the affected animals and in samples of the maize silage. The second dairy farm reported a morbidity and mortality of respectively 51 % and 49 %. Here, the diagnosis was confirmed by identification of the C. botulinum type D toxin in liver and mixed-forage samples. A chicken cock housing a nearby water well that provided the drinking water for the cows was found positive for the same type of toxin, and was hence seen as the main contributor to the catastrophe. On the third farm, a mixed swine-dairy herd, the highest morbidity and mortality was seen, being respectively 100 % and 96 %. The farm had one ensilaged grass pack in which possibly a wild animal was encaptured while moving. A sample of this grass silage was positive for the C. botulinum type D toxin. The fourth poultry-dairy mixed farm previously piled up poultry litter on the place where actually the wet brewer grains fed to the dairy cows were stored. On a total of 85 cows, 67 died leading to a mortality of 79 %. Neurotoxin type D was isolated in a wet brewer grain sample leading to the C. botulinum diagnosis. The presented outbreaks here encouraged Belgian farmers, veterinarians, the Flemish Animal Health Service (DGZ-Vlaanderen) and the FASFC on describing risk factors for botulism more profoundly and elaborated a strategy leading to a quicker diagnosis and a better approach of a C. botulinum outbreak in cattle. These will be discussed in detail. Key words: Clostridium botulinum outbreak, cattle, poultry 48 Molecular Mechanism of Action of Pentosan Polysulfate on the Cellular Prion Protein V. Stadnyk, C. Mayor,V. Vlizlo Institute of Animal Biology UAAS, Scientific Center for Prion Infections Study, Lviv, Ukraine Objectives of study: It was shown by our previous researches, that pentosan polysulfat has the ability to inhibit the expression of cellular C prion protein (PrP ) in the prion-replicative organs of laboratory animals, that it can be used for the prophylaxis of prion infections, in particular C BSE. But the molecular mechanism of action of this preparation on PrP until now is not clear. The aim of our work was to find out the molecular C mechanisms of pentosan polysulfate influencing on PrP . Materials and Methods: for the research of co-operation of pentosan C polysulfate and PrP were used the ELISA method, mapping of amino C acid residues of PrP , which co-operate with pentosan polisulfate were carried out by bioinformatical methods and UV-spectroscopy. C Results: It is set that PrP and pentosan polysulfate specifically cooperate in vitro. Except this, it is shown that for this co-operation the sulfate groups of pentosan are responsible and aminogroups of Arg148 and Arg151. Conclusions: it is set that inhibiting influence of pentosan polysulfate C on PrP is predefined, probably, by the direct co-operation of these molecules. Key words: prion protein, pentosan polysulphate 49 Distribution of Abnormal Prion Protein in the Brain of Goats E. Bouzalas , C. Dovas , S. Kritas , M. Papanastasopoulou , 2 1 1 D. Papakostaki , E. Chatzinasiou , G. Koptopoulos 1 Aristotle University of Thessaloniki, School of Veterinary Medicine, Department of Microbiology and Infectious Diseases, Thessaloniki, Greece 2 Veterinary Institute of Infectious and Parasitic Diseases, Thessaloniki, Greece The objective of this study was to assess the distribution of PrP in the brain of goats raised in a flock with high incidence of scrapie. Scrapie is a progressive fatal neurodegenerative disease affecting both sheep and goats. Its diagnosis is mainly based on the detection of the

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abnormal prion protein (PrP ) in the medulla oblongata at the level of the obex. The study was undertaken in a semi-extensive flock of goats comprised of 250 animals, 105 of which were older than 12 months. All animals were euthanized and samples from medulla oblongata were taken from 86 out of the 105 goats. Additional samples were collected from pons, cerebellum, midbrain at the level of superior colliculus, diencephalon at the level of optic chiasm and frontal lobe of cerebrum Sc to study the distribution of PrP in these animals. The samples were examined by a rapid ELISA (Enzyme-Linked-Immunosorbent-Assay) test (EC No 999/2001) and an immunochromatography assay and positive samples were additionally confirmed with Western blot (WB) and/or immunohistochemistry (IHC). Examination of the obex with the Sc rapid ELISA test revealed the presence of PrP in 12 goats (14%). Five of them had shown also clinical signs such as disorientation, ataxia, apathy and sometimes aggressiveness. In 10 out of those 12 animals, Sc PrP was detected by both methods in other parts of the brain besides Sc obex. Interestingly, in 3 goats other than the obex positive ones, PrP was found in other parts of the brain and mainly in the hypothalamus (diencephalon). It was concluded that these findings might question the use of obex as the exclusive sampling site for detecting the presence of Sc PrP in goats. Sc Key words: scrapie, PrP , distribution, brain, goats 50 Dynamics and Distribution of Prion Infectivity in Sheep Blood C. Lacroux , N. Morel , S. Simon , H. Cassard , F. Corbiere , 1 1 3 2 1 J. Mathey , F. Schelcher , F. Lantier , J. Grassi , O. Andréoletti 1 INRA ENVT UMR 1225, Interactions hôtes-agents pathogénes, Toulouse, France 2 CEA, SPI, Saclay, France 3 INRA, IASP, tours, France In sheep prion transmission has been observed by transfusion of whole blood sampled at the end of the first half of the incubation phase. However, in this species, dynamics and distribution of the infectivity presence in blood remained poorly documented. We first characterized infectivity dynamics and distribution in blood from sheep experimentally challenged by the oral route with a "rapid" strain. In this model clinical onset is observed as soon as 160 days post challenge. Inoculated animals were sampled every month and blood was fractionated into buffy-coat, Ficoll cells and plasma. Moreover T CD4 and CD8 lymphocytes, B lymphocytes (CD45r), Mono-nucleated phagocytes (CD14) and Negative fraction (containing CD4-, CD8-, CD45r- and CD14- cells) were sorted from Ficoll preparations. Each fraction was immunoprecipitated before concentration under small volume and IC inoculation to Tg338 mice (transgenic ovine PrP gene). Bioassay revealed that, infectivity (i) can be detected in this model as early as D60 post challenge (ii) is mainly linked to CD14 positive cells. Moreover this experiment provided elements indicating that infectivity presence in blood is correlated to PrPsc presence in secondary lymphoid tissue. In a second experiment, we investigated sheep naturally infected with scrapie (Langlade model). In this model, blood transfusion allowed to transmit disease as early as 3 months of age and infectivity presence in blood persisted all along the incubation phase. Moreover we demonstrated that animals contracting scrapie through transfusion were contaminant (through blood) 3 months post transfusion. In this model first results of bioassay in Tg338 mice confirmed that most infectious blood fractions are the mononucleated cells and more particularly CD14. Finally, the efficiency of contamination by blood transfusion was established by comparison to IV inoculation using a titrated (in Tg338 mice) inoculum. Results revealed that a 200 ml whole 5 blood transfusion was as efficient as an IV inoculation with 510 DL50, in contrast with the low infectivity level observed by direct blood bioassay in mice. This element could suggest that infectivity in whole blood is supported by a highly efficient mechanism. Both dynamics and nature of the infectivity positive fractions are of interest with regards to (i) the development and evaluation of diagnostic tests on live animals and (ii) human blood sanitary policy. Key words : prion, scrapie, sheep, blood, infectivity 51 Retrospective Study of BSE Surveillance in Canada from 1992 to 2005 J. Kellar , J. Paré , L. Myers 1 Canadian Food Inspection Agency, Animal Products Directorate, Ottawa, Canada 2 Canadian Food Inspection Agency, Epidemiology and Surveillance Section, Saint-Hyacinthe, Canada

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Canadian Food Inspection Agency, Epidemiology and Surveillance Section, Ottawa, Canada

Bovine Spongiform Encephalopathy (BSE) was first diagnosed in Canada in 1993, in a cow imported from the United Kingdom. In 2003, the first indigenous case of BSE was diagnosed. To the end of 2007, a total of 12 indigenous cases of BSE had been diagnosed in cattle born in Western Canada. From the time Canada made BSE reportable in 1990, surveillance has evolved from passive to active, with a markedly greater intensity following the discovery of the first indigenous case. To assess Canada's true prevalence of BSE and the effectiveness of its surveillance, a retrospective study was designed based on the random selection of 3,712 samples from the 95,383 cattle brain samples submitted for BSE testing between 1992 and 2005. The sampling plan was based on an a priori estimation of the proportional distribution of National, Western and Eastern BSE surveillance samples by Office International des Epizooties (OIE) surveillance stream (clinical suspect, casualty slaughter, fallen stock, and routine slaughter). For each sample, data was obtained by record tracing and interviewing of identifiable current and original owners. Information recorded comprised sampling site and date, age of the animal, date of birth, breed, production type, province of sampling, province of birth, clinical signs preceding and reason for death, diagnostic test used and test result. Each sample was classified by OIE surveillance stream. Analyses on a National, Western and Eastern basis highlighted regional variability among surveillance strategies. Reflecting underlying population demographics and market flows, Eastern Canadian surveillance emphasized deadstock/renderer sampling (70.5%) while surveillance in Western Canada focused on on-farm sampling (84.1%). Sampling in Western Canada was predominantly from beef cattle (78.3%), with a mean age at sampling of 8.1 years. In Eastern Canada, the majority of samples originated from dairy cattle (84%), with a mean age at sampling of 4.7 years. The majority of samples examined were classified as casualty slaughter (63.7% in Eastern Canada and 80.4% in Western Canada). National, Eastern Canada and Western Canada prevalence estimates were determined. Results of the study have contributed to a broader assessment of the natural history of BSE in North America. 86 Pregsure Trivalent Induces Much Higher Neutralising Antibody Titres against BVDV-2 Strains than Monovalent BVD Vaccines R. Raue, D. Hurst, J. Salt Pfizer Animal Health, R&D, Sandwich, United Kingdom Objectives: Bovine viral diarrhoea virus (BVDV) is one of the most common and economically important viruses causing infections in cattle. Recently, severe outbreaks of BVDV-2 have been described in the UK. In addition, isolation of BVDV-2 has been reported in most European countries. As vaccination against BVDV is most often performed with inactivated vaccines, neutralising antibodies can be a good in vitro indicator of the ability to protect against a certain BVDV strain or type. This study was designed to compare the neutralisation antibody titres induced after vaccination with a novel vaccine PregSure Trivalent (Pfizer Animal Health), Bovilis BVD (Intervet) and Bovidec (Novartis Animal Health) against a panel of BVDV-1 and 2 strains. Materials and Method: Serum samples were collected from calves 3 weeks after vaccination either with PregSure Trivalent, Bovilis BVD or Bovidec according to the recommendations of the manufacturer. They were tested in a serum neutralisation test against 11 BVDV-1 strains, representing genotypes 1a, 1b and 1d to 1i, as well as 4 BVDV-2 strains from Europe and the US. Results: The geometric mean titre in log2 for the 11 BVDV-1 strains were between 7.3 and 10.8 for PregSure Trivalent, 5.7 and 9.5 for Bovilis BVD and 4.0 and 8.2 for Bovidec. With all vaccine sera BVDV-1h strain G-AU was neutralised best and BVDV-1d strain PI810 worst. The geometric mean titre in log2 for the 4 BVDV-2 strains where between 8.5 and 10.1 for PregSure Trivalent, 1.9 and 5.1 for Bovilis BVD and 1.2 and 2.2 for Bovidec. Therefore, PregSure Trivalent serum samples showed an equal ability to neutralise BVDV1 and 2 strains whereas Bovilis BVD and Bovidec serum samples showed a reduction in the ability to neutralise BVDV-2 strains by at least 4 log2. In some cases Bovilis BVD and Bovidec vaccine sera were not able to neutralise BVDV-2 strains at all. Conclusions: The results of this study demonstrate that the level of neutralising antibodies after vaccination with PregSure Trivalent is

remarkably higher against BVDV-2 strains than those of monovalent inactivated BVD vaccines. In combination with the demonstration of 12 month protection against BVDV-1 and 2 by challenge, this data indicate that the novel vaccine PregSure Trivalent gives a comprehensive cover against all types of BVDV field infections. Key words: PregSure Trivalent, BVDV-1, BVDV-2, neutralisation 87 Monitoring of a BVDV Infection in a Vaccinated Herd by Testing of Milk for Antibodies against NS3 B. Makoschey , H. Kuijk , J. Mars , W. bij de Weg , P. Franken 1 Intervet-Schering Plough, Int. Marketing, Boxmeer, Netherlands 2 Intervet-Schering Plough, Intervet Netherlands, Boxmeer, Netherlands 3 Animal Health Service Deventer, Deventer, Netherlands Objectives: It has been demonstrated earlier, that an inactivated vaccine against the bovine viral diarrhoea virus (BVDV) in combination with a suitable test for antibodies against non structural proteins (NS) exhibits properties of a marker vaccine: animals usually remain seronegative after vaccination but develop NS specific antibodies after field virus infection. In this study, it should be established, whether the NS3 antibody testing of individual or bulk milk samples is likewise suitable for differentiation between BVDV vaccinated and field virus infected animals. Materials and Methods: The study was performed on a Dutch dairy farm. After being certified as BVDV-virus-free in 2002, the farm experienced a new outbreak of BVDV in 2005. A herd vaccination R program was initiated using an inactivated BVDV vaccine (Bovilis BVD, Intervet International). Seventeen cows were selected for further follow up: blood samples, individual milk samples and bulk milk samples were collected at pre-set time points. BVDV neutralizing antibody titers were measured and the NS3 antibody response was measured in a commercial ELISA test (Cedi-testR BVDV, Cedi Diagnostics, Lelystad, The Netherlands). Results: All animals that were seronegative before vaccination developed BVDV neutralizing antibodies after immunization. Three of the ten vaccinated animals showed an increase in the titers of BVDV neutralising antibodies, between study days 56 and 210, indicating that they experienced a field virus infection. All samples from animals that were vaccinated but not infected were tested negative for NS3 specific antibodies at all time points. The samples from the three animals that were infected after the vaccination were tested negative for NS3 specific antibodies at Day 0, 28 and 56, but were positive thereafter. In contrast, all serum samples taken from animals that have been infected with BVDV field virus were tested positive for NS3 specific antibodies. The overall majority (37/44) of the milk samples for these animals were also tested positive in the NS3 ELISA. Likewise, the bulk milk samples were positive in the NS3 ELISA, with the exception of the sample taken at study day 28. Conclusions: The results obtained demonstrate that a field virus infection with BVDV can be monitored by measurement of NS antibodies in blood or (bulk) milk samples, even in a vaccinated herd. 88 Transmission of Bovine Viral Diarrhoea Virus (BVDV) from Persistently Infected Calves to Naive in-contact Cattle is Very Rapid and Efficient B. Makoschey , M. Janssen 1 Intervet-Schering-Plough, Int. Marketing, Boxmeer, Netherlands 2 Intervet-Schering Plough, R&D, Boxmeer, Netherlands Objectives: The transmission of bovine viral diarrhoea virus (BVDV) from persistently infected (PI) calves to in-contact animals is generally considered to happen very efficiently. It should be determined, whether naive animals housed in adjacent pens to a PI are readily infected or whether housing in the same pen is required for efficient infection. Materials and Methods: Four heifers at the age of 10 months or older, negative for antibodies against BVDV, and free of BVDV and one three months old PI calf were included in the study. The heifers were housed as one group in a pen. The PI calf was housed in an adjacent pen. The animals were allowed to have direct contact through the fence. Blood samples were taken weekly and BVDV neutralising antibodies were determined. Results: All four heifers seroconverted within four weeks after introduction of the PI calf. These results, indicate that the infection with BVDV had occurred during the first week after the first contacts with the PI animal.

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Infectious and Zoonotic Deseases (Public Health) 77

Conclusion: It can be concluded that transmission of BVDV from PI animals to naive in-contact animals occurred in a very efficacious and fast manner, even if the animals are housed in adjacent pens. The practical implications of these findings for BVDV control programs will be discussed. 89 BVDV Type 1 and 2 Fetal Protection following the Use of a Modified Live Viral Vaccine W. Xue, D. Mattick, L. Smith, J. Maxwell Intervet-Schering Plough, R&D Desoto, Desoto, United States Objectives: Previously, it has been demonstrated that pre-breeding R vaccination with a modified live virus (MLV) vaccine (Vista 5 SQ, Intervet International) containing BVDV Type 1 and Type 2 strains, offers protection against persistently infected fetuses when dams are challenged with BVDV type 1 and type 2 in the first trimester. The animal trials presented here were designed to demonstrate the efficacy of the MLV vaccine in protecting fetuses from infection with BVDV type 1 or type 2, when pregnant cows were challenged in the last trimester, at 170 days of gestation. Materials and Methods: Heifers, seronegative to BVDV, were vaccinated at pre-breeding, and then bred naturally 4 weeks postvaccination. Eighty-three pregnant heifers were challenged with either type 1 (37 in total with 25 vaccinated and 12 controls), or type 2 (46 in total with 28 vaccinated and 18 controls) non-cytopathic (ncp) BVDV field isolates at about 170 days of gestation. Fetuses were collected from the challenged heifers 60 days post-challenge (for type 2 BVDV challenge), and newborn calves were collected prior to taking colostrum (for type 1 BVDV challenge). Serum samples were taken from the fetuses or calves for a BVDV serum neutralizing (SN) antibody test. Tissue samples of thymus, lung, spleen and kidney were collected from the fetuses or calves, and were used for virus isolation. Results: Results of SN antibody tests revealed that 100% of fetuses or calves from control heifers developed SN antibody to BVDV. However, only 4 out of 22 calves born to vaccinated heifers were positive for SN antibody in the type 1 BVDV challenge study, and 7 of 28 fetuses from vaccinated heifers were positive for the SN antibody in the type 2 BVDV challenge study. Virus isolation from calves' tissue samples demonstrated the presence of type 1 BVDV in 5 of 12 calves from control heifers and none of the calves from vaccinated heifers. The type 2 BVDV was isolated from 17 of 18 fetuses in the control group, and 2 of 28 fetuses in the vaccinate group. R Conclusions: Results demonstrated that MLV vaccine Vista 5 SQ prevented fetal infection by at least 82% to type 1 BVDV, and 75% to type 2 BVDV, when the dams were exposed to BVDV at about 170 R days of gestation. Vaccine Vista 5 SQ is the first MLV BVDV vaccine that has proven to prevent fetal infection in the last trimester of pregnancy. 90 Fetal Protection against Bovine Viral Diarrhea Virus Type 1 Infection Using One Shot of the Mucosiffa Modified-live Virus Vaccine G. Meyer , M. Deplanche , D. Roux , M. Moulignie , 1 1 3 1 D. Raboisson , N. Hagen-Picard , P. Mathevet , F. Schelcher 1 National Veterinary School of Toulouse, Pathology of Ruminants, Toulouse, France 2 National Institure of Agronomical Research, Animal Health Department, Toulouse, France 3 Merial Animal Health Limited, Lyon, France Objective: To evaluate the efficacy of a commercially available live Bovine Viral Diarrhea virus (BVDV) vaccine to protect against fetal infection in pregnant cattle. Method: Thirty beef heifers were allocated into 2 groups. One group (18 heifers) was vaccinated against BVDV with one shot of a R commercially (Mucosiffa , Merial) live vaccine according manufacturer's recommendations, and the other group (12 heifers) 3.5 served as nonvaccinated control cattle. The vaccine contained 10 TCID50 per vaccination dose (2.0 ml) of cp BVDV-1 Oregon C 24 V strain. One month later, estrus was induced and the heifers were bred. Three cycles of insemination were performed. Pregnancy was confirmed by transrectal ultrasonographic examination and bovine pregnancy-associated glycoprotein quantification. Four months after

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vaccination, 13 vaccinated and 9 non vaccinated pregnant heifers were 6 intranasally challenged with 10 TCID50 of the BVDV-I 22146/Han strain, on days 49-96 of gestation. Seroconversion, clinical signs, mortality, abortion rate, hematology, viremia, and fetal infection were studied. Results: By ELISA or neutralization assay, all vaccinated animals seroconverted to BVDV four weeks after immunization. Neutralizing antibodies reached a plateau two month after vaccination with titers ranging from 5970 to 14159 ED50/ml. Challenge was followed by a rise of 0.57 log of neutralizing antibody titer for vaccinated animals and by seroconversion for control heifers. Post-challenge, all heifers were free from clinical signs of BVD. Viremia was not detected by real time RT-PCR for 18 days post-challenge in any of the vaccinated heifers. In contrast, 100% of the control animals were BVDV viremic on at least one day between days 4 to 18 post-challenge. Transient leucopenia with lymphopenia was observed for heifers of the control group but not of the vaccinated one. BVDV type 1 infection led to abortion only in one unvaccinated heifer, 47 days after challenge and virus was detected by real time RT-PCR in the spleen of aborted fetus. Three months after challenge, all fetuses were harvested from heifers and tested for evidence of BVDV infection. BVDV was detected with high viral charges by qRT-PCR in the spleen, thymus, cerebellum and placenta from 8 of 8 fetuses obtained from control heifers but from none of the fetuses obtained from the 13 vaccinated heifers. R Conclusion: The commercial monovalent Mucosiffa live vaccine is safe and fully protective against fetal BVDV type 1 infection and abortion. Key words: bovine viral diarrhea, vaccination, fetal protection 91 Impact of Live and Killed Marker IBR Booster Vaccinations on BoHV-1 Shedding after Primovaccination with Live Marker IBR Vaccine M. Pearce, I. Vangeel Pfizer Animal Health, Sandwich, United Kingdom For BoHV-1 control it is important to minimise virus shedding from infected cattle. It has been suggested that killed vaccines are more effective at suppressing BoHV-1 shedding than live vaccines. This study compared the impact of live and killed marker IBR booster vaccines on BoHV-1 shedding after challenge and reactivation following primovaccination with live marker IBR vaccine. Twenty BVDV free calves aged 12-14 weeks with no antibodies against BoHV-1 and BVDV were given 2 doses of live IBR vaccine IM 3 weeks apart. Six months after primovaccination, 10 calves received one IM dose of live IBR vaccine; the other 10 calves received one dose of killed IBR vaccine. Live vaccine was Rispoval® IBR marker vivum; killed vaccine was Rispoval® IBR marker inactivatum. Ten control calves received saline IM at the time of each vaccination. Six months after booster vaccination all cattle were challenged 8.0 intranasally with 10 TCID50 BoHV-1, Iowa strain. Six weeks after challenge, calves were given dexamethasone at 0.1 mg/kg bodyweight IV daily for five days. Cattle were blood sampled weekly for three weeks after booster vaccination and tested for neutralising antibodies against BoHV-1. After challenge and reactivation, nasopharyngeal swabs were taken daily for 21 days and tested for the presence of BoHV-1 by virus isolation. Boosting with inactivated vaccine gave higher titres of BoHV-1 neutralising antibodies (P<0.05) 14 and 21 days after booster vaccination. Log2 peak geometric mean titres were 13.2 in cattle boosted with inactivated vaccine compared with 11.6 in cattle given live vaccine. Vaccination reduced duration of shedding after challenge (P<0.001), and reduced total virus shed (P<0.001) and duration of shedding (P<0.001) after reactivation. After challenge, there was no significant difference in peak shedding, total virus shed or duration of shedding between the two vaccinated groups. After reactivation no detectable viral shedding occurred in cattle given the killed vaccine booster, but shedding occurred five and six days after dexamethasone administration commenced (log2 TCID50 0.37 and 2.54 (P<0.05) respectively) in cattle given the live vaccine booster. However, differences in total virus shed and duration of shedding between the two vaccinated groups were not significantly different. In conclusion, there was no demonstrable difference in viral shedding patterns after challenge or reactivation between cattle given live or killed booster vaccinations. Key words: IBR, BoHV-1, vaccine, shedding

78 XXV. Jubilee World Buiatrics Congress 2008

131 Evaluation of the Exhaustion Dynamics of MDA in IBR Cases I. Szabó , T. Molnár , T. Abonyi 1 Pfizer NEER Region, Paris, France 2 MgSzH Diagnosztikai Igazgatóság, Budapest, Hungary 3 Pfizer Ltd, Budapest, Hungary Infectious bovine rhinotracheitis (IBR) causes substantial economic losses in cattle herds. The key elements of successful eradication are: (a) in IBR-free herds, preventing the introduction of infection and maintaining disease-free status, (b) in infected herds, starting an eradication programme as soon as possible, determining the spread of virus within the herd, applying a management-adapted immunisation programme, and breaking the chain of infection in a reliable manner. The knowledge of maternal immunity status is essential for accurately determining the spread of virus and the age group in which basic immunisation should be started. In eradication programmes using marker vaccination, it is important to prevent the infection of the progeny with virulent virus. The duration of maternally derived immunity is extremely important in terms of calf vaccination. The primary objective of this work was to collect data on the duration of maternal immunity in Hungary, where about 75% of the large dairy herds are infected with virulent IBR virus at present. In 17 large cattle herds infected with IBR virus, located in Hungary, blood samples were taken from 5-10 one- to six-month-old calves per age group. The IBR antibody levels of the blood samples were determined by a discriminative ELISA. The results clearly show that in 10% of the herds the maternal antibodies disappear already by 2 months of age, while in 19%, 44% and 73% of the herds the cessation of maternal immunity can be observed at 3, 4 and 5 months of age, respectively. On two farms the maternally derived protection was present even at 6 months of age. The findings on some farms indicate that after the disappearance of maternal antibodies by 3-4 months of age, in the presence of conditions facilitating the spread of virulent virus calves may become infected with virulent IBR virus already by 6 months of age. These findings are important with a view to the successful In conclusion, for the elaboration and successful 132 Differentiation of Malignant Catarrhal Fever, Mucosal Disease and Bluetongue Based on the Retrospective Study of 115 Cases R. Bexiga , H. Guyot , C. Saegerman , A. Mauroy , F. Rollin , 2 1 1 1 1 E. Thiry , A. Philbey , D. Mellor , D. Barrett , K. Ellis 1 University of Glasgow Veterinary Faculty, Animal Production and Public Health, Glasgow, United Kingdom 2 University of Liege, Clinic for ruminants, Liege, Belgium The clinical differentiation of bluetongue (BT) in the form currently seen in northern Europe is highly relevant, especially from malignant catarrhal fever (MCF) and mucosal disease (MD), two endemic diseases in most of Europe. The aim of this study was to characterise the clinical findings typical of cases of MCF and MD, to compare these to clinical findings characteristic of the recent BT outbreak in northern Europe. Cases of MD (n=66) and MCF (n=11) were selected from records of cattle admitted to the University of Glasgow Veterinary School, from January 1989 to June 2007. Selected cases of MD had a postmortem diagnosis of MD, were antigen positive and antibody negative for bovine viral diarrhoea virus (BVDV) by ELISA. Selected cases of MCF had a postmortem diagnosis of MCF, were positive for antibody against ovine herpesvirus-type 2 (OHV-2) by an immunofluorescent antibody test or for OHV-2 nucleic acid by PCR and were antigen negative for BVDV. Cases of BT (n=38) were selected from records of cattle examined by the ambulatory clinic of the Faculty of Veterinary Medicine of Liege from August 2006 to February 2007. Selected cases had a clinical diagnosis of BT and a positive test result either for detection of antibody by competitive ELISA or viral nucleic acid by RTPCR. Case data were summarised to determine the most common presenting signs for each condition. A Freeman-Halton extension of the Fisher exact test was performed to test for significant differences in frequency of clinical signs between the three conditions. Clinical signs for which there was a significant difference in the proportion of cases presenting in each category were appetite, demeanour, size for age, faecal appearance, adventitious lung sounds, corneal opacity, nasal discharge, halitosis, bilateral lymph node enlargement and interdigital ulceration (P<0.001). This significant difference could be attributed largely to BT cases for 4 clinical parameters, which yielded moderate to

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good sensitivities and specificities when evaluating the absence of changes in these categories in cases of BT: appetite (95% Se, 64% Sp), demeanour (74% Se, 79% Sp), faecal appearance (92% Se, 68% Sp) and lymph node size (95% Se, 66% Sp). Based on data available for this study, whenever a clinical case suggestive of one of the conditions studied presents with normal appetite, normal demeanour, normal faecal appearance or absence of bilateral lymph node enlargement, there would be a high suspicion for BT. 133 Evolution of the Clinical Expression of Bluetongue in Belgian Cattle during Year 2006 vs 2007 H. Guyot , A. Mauroy , F. Rollin , E. Thiry , B. Losson , 2 C. Saegerman 1 University of Liege, Clinical Department of Food Animals, Liege, Belgium 2 University of Liege, Department for Infectious DiseasesEpidemiology, Liege, Belgium In August 2006, Belgium notified its first cases of Bluetongue (BT), serotype 8, in cattle and sheep. The disease was also observed at this time in the neighboring countries. The resurgence of BT was observed in Northern Europe in 2007. The aim of the study was to compare clinical signs of BT observed in 2006 vs 2007 in Belgian cattle. The description of clinical signs was based on the observation of 38 and 39 cows in 2006 and 2007, respectively. BT cases were only included if they were confirmed by one or both laboratory diagnostic tests (competitive ELISA test and/or RT-qPCR). The inventory of clinical signs was made with a standardised clinical form for BT. This form is divided into general, cutaneous, locomotor, digestive, respiratory, neurological and reproductive clinical signs. Case data were summarised to determine changes in clinical presentation of BT between 2006 and 2007. A Fischer's exact probability test was performed to compare (P<0.05) the frequency of clinical signs between the two years. Regarding general clinical signs, hyperthermia and tiredness were more often observed in 2007, compared to 2006. All clinical signs about skin and annexes were not significantly different between the two years. Locomotor signs such as prostration, incapacity to get up, reluctance to move, lameness and amyotrophy were more frequent in 2007. Loss of appetite, difficulties in grasping feed, salivation and drooling were the digestive signs more often observed in 2007. A purulent nasal discharge was the only respiratory sign more commonly observed in 2007. Apathy, generalised weakness and paresis or paralysis were more often encountered in 2007. The most important changes between the two years concerned reproduction. A higher incidence of abortion, premature calving and stillbirth was observed during 2007 outbreak. The frequency of most of the clinical signs of BT was higher in 2007 in Belgian cattle. Confirmed cases of BT in Belgian cattle were only 296 in 2006 compared to 4187 in 2007. These data do not represent the real situation of BT infection because the farmers do not notify all cases. Nevertheless, it seems that the 2007 outbreak was more severe regarding the number of cases and the frequency of clinical signs. The mild winter and wet 2007 summer might have favored the persistence of the vectors. Keywords: Bluetongue, cattle, clinical signs, emergence, resurgence 134 Oesophageal Paresis in Cattle Associated with Blue Tongue Serotype 8 Infection B. Pardon , V. Vandenberge , R. Ducatelle , P. Deprez 1 Faculty of Veterinary Medicine - University of Ghent, Department of Internal Medicine and Clinical Biology of Large Animals, Merelbeke, Belgium 2 Faculty of Veterinary Medicine - University of Ghent, Department of Pathology, Merelbeke, Belgium During the last Blue Tongue (BTV) outbreak in Belgium, four calves from four different herds were referred to the clinic with complaints of regurgitation, alimentary nasal discharge, salivation, cough, anorexia, and progressive weight loss since 5 to 7 days. All calves were between 7 and 9 months old and were on pasture in groups of 7 to 20 animals. All cases were found within a 10 kilometres radius. Several practitioners reported similar cases in adjacent regions. Endoscopic examination displayed a dilatation of the distal oesophagus. The lower oesophageal sphincter had a normal tone. The calves received supportive therapy, procaine penicillin and also erythromycin as a prokinetic agent. Two calves died and the other

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Infectious and Zoonotic Deseases (Public Health) 79

calves recovered after 4 to 10 days, respectively. Repeated endoscopic examinations demonstrated that the oesophageal tone gradually normalised. Oesophageal paresis in cattle has been described as a sequel to oesophageal obstruction or caused by metabolic, neurological or neuromuscular disorders. Due to the absence of oesophageal obstruction, hypocalcemia or signs of encephalitis, Clostridium botulinum type B intoxication was initially suspected. In rumen fluid and faeces of one calf, neurotoxins belonging to group C/D and a toxigenic C. botulinum strain belonging to type A,B or E were found. In rumen fluid and faeces of the other calves a substance toxic for mice was isolated, that was not neutralisable by botulinum antitoxins. Because of the BTV serotype 8 epidemic in the region, the calves were tested for BTV by PCR: all calves were antigen positive. At necropsy only aspiration pneumonia of the cranial lung lobes was found. Histology of the oesophagus revealed atrophy and fibrosis of the tunica muscularis. The entire oesophagus was affected, but the pathology was most prominent in the distal part. In this segment oval calcified structures (80 x 50 µm) were found, resembling disintegrated and calcified sarcocysts. No vasculitis nor neuronal damage (synaptophysin staining) could be demonstrated. Because of the inconclusive results of the C. botulinum analysis, the presence of histological changes in the oesophageal musculature and the demonstration of BTV infection, a tentative diagnosis of oesophageal paresis caused by BTV was formulated. In literature this phenomenon has incidentally been reported previously and therefore the clinical symptoms of regurgitation caused by oesophageal pathology may be included in the list of BTV-induced pathology in the future. Key words: regurgitation, oesophageal dilatation, blue tongue serotype 8, cattle 135 Seroepidemiology of Bovine Leukemia Virus (BLV) Infection in the North Eastern Provinces of Iran A. Haghparast , Gh. Mohammadi , Sh. Mousavi 1 Faculty of Veterinary Medicine, Ferdowsi University of Mashhad, Department of Pathobiology, Mashhad, Iran 2 Faculty of Veterinary Medicine, Ferdowsi University of Mashhad, Department of Clinical Sciences, Mashhad, Iran Bovine Leukemia Virus (BLV) is an exogenous c-type oncovirus in the retroviridae family that causes a chronic infection in cattle, which develops in three possible pathological forms: asymptomatic course, persistent lymphocytosis and lymphosarcoma. Once infected, cattle remain virus carriers for life and start to show a serological reaction within a few weeks after infection. This disease causes significant economic losses associated with the costs of control and eradication, loss in milk production and difficulties in exports. The Agar Gel Immuno Diffusion (AGID) and ELISA tests have been the serological choice assays for routine diagnosis of serum samples. However, due to the higher sensitivity of ELISA, in recent years this method has replaced the AGID in seroepidemiological studies for control and eradication programe. Prevalence studies on BLV infection has not been conducted in the eastern part of Iran. This study was designed to determine the prevalence of BLV in Khorasan Razavi and Khorasan Shomali provinces which are the main provinces located in the north east of Iran. During summer 2007, 429 blood samples were randomly taken from industrial dairy herds of these two provinces and subjected to ELISA test (indirect sandwich ELISA). 25.41% of these samples showed positive reaction. The positive results in Khorasane Razavi and Khorasan Shomali provinces were 29.83% and 1.5% , respectively. Moreover, 64.7% of the herds tested, had at least one positive cow. The overall true prevalence was 24.63%. Statistical analysis revealed a direct correlation between age and occurrence of BLV. Significant correlation was also found between herd size and BLV infection. In another words, the large herds had the most BLV positive cases. Moreover, number of parity was also shown to have a direct correlation with BLV seropositivity. Results of Odd ratios analysis which is an indication of possible risk factors, confirmed the above findings. Taken together, the results presented in this study along with similar studies in recent years in Iran confirm an alarming increase in prevalence of BLV infection among dairy herds. Therefore, control strategies based on test and implementation of corrective management should be taken in order to downgrade and possibly eliminate this economically hardship disease. Key words: BLV, seroepidemiology, Khorasan Razavi, Khorasan Shomali, Iran

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136 Individual and Herd Prevalence of Bovine Leukemia Virus Infection in Chilean Dairy Herds G. Monti, M. Grau Universidad Austral de Chile, Institute Preventive Veterinary Medicine, Valdivia, Chile Objectives of study: The aim of this study was to assess herd and within-herd prevalence of Bovine Leukemia Virus (BLV) infection in Chilean dairy herds. Materials and Methods: Herd and individual level prevalence was estimated with a cross-sectional random sample of 80 dairy herds in the main dairy area of the country (X° region) performed during 2007. Sample was stratified according to herd size in large, medium and small and it was considered the proportion of each class in the population of the herds of the area (2%, 13% and 85% respectively). All selected herds were commercial dairy herds that are registered at the regional offices of the Servicio Agrícola y Ganadero (SAG) from the Ministry of Agriculture of Chile. SAG also has records of the Milk Collection Centers (CALS) where smallholders deliver their milk. To avoid interferences with passive antibodies animal older than 6 months will be candidates to be tested. A commercial ELISA (IDEXX-BOMMELI) for serum samples was used for assess BLV status. The herd and individual-level true prevalence were estimated by using a Bayesian approach. Briefly, we modelled that n animals were sampled randomly from herds. Model estimate prevalence distribution based in the result of one test applied to individual animals, assuming that prevalence is a mixture of point mass at zero and an continuous Beta distribution on (0,1). Independent prior distribution were assumed for test sensitivity and specificity and modelled as beta distributions also. Results: The results showed a high degree of variation in herd and cow-level of BLV prevalence. The posterior median and 95% credibility interval for Herd-prevalence was 48% (20.9, 78.4). Also the predictive probability that a randomly selected herd in the region had prevalence less/equal than 5% is 70% and prevalence less/equal than 50% is 98%, and it is less that those of neighboring countries. Larger and specialized dairy herds showed larger prevalences. Conclusions: The cross-sectional study resulted in an estimation of the true prevalence estimates of BLV for one of the main dairy regions in Chile. The results showed the degree of variation in herd and cow-level BLV prevalence suggesting that a compulsory control program is feasible and small farmers have good opportunities to add value to their production by enrolling a free-herd certification schema. Key words: Bovine Leukemia Virus, prevalence, Bayesian approach 137 Q Fever (Coxiella Burnetii) Eradication in a Dairy Herd by using Vaccination with a Phase 1 Vaccine (Coxevac®, Ceva S.A.) Ph. Camuset , D. Remmy 1 Cabinet Vétérinaire, Yvetot, France 2 CEVA Santé Animale, Libourne, France Objective of the study: In a dairy herd followed for fecundity since 1985, Coxiella burnetii was found in April 2002 and March 2004. That analysis was initiated after a deterioration of the fertility results and early and late abortions of the dairy cows. In a second step, a protocol aiming to eradicate the infection by culling and vaccination was performed. Material and Methods: The herd is composed of about 30 dairy cows producing around 9000 kg of milk per lactation. Since Sept. 2004, the dairy cows and the heifers have been vaccinated 2 times/3 weeks of interval by sub-cutaneous route with the vaccine Coxevac® (CEVA S. A.), preferentially at W2 and W5 post partum (cows) and before first insemination (heifers). A booster injection between W2 and W5 postpartum was yearly planned. Post-partum, an individual diagnostic was initiated with research of the bacteria by real time PCR in the milk mixture of the 4 quarters at D0, D3, W1, W3 and W8 and in the vaginal mucus between D0 and D3. Results-discussion: During the first year of study, among the 46 cows studied (10 first lact.), 15 cows (2 first lact.) excreted at least one time the bacteria, 10 several times. The third year, only 3 cows excreted (no first lact.); among them, two that have excreted several times have been culled. During the study period, only one vaccinated cow, not excreting the previous lactation, became a new excreting (only in the vaginal mucus); on the other hand, four cows seem healthy again, they had only excreted in a previous lactation around calving always in vaginal mucus, sometimes in milk. They have never re-excreted later. For

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80 XXV. Jubilee World Buiatrics Congress 2008

fertility and fecundity, the performances (% of anoestrus, early and late metritis and abortions) are better since the use of the vaccination and the culling of the excreting cows. Those results must be put in perspective according to the sensibility of the Coxiella b. diagnosis because of the intermittence of shedding. For the performances of reproduction in high production dairy herds, the influence of herd management and feeding is able to put in perspective the only influence of the eradication of Q fever in that herd. Conclusion: Either for human health or for zootechnic and economic reasons, the use of a diagnostic method towards Coxiella burnetii shedding animals, their vaccination then their progressive elimination appear to be a feasible process by using the vaccine Coxevac® (CEVA S. A.). Key words: Q fever, bovine, vaccination, Coxiella burnetii 175 Serological Testing of Primiparous Cows and Young Cattle before Pregnancy is Useful to Detect Acute Q-Fever Infection within Herds J. Boettcher, A. Vossen, A. Gangl, N. Meier, A. Schneppe, G. Wittkowski Bavarian Animal Health Service, Poing, Germany As Q-Fever is a zoonosis knowledge about acute infection within cattle herds is of special interest. To estimate prevalence of Q-Fever in Bavaria 21858 sera from 765 herds were tested in ELISA (Idexx; plates coated with phase I and II-antigen) for antibodies. Seroprevalence (SerP%) was 14.5%±0,2% and 62.5%±1,4% of the herds were seropositive. Testing 799 cows in autumn 2003 and 2004 revealed a rate of seroconversion of 4.6% (CI95% 2.9-5.9%) p.a. To detect age-related effects 2819 sera were analysed. Decreasing SerP% until 6 month of age (mona) is explained by maternally derived antibodies. A low level SerP% (<2%) was observed between 6-24 mona, while a sharp increase in SerP% was characteristic for animals between 24 and 36 mona. Two antigens (phase I and II) of Coxiellae are of special interest regarding discrimination of acute and + - + chronic infections. In humans a phase I /phase II (phI /II ) + + serological pattern is indicative for acute and phI /II -pattern for chronic infection. Therefore sera from a dairy experiencing an acute outbreak in summer 2005 were analysed for antibodies directed against phase I- and II-antigens, respectively. Most remarkably - + between 6 and 24 mona exclusively animals of the phI /II -pattern + + were found (SerP%=10%). After 24 mona phI /II -SerP% increased - + slowly to 35% in animals older than 3.5 years. PhI /II -SerP% reached its maximum (38%) about one year earlier, it preceded the + + phI /II -peak. Analysis of sera from this herd between May 2004 and + + - + May 2006 revealed that SerP% of phI /II - and phI /II -antibodies + + were inversely correlated. Each time when phI /II - SerP% dropped - + below 30%, a sharp increase of phI /II -SerP% was seen. These data are the basis for the hypothesis that primiparous cows are a hotspot of infection; consequently the risk of human infection is increased. PhI + /II -pattern in cattle of 6-24 month of age might result from infection at birth. A dormant stage of Coxiellae might be responsible for a lack of phI-reactivity. During first pregnancy Coxiellae got activated and are invading the uterus followed by shedding of Coxiellae at + + parturition. Waves of inversely correlated SerP% of phI /II - and phI + + + /II -antibodies are best explained by immunity due to phI /II pattern. Financially supported by the Free State Bavaria and the Bavarian Joint Founding Scheme for the Control and Eradication of contagious Livestock Diseases. Key words: Q-Fever, serology, phase I/II-antibodies 176 Seroepidemiological Study of Crimean Congo Hemorrhagic Fever in Khorasan Provinces, Iran Dr. Mostafavi , Dr. Bokaei , Dr. Haghdoost , Dr. Keivanfar , 4 5 6 7 Dr. Gooya , Dr. Meshkat , Mr. Davari , Dr. Chinikar 1 Faculty of Veterinary Medicine, University of Tehran, Epidemiology, Tehran, Iran 2 Kerman University of Medical Sciences (KUMS), Epidemiology, Kerman, Iran 3 Faculty of Veterinary Medicine, University of Tehran, Microbiology and Immunology, Tehran, Iran 4 Center for Disease Control, Ministry of Health, Tehran, Iran 5 Veterinary Organization, Ministry of Jihad-e-Keshavarzi, Center of Animal Disease Control, Tehran, Iran

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Center for Disease Control, Ministry of Health, South Khorasan, Qaen, Iran 7 Pasteur Institute, National Reference Laboratory of Arboviruses and Hemorrhagic Fevers, Tehran, Iran Crimean-Congo Haemorrhagic Fever (CCHF) is an arboviral zoonotic disease that is asymptomatic in infected animals, but a serious threat to humans. Humans become infected with Crimean-Congo Haemorrhagic Fever Virus (CCHFV) either through bites of infected ticks, or by direct contact with virus contaminated tissues or blood of animals. CCHF now occurs sporadically throughout much of Africa, Asia, Europe, and recent outbreaks of CCHF have occurred in Afghanistan and Pakistan. Since 1999, cases of CCHF have been reported from different parts of Iran. Nonetheless, most of human reports came from the Southeast of Iran; therefore we analyzed samples from the Northeast provinces to check the geographical extension of CCHF. From October 2001 to the end of 2007, 10 (15.9%) of 63 probable human patients confirmed to be infected with CCHFV using specific enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) tests in Khorasan provinces, Northeast of Iran, close to Afghanistan and Pakistan. No specific antibodies against other viral fevers such as Yellow fever, Rift Valley Fever, Dengue 2, and Chikungunya were detected in the sera of these probable cases. During the years 2003 to 2005, of 448 animal sera that collected form different townships of this region, IgG antibodies were noted in 77.5% (95% CI: 72.5-82%) of 298 sheep samples and 46% (95% CI: 38.1-54.0%) of 150 goat samples. Sexes were infected equally. There was an age-related increase in seroprevalence of sheep and goats. Regardless whether CCHF is newly enzootic or has long established enzooticity, the potential exists for sporadic or clustered outbreaks of CCHF in humans, so persons in close contact with animals and also health care workers should be alarmed. Key words: Crimean-Congo Haemorrhagic Fever (CCHF), RT-PCR, ELISA, Khorasan provinces, Iran 177 An Outbreak of Epizootic Hemorrhagic Disease Virus among Beef & Dairy Cattle in Israel M. Kedmi , N. Galon , H. Yadin , V. Bombarov , C. Batten , 4 4 2 P. Mertens , S. Anthony , E. Klement 1 Hachaklait, Clinic for ruminants, Kayseri, Israel 2 Koret School of Veterinary Medicine,Hebrew University, Department for Infectious Diseases- Epidemiology, Rehovot, Israel 3 Kiimron Veterinary Institute, Virology, Beit-Dagan, Israel 4 Institute of Animal Health, Reference Laboratory, Pirbright, United Kingdom Epizootic hemorrhagic disease virus (EHDV) is an arthropod-borne virus, which is a member of the Orbivirus genus of the family Reoviridae. At least eight serotypes of this virus have been reported worldwide most of which affect wild cervids during the summer of 2006, an outbreak of epizootic hemorrhagic disease in cattle caused by EHDV serotype-7 (EHDV-7) occurred in Israel. The disease was reported from the end of August 2006 until mid November. The disease was characterized by a variety of clinical signs, including mucosal bleeding, ulcers, salivation and swollen, injured teats. Recumbency and mortality were rare (<1.5%). The majority of affected cows did not show any clinical symptoms except significant decrease in milk production. Overall, 83 dairy cattle herds and 22 beef cattle herds were affected. No morbidity was reported in wild or small domestic ruminants. In this report we describe the outbreak spread in terms of time and space. We aim to characterize the economical losses caused by EHDV and to define the involvement of small domestic and wild ruminants in this outbreak.During the outbreak serum samples were collected from more then 2000 ruminants (dairy and beef cows, sheep and goats) from affected and unaffected herds. Sera were serologically screened for exposure to EHDV and BTV by specific competitive enzyme- linked immunosorbent assays (ELISAs). Milk production and morbidity were monitored using specific herd managment softwares. Serological evidence for exposure was found only in cows and buffalo, while no seropositive domestic and wild small ruminants were detected. The rate of exposure was high (80%-100%) in the Jordan valley and decreased progressively along the trail of the outbreak towards the coastal plane (0%-20%). Disease spread apparently followed the local winds, which blew from East to North and West. A loss of 100 liter per milking cow was recorded in affected herds. We estimated that over 800 cows died or were culled as a direct result of the

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disease. Thus losses were estimated to exceed 2M $US. This is the first report of an outbreak of EHDV-7 is recorded in cattle in Israel. Key words: EHDV, BTV, spatial, wild ruminanta, wind 178 Evaluation of a New Respiratory Syncytial Virus Vaccine Candidate G. Meyer , M. Deplanche , C. Dubuquoy , M. Soulestin , 3 3 3 4 3 G. Durand , N. Castagne , J. Bernard , P. Bernardet , S. Riffault , 3 JF. Eleouet 1 National Veterinary School of Toulouse, Pathology of Ruminants, Toulouse, France 2 National Institure of Agronomical Research, Pathology of Ruminants, Toulouse, France 3 National Institure of Agronomical Research, Animal Health Department, Jouy en Josas, France 4 National Institure of Agronomical Research, Animal Health Department, Tours, France Objective: To evaluate the efficacy of a new vaccine candidate against Respiratory Syncytial Virus (RSV) based on N protein nanoparticles. Method: The vaccine was constituted by sub-nucleocapsid ring structures (N SRS), each ring being composed by 10 or 11 RSV nucleoproteins surrounding a bacterial RNA of 70 bases (Patent WO 2006/117456). Twenty four one-month old calves, free of bovine RSV antibodies, were divided in three groups of 8 calves. The first group was vaccinated twice (4 weeks interval) with 2 mg of N SRS protein with Montanide-ISA70 VG adjuvant by the intramuscular route. The second group received at four weeks interval the same vaccine formulation and 10 mg of N SRS protein with Montanide-IMS 4112 VG adjuvant by the intranasal route. The last group served as nonvaccinated control calves. One month later, all calves were challenged 5 by intranasal and intratracheal routes with 10 UFP of the BRSV 3761 strain. Clinical examination was performed for 21 days post challenge. Virus infection was studied by real time PCR on nasal swabs from 3 days before to 21 days post challenge. Bronchoalveolar lavages and lung biopsies were obtained from two calves per group euthanized at 6 days post challenge and were used for virus quantification. RSV antibody response (ELISA) was studied and lymphoproliferation studies were performed on maxillary, mediastinal and tracheobronchial lymphatic nodes. Results: All vaccinated calves developed anti-N antibodies in blood and nasal secretions following vaccination. A memory cellular response was also detected in the sub-maxillary lymph node of vaccinated calves. Clinical examination demonstrated a moderate pathology for each group with significant differences on days 5 and 6 post challenge between the two vaccinated groups and the control one. Viral quantification showed lower viral loads in nasal secretions for the two vaccinated groups, with significant differences at days 3, 4 and 11 post challenge. No significant differences were observed between the two vaccinated groups for clinical signs and virus load parameters. Conclusion: Vaccination with N SRS partially protects calves against challenge with a wild bovine RSV strain. No differences were observed between the two vaccinated groups, suggesting that intranasal vaccination did not prime mucosal immunity in our experimental conditions. Studies are underway to improve the mucosal route of vaccination as well as immunogenicity of the candidate RSV vaccine. Key words: Respiratory Syncytial Virus, vaccination 179 Efficacy of an Inactivated BTV-8 Vaccine against a Virulent BTV-8 Challange in Sheep C. Hamers , M. Blanchet , A. Chossat , L. Besancon , C. Cariou , 2 1 E. Hanotel , S. Goutebroze 1 MERIAL SAS, Clinical Development, Saint Vulbas, France 1 MERIAL SAS, Bio-Analytical Development, Lyon, France Introduction: Bluetongue virus (BTV) causes an infectious, noncontagious, Office International Epizooties (OIE) and BTV cycle can be interrupted by the systematic vaccination of susceptible species, particularly sheep and cattle. Here, we demonstrated in a vaccination / challenge study that single administration of an inactivated vaccine containing purified BTV serotype 8 (BTV-8) was able to prevent clinical signs and viraemia in sheep. Material & Methods: Eleven weaned lambs were randomly allocated to one group of 5 vaccinates and one group of 6 control sheep. Vaccinates were subcutaneously treated once on day 0, with 1 mL of

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BTVPUR* AlSap8 (MERIAL). The other group was left unvaccinated and served as control. Thirty one days after vaccination, all sheep were challenged with a virulent BTV-8. All sheep were then monitored for rectal temperature and clinical signs from 5 to 14 days after challenge. Furthermore, viraemia of all sheep was monitored by quantitative RTPCR on whole blood samples on days 5, 7, 9, 12 and 14 after challenge. Results: Hyperthermia : Vaccinates were significantly protected from hyperthermia. Average maximal hyperthermia was 41.2°C in the controls and 39.9°C in the vaccinates. Clinical scores: Vaccinates were significantly protected from clinical signs. Average global clinical score was 28.8 in the control group and 1.2 in the vaccinated group. Viraemia (qRT-PCR): All controls were found positive (moderate to high copy numbers) at all time points. None of the vaccinated animals was ever detected positive. Conclusion: In the present study, single vaccination with the product tested provided full and significant clinical and virological protections against a BTV-8 challenge. These results show that BTVPUR* AlSap 8 may be used for BTV-8 disease prevention (clinical protection) and for epidemiological control of BTV-8 (virological protection). * BTVPUR AlSap is a registered trademark of the MERIAL group in the European Community. Key words: Bluetongue, BTV-8, inactivated vaccine, efficacy, sheep 180 Efficacy of an Inactivated BTV-8 Vaccine Against a Virulent BTV-8 Challange in Cattle C. Hamers , M. Blanchet , F. Teppe , L. Besancon , A. Meyet , 1 S. Goutebroze 1 MERIAL SAS, Clinical Development, Saint Vulbas, France 2 MERIAL SAS, Bio-Analytical Development, Lyon, France Introduction: Bluetongue virus (BTV) causes an infectious, noncontagious, Office International Epizooties (OIE) and BTV cycle can be interrupted by the systematic vaccination of susceptible species, particularly sheep and cattle. Here, we demonstrated in a vaccination / challenge study that administration of an inactivated vaccine containing purified BTV-8 was able to prevent cattle from clinical signs and viraemia. Material & Methods: Ten cattle were allocated to 2 groups of 5 animals. One group was subcutaneously vaccinated twice (day 0 and 28) with 1 mL of BTVPUR* AlSap8 (MERIAL). The other group was left unvaccinated and served as control. On day 51, all animals were challenged with a virulent BTV-8. All cattle were then monitored for rectal temperature and clinical signs from 5 to 28 days after challenge. Furthermore, viraemia of all animals was monitored by quantitative RT-PCR on whole blood samples, three times a week during the monitoring period. Results: Hyperthermia: All controls presented an elevated rectal temperature, whereas rectal temperatures remained normal for all vaccinates. Clinical scores: Vaccinates were significantly protected from clinical signs. Average global clinical score was 59.0 in the control group and 10.8 in the vaccinated group.Viraemia (qRT-PCR): All controls were found positive (moderate to high copy numbers) at all time points but at day 56 none of the vaccinated animals was ever detected positive. Conclusions: In the present study, vaccination with BTVPUR* AlSap 8 provided full and significant clinical and virological protections against a BTV-8 challenge. These results show that BTVPUR* AlSap 8 may be used for BTV-8 disease prevention (clinical protection) and for epidemiological control of BTV-8 (virological protection). * BTVPUR AlSap is a registered trademark of the MERIAL group in the European Community Key words: Bluetongue, BTV-8, inactivated vaccine, efficacy, cattle 181 Rapid Detection of Rift Valley Fever Virus Based on Viral Small (S) RNA Genome Segment I. Aradaib , M. Al-dubaib 1 The National Ribat University, Medical Lab. Sciences, Khartoum, Saudi Arabia 2 Qassim University, Veterinary Medicine, Buridah, Saudi Arabia A reverse transcriptase Polymerase Chain Reaction (RT-PCR)-based assay was developed and evaluated for detection of Rift Valley Fever Virus (RVFV) ribonucleic acid (RNA). A pairs of oligoribonucleotide primers selected from the small (S) RNA genome segment of RVFV, were designed and used for PCR amplification. The primers resulted in

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82 XXV. Jubilee World Buiatrics Congress 2008

amplification of a primary 490 base pair PCR product. The PCR products were amplified from RNAs extracted from RVFV field isolates and vaccine strains, propagated in Vero cell cultures. Amplification products were not detected when the RT-PCR-based assay was applied on RNA from other related hemorrhagic fever viruses including Crimean Congo Hemorrhagic fever; dengue virus; epizootic hemorrhagic disease virus and total nucleic acid extracts from uninfected Vero cells. The described RT-PCR-based assay provides a rapid, sensitive and specific assay for detection of RVFV in cell culture and should be recommended for inclusion during outbreaks of the disease among humans and susceptible livestock. Key words: viral hemorrhagic fevers, RVFV, sRNA, RT-PCR 182 Prevention of Coxiella burnetii Shedding in Intedted Dairy Herds Using a Monovalent Vaccine Containing Phase-1 Coxiella burnetii R. Guatteo , F. Beaudeau , A. Joly , D. Remmy , H. Seegers 1 National Veterinary School of Nantes, Unit of Animal Health Management, Nantes, France 2 Union Bretonne des Groupements de Défense Sanitaire, Vannes, France 3 CEVA Santé Animale, Libourne, France Objective of study: The main aim of this study was to assess the efficacy of a monovalent vaccine containing phase I Coxiella burnetii to prevent Coxiella shedding in susceptible (ie considered as non infected) dairy cows within infected herds in comparison to a placebo. Materials and Methods: To reach this goal, 336 dairy cows and heifers, from 6 infected herds with repeated abortions due to Coxiella burnetii, were followed up over a one-year period. Before treatment (ie vaccination or placebo), the status of the cows towards Coxiella burnetii infection was determined on the basis on PCR results on milk, vaginal mucus and faeces and serological analysis performed 2 weeks apart. A cow was considered as susceptible when all PCR and serological results were negative, and was considered as infected otherwise. Then, the allocation of treatments was performed randomly within pregnant and non pregnant cows. After treatment (D0), the animals were subject to systematic samplings (milk, vaginal mucus and faeces) on D90, D180, D270 and D360 to detect a putative shedding using PCR. Moreover, under the assumption that calving period is at maximum risk of shedding, the same samples were taken within 15 days after calving. An animal was considered as a shedder at a given time t, if at t, it was found positive-PCR tested at least once among the taken samples. The effect of treatment on the risk for an animal of becoming shedder was assessed using survival analysis techniques (Cox regression model). Results: Almost all heifers were detected as susceptible before treatment. When vaccinated while not pregnant, an animal had 0.2 (P<0.05) times higher (i.e. 5 times lower) risk of becoming a shedder than an animal receiving placebo. An animal which was vaccinated while pregnant had a similar risk of becoming shedder than an animal receiving placebo (HR = 0.9; P<0.05).No significant difference in the risk of becoming a shedder was found between primiparous and multiparous animals. There was no significant farm effect in this multicentric trial. Conclusion: In this original trial, carried out under field conditions and in infected herds, a very strong protective effect towards Coxiella shedding in susceptible animals was found. These results highlight the interest of implementing vaccination, if possible, in non infected herds, or at least in heifers located in infected herds, under the assumption that they are still susceptible. Key words: Coxiella burnetii, dairy cow, vaccination, shedding, survival analysis 183 Development of Multiplex-PCR Techniques for Detection of Abortive Bacterial Infections of Sheep B. Sareyyupoglu , K. Diker , S. Güngördü , Z. Cantekin , R. Jahed 1 Faculty of Veterinary Medicine, Ankara University, Department of Microbiology, Ankara, Turkey 2 Faculty of Veterinary Medicine, Mustafa Kemal University, Department of Microbiology, Hatay, Turkey One of the most important problems of sheep breeding in Turkey is bacterial abortion. Amongst the bacterial infectious diseases causing ovine abortions in Turkey are brucellosis, campylobacteriosis,

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chlamydiosis, listeriosis and salmonellosis. More than 9 genera are known to be regularly encountered in ovine abortion cases. Some of these are zoonotic and a threat for human health. Despite conventional routine isolation and identification methods used for laboratory diagnosis of these infections, problems are encountered such as lack of equipment, requirement of strict isolation environment, facility and biosecurity precautions in diagnostic laboratories and these methods are laborious and time consuming. Recently, polymerase chain reaction (PCR) and other PCR based molecular detection techniques have been used in the diagnosis of infections. Since different bacteria cause ovine abortions, diverse PCR applications should be performed for the molecular detection of each of these. Problems with conventional identification methods like time, labor and cost can be solved by multiplex-PCR applications. In this project, original PCR and multiplex-PCR techniques were developed for the molecular detection of abortive bacterial infections of sheep. Thirty nine pair of oligonucleotide primers was originally designed for use in PCR and multiplex-PCR techniques. Following the selection of appropriate primer pairs, four different multiplex-PCR techniques (2 multiplex for the detection of Brucella spp., Campylobacter spp. and C. abortus, one dublex-PCR for the detection of Brucella spp. and Campylobacter spp., and 1 multiplex for the detection of Listeria spp., Leptospira spp. and C. burnetii) were developed. Another multiplex-PCR technique for Chlamydophila spp. and Salmonella spp. detection were also established with the known primers. These techniques are believed to be unique in the world. This research has been sponsored by TUBITAK under project TOVAG104O237. Key words: bacterial abortion, molecular detection, multiplex-PCR, primer design, sheep

POSTER ABSTRACTS

634 Diagnostic Studies on Caprine Brucellosis at Assiut Governorate, Egypt T. El-Allawy , A. Ahmed , M. Hussien , M. Sayed , M. Abdel-Rady 1 Faculty of Veterinary Medicine, Assiut University, Animal Medicine, Assiut, Egypt 2 National Research Center, Department of Food Hygiene, Assiut, Egypt 3 National Research Center, Microbiology, Assiut, Egypt A total of 2024 blood serum samples of goats were collected from different localities of Assiut Governorate and screened by RBPT and BAPAT. Positive samples to RBPT and/or BAPAT were further tested by TATa, TATm, MET, Riv. T and CFT. 19 samples out of 2024 samples were diagnosed as true positive with an overall incidence rate of 0.94%. The incidence rates of the different localities were 1.8%, 1.55%, 1.85%, 1.13% and 1.53% in Assiut, Sahel-Saliem, Sedfa, El-Qusseia and Dairout, respectively. The highest incidence was recorded in Sedfa (1.85%) while the lowest in El-Qusseia (1.13%). No positive results were detected in El-Fateh, Abnoub, ElBadary, Abo-Teeg, El-Ghanaiem and Manfalut.The efficiency percentages were 100%, 100%, 96%, 96%, 100%, 99.6% and 100% for RBPT, BAPAT, TATa, TATm, MET, Riv.T and CFT, respectively. The agreement rates of different serological tests with the CFT were 100%, 100%, 96%, 96%, 100% and 99.6% for RBPT, BAPAT, TATa, TATm, MET and Riv.T, respectively. A total of 293 milk samples were also collected from lactating goats in different localities and subjected to the Milk Ring Test (MRT). 2 samples only were positive with a percentage of 0.68%. The results of serum tests of these 2 samples were as follows, one was positive in all tests and the other one was positive in RBPT, BAPAT, MET, Riv.T and CFT and negative in TATa and TATm. Goats must be taken into our consideration during the work of the eradication programmes as it can transmit the infection to other free areas (mobile flocks) and / or to other animal species such as cattle and buffaloes (common pasture). Because it is easy to perform and interpret, its high sensitivity and its good correlation with the serological tests, the Milk Ring Test (MRT) was found to be a good and reliable field test so it can be used as a rapid screening test to diagnose the disease in lactating goat. Also the MRT is an easy, rapid and reliable test if done carefully. Key words: diagnosis, caprine, seological tests, brucellosis

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635 Prevalence of the Pulmonary Lesions of the Bovines of the Area of Tiaret B. Mabrouk, T. Madjid, M. Toufik, M. Bakir Batna University, Department of Veterinary Sciences, Batna Algeria, Algeria The seizures partial or total of the carcasses or the bodies by the veterinary services constitute a true fear for the butchers. A study undertaken in the slaughter-houses of the wilaya of Tiaret (Western Algeria) during the last two years and a great incidence of respiratory lesions at the bovines was found. This study shows the prevalence and the frequency of a series of lesions raised on the lungs of 870 bovine autopsiés during medical inspections and 744 pathologies were diagnosed. The seasonal distribution of the lesions made it possible to put forward the effect of the season on the appearance of the lesions. The cyst hydatic constitutes pathology the most met (n=330 either 42.64%) followed 111 cases of emphysema (or 14.35%) and 61 cases of congestion (or 7.89%). The histopathologic study of the fragments of injured lungs taken at the slaughter-house highlighted tissue deteriorations. According to the localization of the lesions, one noted that the right lung is touched the most for 92.60% of the cases that the left lung as well as the craniaux lobes for 87.05% comprativement with the caudal lobes of the lungs. Key words: pulmonary lesions, histopathology, bovine 636 A Retrospective Study of Bovine Tuberculosis Eradication Project and Human Tuberculosis M. Sakha , N. Ezatpanah , A. Rohitoorbati 1 Islamic Azad University, Science & Research Branch, Internal Medicine, Tehran, Iran 2 National Research Center, Internal Medicine, Kerman, Iran 3 Faculty of Veterinary, Internal Medicine, Kerman, Iran Introduction: Tuberculosis (TB) is a chronic and zoonotic disease, had known for many years. The cause of the disease is acid-fast bacillus of mycobacteriacea, mycobacterium tuberculosis. Weakness, emaciation and chronic respiratory symptoms are common signs of the disease. The main group of human TB is pulmonary TB and the other is extra pulmonary TB. It seems that many cases of extra pulmonary TB are related to animal tuberculosis. This is a retrospective study made during the last ten years (1991- 2000) on the incidence of human and bovine TB and evaluated the bovine TB control project to human TB incidence in city of Kerman, IRAN. Bovine TB control project is a national project that has performed for many years in IRAN. Materials and Methods: All data were taken from the veterinary office in Kerman for bovine TB control project and the only centre of human tuberculosis control for human cases. Data were analyzed and variables of age, gender, root of infection and social situation were considered in human section. The bovine positive cases have been omitted and sent to the slaughter house before. Results and Conclusion: This study showed that the most patients were over 60 years old and then 21 to 30 years old. Women showed more incidence of the disease. The prevalence of the extra pulmonary TB in the first year of the study was11.9 in 100,000 and two years later was relatively higher than. In remaining years the prevalence showed a decreasing slope and reached to half of early year. Among the 219375 cows tested for TB in ten years,`65 cows were reactor. The most incidence was found in the first and second years, that is, 37 and 18 cases, respectively. In later years, there was obviously a decreasing slope with only one or two reactors in 1993 to 1998 and no cases in 1999 and 2000.There is positive relation between decrease in human TB (extra pulmonary TB ) and the control of bovine TB in Kerman, so that with the obvious decreasing of bovine reactors in last year of this study, human cases also had a decreasing slope, although the human extra pulmonary TB is nearly one third of total human cases and it needs supplemental projects to control of human tuberculosis. Key words: tuberculosis, bovine, human, retrospective 637 Bovine Theileriosis: Evaluation of 372 Cases during Spring and Summer in South of Iran M. Sakha , M. Radfar , M. Janbaz 1 Islamic Azad University, Science & Research Branch, Internal Medicine, Tehran, Iran 2 Faculty of Veterinary Medicine, Parasitology, Kerman, Iran 3 Faculty of Veterinary Medicine, Internal Medicine, Kerman, Iran

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Bovine theileriosis caused by Theileria annulata is a dangerous and important disease in tropical and subtropical areas. There are large economical losses in these areas every year including Iran. The vector of the disease in Iran, the tick of Hyalomma is found in the most parts of the country. The disease is more seen in spring and summer. In this study, the prevalence of the disease in terms of month of the year, age, sex and animal breed were studied. Results showed that the maximum and minimum prevalence of the disease was in May and June, respectively, during the studied months. In relation to age, 76.2 percent of the infected cattle were under a month old. In relation to the breed, infection of different breeds of Holstein, crossbred and native cattle were 1.6, 94.4 and 4.0 percent, respectively. In relation to sex, 55.4% of males and 44.6% of females were affected. From the aspect of clinical signs, nearly all cattle showed anorexia and enlargement of lymph nods. 58% of blood smears showed less than 10% intraerythrocytic form of theileria in microscopic high power field, 38% showed more than 10 percent and the rest was uncertain to diagnose. It is concluded that in spite of all the means of prevention and vaccination, the disease has been known as the most important disease in the region especially in the warm months of the year parallel to the activity of hyalomma. The dominant affected breed was crossbred, because the most of the referred animals was crossbreds. Calves were the most affected in this study because most of our cases were calf and cattle were more treated at the farms. The infected RBC were not correlated to the severity of the disease. 638 New Approaches in the Prophylaxis of Prion Infections V. Vlizlo, P. Verbitskii, C. Mayor, V. Stadnyk Institute of Animal Biology UAAS, Scientific Center for Prion Infections Study, Lviv, Ukraine Objectives of study: In nowadays very much the attention is spared to the problem of the treatment of prion infections. Most efforts are concentrated on the search of preparations which would block SC occurance or reduce the level of already present PrP . At the same time, very little attention is spared to the problem of prophylaxis of the prion infections. Although work in this direction is rather active, taking into considiration a creation of transgenic cattle in which the gene of Prnp is knocked-out. But still it is unknown, how exactly will react an C organism on absence of such important protein as PrP . That is why we consider, that more perspective direction is a search of preparations C SC which would reduce the level of PrP , in the same time depriving PrP of substrate for replication. These preparations can be applied in risky herds, with suspicious or sick animals are founded, for the reduction of risk of prion infections among the healthy individuals. That is why the purpose of our work was a search of preparations which would reduce C the level of PrP in the organs of the prion-replicating system of laboratory animals. Materials and Methods: Work was done on laboratory rats treated with heparine and Petosan Polisulfate. After decapitation in the proper C organs were probed: general maintenance of PrP , type of expression of its izoforms, maintenance of zinc and copper, activity of C superoxidedismutase, evaluation of DNA damage, ELISA test of PrP interaction with pentosan polisulfate (PPS), determination of UVC spectrums of PrP at interaction with PPS. Results: It is set that heparine and PPS is able to reduce the level of C PrP in the organs of the prion-replicating system; unlike heparine, it is shown that PPS operates in far less concentrations and does not influence on the state of the antioxidant system. It is also shown that C action of PPS on PrP proportionally depends on the concentration of preparation. In the case of heparine such dependence was not discovered. It is set that both preparations do not influence on genomic DNA, which is proved by the absence of doblestrain DNA breaks. C Conclusions: It is set an inhibiting action of PPS and heparine on PrP in vivo. Key words: prion infections, cellular prion, pentosan polysulphate 639 Brucellosis Surveillance in Jordan, 2001-2006 B. Barqawi , Z. Mashhadani , B. Hijjawi 1 Central of Disease Control & Prevention-CDC, Field Epidemiology Training Program-FETP, Amman - Jordan, Jordan 2 MOH, DCD, Amman - Jordan, Jordan Background: Brucellosis is a zoonotic bacterial disease that often infects livestock animals (sheep, goats, and cattle). Animals often shed

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organisms during birth. Humans become infected by coming into contact with animals or animal products contaminated with the bacteria. The objective of this study was to examine brucellosis surveillance and livestock immunization data in order to make recommendations that can improve brucellosis prevention and control programs in Jordan. Methods: All health directorates in Jordan electronically report the number of cases of confirmed and probable human brucellosis to the Ministry of Health weekly. Confirmed human cases of brucellosis are diagnosed by standard Coombs tube agglutination test. Clinically compatible cases with supportive serology (Rose Bengal test) and epidemiologically linked to confirmed cases are defined as probable cases. The Veterinary Directorate, Ministry of Agriculture collects data on livestock and immunization. The study period was 2001 to 2006. Results: 1,137 cases of human brucellosis were reported in Jordan during 2001 to 2006. The mean yearly number was 190. Mean yearly human brucellosis incidence was highest in Mafraq (30/100,000 population) and Karak (12) Governorates. Mafraq (4:1) and Karak (3:1) have the highest mean animal-to-human density of sheep and goats in Jordan. Human cases vary by month of occurrence; most cases (651; 57%) were from May to August and few cases (214; 19%) were from October to February. Immunization of sheep and goats in Jordan increased from 15% in 2002 to 46% in 2006. Conclusions: Human cases of brucellosis cluster geographically in areas with high density of sheep and goats. Occurrence of human cases parallels the spring-summer birthing pattern in animals. Human brucellosis is highly underreported. WHO burden of disease studies estimate there may be 3,000 human cases a year in Jordan. There does not appear to be any recent change in the incidence of human brucellosis or animal disease. New surveillance methods to identify more cases will be explored, such as following up recipients of antibiotic prescriptions typically used in brucellosis treatment. The Ministry of Agriculture and Ministry of Health increased immunizations to almost 2 million animals in 2006, but effective prevention will require sustained efforts to maintain high immunization levels, particularly in Mafraq and Karak. 640 Comparison of Biochemical Profiles of Hydatid Cyst Fluids of Echinococcus granulosus from Different Hosts in Iran A. Eslami , P. Khazraii , M. Heidarpour , Hr. Mohamadi 1 Faculty of Veterinary Medicine, University of Tehran, Department of Parasitology, Tehran, Iran 2 Faculty of Veterinary Medicine, University of Tehran, Department of Clinical Sciences, Tehran, Iran Hydatidosis (echinococcosis) is caused by the cystic larval. Stage of Echinococcus granulosus, is recognized as being one of the world's major zoonoses. Hydatidosis is endemic in Iran where many domestic animals including sheep, goat, cattle and camel act as intermediate hosts, while dog coyote and wolf act as final host of E. granulosus. There are nine host adapted strains of E. granulosus. The strain characterization is particularly important in regions where more than one species of live stock intermediate host exist and where there is the possibility of different cycles of transmission and source of infection for humans. Biochemical studies are useful in differentiating strain variations of E. granulosus in different countries. 15 sample of hydatid fluids were collected from the lung and liver of each host (i.e. sheep, cattle and buffalo).The concentrations of triglycerids, cholesterol, glucose, urea, creatinine, uric acid, total protein, magnesium, calcium, + phosphorous, LDH, ALP, AST, CPK, GGT, Na and K were determined. The biochemical profiles of lung and liver in these hosts were compared with each other. Hydatid fluids from the lung cysts of sheep contained significantly more glucose, uric acid, magnesium and LDH than those of cattle; but there were no significant differences on the concentration of other factors in hydatid fluids from lung cysts of these two species. There were other similarities and significant differences in the biochemical composition in hydatid cyst fluids of different hosts which can assist in the characterization of the host origin of infectivity in human being. 641 Vaccinia Virus in Humans and Cattle in Southwest Region of Sao Paulo State - Brazil J. Megid , C. Appolinario , R. Carreira , H. Langoni , M. Pituco , 2 L. Okuda 1 UNESP- School of Veterinary Medicine and Animal Sciences, Veterinary Hygiene and Public Health, Botucatu, Brazil

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2

Instituto Biológico, Laboratorio de Viroses de Bovideos, Sao Paulo, Brazil

Although human vaccine was erradicated in the world and the vaccination programs stopped, outbreaks of Vaccinia Virus has been reported in several states and regions of Brazil from which different types of Vaccinia Virus (VV) were isolated and characterized. Recently, Trindade et al (2007) studied the origin and diversitiy of Brazilian Vaccina Virus isolated evaluating the possibility that those could represent an escaped vaccinal strain. The authors reinforce the persistence of vaccinia virus in Brazil and other parts of the world despite the smallpox eradication campaign and the concept that the vaccinal strain could no more exist in nature. Objective: In this paper we report another cases of Vaccinia Virus infection in cattle and humans in the southwest region of Sao Paulo state in Brazil. The disease was observed in several small dairy farms with manual milking. Lesions varying from vesicules to ulcers were observed in the teats of the cattle and mouth of one calf. Milkers presented lesions in the hands and one child, 11 years old, presented lesions in mouth and nose. The age of the milkers ranged from 47-63 years old. Material and Methods: Crusts and swabs were collected from the animals that presented lesions in initial and healing stages.Crusts and swabs were maintained in Eagle Minimal Essential Medium (MEM) with penicilyn (500U/mL) and mycostatin (10 µg/mL) and maintained in refrigeration and sera were freezed. The material was processed for virus isolation and PCR assays. Results: A Vaccinia Virus was isolated in cell culture from all crusts and confirmed by PCR assay. Conclusion: Outbreaks of Vaccinia Virus in cattle in Brazil has been frequently reported. Lesions are observed in animals as well as in humans older than 40 years previously vaccinated against variola. This paper reports the characterization of new focus of Vaccinia Virus in Brazil and reinforce the susceptibility of humans previously vaccinated against variola to the circulating strain of vaccinia virus. Key words: Vaccinia Virus, cattle, human, outbreaks, public health 642 Mycoplasma bovis Sanitation Program in a Dairy Herd using Baytril 100/Max and Biosecurity Measures D. Gevaert , P. Passchyn 1 Bayer SA-NV, Animal Health Division, Brussels, Belgium 2 DAP De Toren, Veterinary Medicine, Torhout, Belgium Objectives of the study: A sanitation program in a dairy herd with bovine respiratory disease (BRD) initiated by Mycoplasma bovis was tested. Materials and Methods: A dairy herd of 117 Holstein Friesian cattle suffered since 2001 from BRD: relapses and growth retardation (calves) + chronic coughing (adults). Tilmicosin and tulathromycin treated calves relapsed after 14 days. Calves are housed individually up to 2 weeks, followed by group housing in 3 age groups (1, 2, 3 months) in the same stable. The calves are vaccinated against IBR, Pasteurella haemolytica, Parainfluenza and BRSV. Sanitation program: Step 1: herd visit + analysis of the spread of M. bovis using ELISA serology. Step 2: enrofloxacin (Baytril 100/Max) treatment + biosecurity measures. Step 3: follow-up visits. Results: Step 1: the M. bovis antibody prevalence increased with age: 25% in calves <3 months; 50% in calves 4-5 months; 79% in young cattle 6-12 months; 69% in cattle 1-2 years; 83% in the adult cows. 77% of the offspring of positive cows were also positive as compared to 42% positive offspring of negative cows. Step 2: Newborn calves were treated routinely on the 2nd day of life with a single administration of 7.5 mg enrofloxacin per kg BW to inhibit vertical M. bovis transmission. Calves developing BRD later on received a single administration of 7.5 mg enrofloxacin per kg BW against horizontal spread. Biosecurity measures: a separate calving stable, formaldehyde disinfection (P3-Incidin 03), freezing of colostrum from seronegative cows, feeding of female calves with artificial milk instead of cow milk. Step 3: The farmer reported in April 2007 conspicuously less respiratory problems although they had not completely disappeared. The administration of Baytril on day 2 also lead to less E. coli neonatal diarrhoea. Calves with signs of BRD aged 1-1.5 months responded well to therapy and only a few relapsed without growth retardation. The accounting data showed a reduction of antibiotic cost against BRD from 1115 euro to 558 euro. Serology of calves aged 6-8 months showed that 5 of 7 calves were positive (71%) as compared to 8 out of 8 (100%) the previous year.

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Conclusions The sanitation program reduced the antibiotic cost by 50%, gave less BRD signs and 29% less seropositive animals. The program must be continued until the M. bovis herd prevalence has dropped to an acceptable level. Key words: Mycoplasma bovis, enrofloxacin, biosecurity 643 Analysis of Pathways for Release of Rift Valley Fever Virus into the Continental United States R. Kasari , A. Carr , V. Lynn , T. Weaver 1 USDA-APHIS-VS-CEAH, National Surveillance Unit, Fort Collins, United States 2 Uniformed Services University of the Health Sciences, Department of Preventive Medicine and Biometrics, Bethesda, United States 3 USDA-APHIS-VS-CEAH, Center for Emerging Issues, Fort Collins, United States Objectives of Study: The primary objectives of this study were to evaluate the feasibility of RVFV to enter the continental United States by various routes as well as identify the states whose domestic and wild ruminant and human populations are most vulnerable for exposure to this virus. Materials and Methods: Pathways analysis methods were used to identify the pathways for release of RVFV into the continental United States. Initially, the agent, host, and environmental factors important in the epidemiology of RVF were used to develop a list of potential pathways for release of RVFV into the continental United States. Five pathways were identified: (1) importation of RVFV-infected animals, (2) entry of RVFV-infected people, (3) mechanical transport of RVFVinfected insect vectors, (4) intercontinental wind-borne transport of RVFV or RVFV-infected insect vectors, and (5) smuggling of live virus. Next, the feasibility of each pathway was evaluated using data from governmental and public domain sources to evaluate animals, commodities, and people that moved from RVF-endemic countries to the continental United States between calendar years 2000 and 2005. Finally, entry points of each feasible pathway into the continental United States were used to identify the domestic and wild ruminant and human populations at risk for exposure to RVFV. Results: Feasible pathways for entry of RVFV into the continental United States are (1) importation of RVFV-infected animals, (2) entry of RVFV-infected people, (3) mechanical transport of RVFV-infected insect vectors, and (5) smuggling of live virus. Conclusions: A pathways analysis provides the requisite information needed to construct an effective targeted surveillance plan for this virus in order to enable rapid detection and response by animal health and public health officials. Domestic ruminant livestock, ruminant wildlife, and people in 14 states (Alabama, California, Florida, Georgia, Maine, Maryland, Massachusetts, Minnesota, New Jersey, New York, Pennsylvania, South Carolina, Texas, and Virginia) appear to be most vulnerable to exposure from RVFV. It should be noted however, that this group of "at-risk" states could be altered entirely, if this viral agent is released illicitly as part of a terrorist act. Key words: Rift Valley Fever, zoonotic disease, pathways analysis 644 Analysis of Pathways for Release of Ehrlichia ruminantium (Heartwater) into the Continental United States R. Kasari , E. Lawrence , H. Howe , L. Farnsworth , E. Freier , 2 2 M. James , S. Miller 1 USDA-APHIS-VS-CEAH, National Surveillance Unit, Fort Collins, United States 2 USDA-APHIS-VS-CEAH, Center for Emerging Issues, Fort Collins, United States Objectives of Study: The objective of this study was to evaluate the feasibility of Ehrlichia Ruminantium by itself or via E. Ruminantiuminfected Amblyomma spp. vector ticks to enter the continental United States by various routes as well as identify the States whose domestic and wild ruminant populations are most vulnerable for exposure to this rickettsial organism. Materials and Methods: Pathways analysis methods were used to identify the pathways for release of E. ruminantium into the continental United States. Initially, the agent, host, and environmental factors important in the epidemiology of E. ruminantium infections in susceptible animal species were used to develop a list of potential pathways for release of this rickettsial agent into the continental United States. Five pathways were identified: (1) importation of E. ruminantium-infected animal species, (2) migrating cattle egrets (Bubulcus ibis) serving as a

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transport host for E. ruminantium-infected Amblyomma spp. ticks, (3) mechanical transport of E. ruminantium-infected Amblyomma spp. ticks by humans and imported non-ruminant animals, reptiles, and other birds, (4) mechanical transport of E. ruminantium-infected Amblyomma spp. tick by fomites, and (5) smuggling of live E. ruminantium. Next, the feasibility of each pathway was evaluated using data from governmental and public domain sources to evaluate animals, commodities, and people that moved from heartwater-endemic African and Caribbean island countries to the continental United States between calendar years 2000 and 2006. Finally, entry points of each feasible pathway were used to identify the States whose domestic and wild ruminant populations were at greatest risk for exposure to E. ruminantium. Results: All five pathways were feasible ones for entry of E. ruminantium or E. Ruminantium-infected Amblyomma spp. vector ticks into the continental United States. Conclusions: Compared to other states, ruminant livestock and ruminant wildlife species in 17 states (Alabama, California, Florida, Georgia, Illinois, Louisiana, Massachusetts, Maryland, Minnesota, Mississippi, North Carolina, New Jersey, New York, Pennsylvania, South Carolina, Texas, Virginia) should be most vulnerable to exposure to Amblyomma spp. ticks, some of which could be infected with E. ruminantium. This group of "at-risk" states could be altered entirely, if the rickettsial agent is released illicitly as part of a terrorist act. Key words: heartwater, Ehrlichia ruminantium, pathways analysis 645 Epidemic of Bovine Viral Diarrhea Virus (BVDV) Infection Originated from 3 Persistently Infected Cattle on a Farm during a 5-year Period M. Tajima , K. Sano 1 Hokkaido University, Veterinary Teaching Hospital, Sapporo, Japan 2 NOSAI Iburi, West branch, Toya, Japan Bovine viral diarrhea virus (BVDV) is distributed all over the world. Persistently infected cattle with BVDV (PI) are a latent source of infection of BVDV in the herd, and can cause serious economic damage. Testing and culling of PI are important procedures for dairy production. We encountered 3 outbreaks of BVDV infection in a farm during a 5-year period, and these were suspected to have caused by 3 individual PI. First epidemic: In May 2002, a 6-month-old Holstein calf was diagnosed as PI because of mucosal disease. A sister cow (primipara) of the PI calf and their dam (PI-1) were diagnosed as PI, and the bulk tank milk (BTM) test was positive for BVDV. After exclusion of all PIs, BTM test became negative for BVDV. The negative results were obtained twice at an interval of 6 months, therefore, the test of BVDV was discontinued. Second epidemic: In January 2005, 2 calves (pneumonia and poorgrowth) were diagnosed as PI. Their dams were BVDV negative and BTM test was positive again for BVDV. All cattle on the farm were examined for BVDV. Six PIs were detected, a milking cow imported from the cattle market in May 2004 (PI-2), 4 Holstein calves and a beef calf. By examination of all neonatal calves thereafter, 14 PIs were identified during a 6-month period. During this period, 50 milking cows were imported into the farm. All of these animals were BVDV negative, but 5 calves produced from these cows were diagnosed as PI. Except for these 5 calves, 9 calves and PI-2 were infected with the same BVDV based on the analysis of E2 diversity of the isolated BVDV, however, the BVDV differed from that involved in the first epidemic on the farm. After exclusion of all PIs, there were no PIs for a 3-month period. Third epidemic: In October 2005, an imported dam (PI-3) and neonatal calf were diagnosed as PI. At this time, the importation of cattle was discontinued, and the examination of newborn calves for BVDV was continued. After 6 months, 2 PIs were detected. All 4 PIs were infected with the same BVDV, and BVDV from PI-3 differed from those of PI-1 and -2. After the final detection of PI, approximately 250 newborn calves were examined for PI during a one-year period (till May 2007). There were no PIs and BTM test was negative every 6 months; therefore, the farm was recognized as BVDV free. Continuous BTM test and examination of newborn calves are the most important and effective procedures for BVDV eradication. Key words: BVDV, eradication, test and cull 646 Paraclinical Findings of Chronic Extensive Peritonitis in a Diarrheic Young Calf N. Atyabi, N. Alidadi, M. Hashemian Faculty of Veterinary Medicine, University of Tehran, Animal Science, Tehran, Iran

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86 XXV. Jubilee World Buiatrics Congress 2008

A 5-month-old calf with a chronic diarrhea and bloat and obvious abdominal pain was referred to the large Animal Teaching and Research Hospital. On physical examination the calf was lethargic and had a rectal temperature of 39.8 °C, respiratory rate 44 breaths per minutes, heart rate 100 beats per minutes and mild tachycardia. The calf had pale mucosal membranes and clinical dehydration. The peritoneal fluid examination showed an exudate with high protein level and degenerated neutrophil which was positive for Salmonella dublin. The fecal samples were negative for parasites and Mycobacterium paratuberculosis, but positive for Salmonella dublin. The leukogram showed significant leukocytosis, lymphocytosis and monocytosis accompanying by neutropenia and a shift to left. A degenerative anemia was diagnosed in the calf which could be a ambiguous prognosis. The animal died despite of relevant fluid and antibiotic therapy, and a chronic extensive peritonitis was found in the postmortem examination Key words: calf, chronic extensive peritonitis, diarrhea, leukogram 647 Molecular Survey on Occurrence of Mycoplasma Mycoides Subspecies Mycoides L.C and S.C in Pulmonary Infection of Iranian Cattle, Goats and Sheep N. Atyabi, Z. Nikusefat, SA. Pourbakhsh Faculty of Veterinary Medicine, University of Tehran, Clinical Science, Tehran, Iran This survey described microbiological and molecular study for detection Mycoplasma mycoides subspecies mycoides (L.C and S.C) in pneumonic lungs of Iranian sheep, goats and cattle. Suspected samples were investigated based on color, consistency and appearance of the cut surface. Total 180 lung samples were collected from slaughter house located in West of Iran, Kermanshah during 2006-2008 and transferred near ice packs to Razi Research Institute. Gross lesions showed hepatization with grey and white lesions (consolidation) and motley appearance with or without fibrin. Minced lung tissues were inoculated to PPLO broth agar (Diffco) with application acetatecellulose filter (0.45-2.5 micrometer). Samples were in 6.8% carried to Co2 incubator. After 10-14 days the yellow tubes were subcultured to PPLO plate. After while, characteristic colony was observed. DNA extractions were based on phenol method and DNA extracted isolates were preserved in 50% glycerol at -20°C.DNA extract of all samples were subjected to generic and species specific PCR based on 16 S rRNA with different set of primers and cycles. The visualized amplicon consisted of 573 bp.40 samples from total 180 lung extract were positive for genus mycoplasma (22.2%),while only 30 samples were positive in culture(16.66%) .There were no significance difference in sex and age between affected animals.(P>0.05) The highest percentage of infection was observed during December (32%) and the lowest was in June (2.5%).There were no evidence of Mycoplasma mycoides mycoides (S.C ) and (L.C) by species specific primer based on CAP-21 PCR in infected ruminants. However, the present work was carried out to study the incidence of two strains of mycoid cluster in Iranian ruminants. Further investigation should be conducted to profile all mycoplasma spp in pulmonary infections. Key words: Mycoplasma mycoides, sub. mycoides (L.C and S.C), ruminant, Iran 648 Application of PCR and Hematological Findings in Replacement Dairy Heifers with ELISA Seropositive to Bovine Leukosis Virus Infection T. Rukkwamsuk, S. Panneum Faculty of Veterinary Medicine, Kasetsart University, Large Animal and Wildlife Clinical Science, Nakhon-Pathom, Thailand Bovine leukosis virus infection was studied in 171 replacement dairy heifers. Blood samples were collected for determination of hematological parameters, detection of bovine leukosis virus using PCR technique, and determination of antibody against bovine leukosis virus infection using indirect ELISA (IDEXX HerdCheck Anti-BLV). Blood samples from seropositive and seronegative heifers were randomly selected to detect bovine leukosis virus using PCR technique. Results revealed that seroprevalence of bovine leukosis virus infection in this group of heifers was 19.3% (33 heifers were seropositive and 138 heifers were seronegative). Most hematological parameters did not differ between seropositive and seronegative heifers. Average total white cell counts were higher for seropositive heifers than for seronegative heifers (P = 0.08). Regarding PCR results, bovine

leukosis virus was not detected from some seropositive heifers but could be detected from some seronegative heifers. However, results obtained from indirect ELISA and from PCR technique had a moderate agreement (Kappa value = 0.60). In conclusion, seroprevalence of bovine leukosis virus infection in replacement dairy heifers was relatively high. Increased total white cell count seemed to be related to bovine leukosis virus infection. Although agreement of serological test using ELISA and virus detection using PCR was moderate, results of these tests were still contradicted in some heifers. Therefore, further study on increasing sensitivity and specificity of diagnostic tests is required. Key words: Bovine Leukosis Virus, hematology, replacement heifer 649 Bovine Leptospirosis in Sardinia: Isolation and Evaluation of Molecular Methods M. Ponti, G. Sanna, G. Carboni, G. Canu, M. Manca, M. Noworol, B. Palmas, E. Marongiu, C. Patta Istituto Zooprofilattico Sperimentale Della Sardegna, Sassari, Italy Previous studies carried out in Sardinia have shown that Leptospirosis affects both humans and farm animals and that Leptospira serovar pomona is responsible for the icterus-haemorrhagic syndrome in calves. In this study, the microscopic agglutination test (MAT) was used to determine the serological titers of the serum samples collected from cattle. Even though definitive diagnosis is provided by culture of pathogenic Leptospira, several molecular approaches were also evaluated, aiming at optimizing the detection of bacteria in bovine samples of urine and blood. Specimens were collected from two dairy cattle herds in the north of Sardinia. Farm A consisted of 580 subjects, out of which 260 were lactating cows, 280 were freerange heifers and 40 were calves. Farm B included 120 animals, out of which 45 were lactating cows and the remaining ones were heifers and calves. Sera were examined by MAT. Urine, blood and tissue suspension were inoculated in semi-solid medium for cultivation and tested by PCR. The Leptospires strains isolated were identified by using monoclonal antibodies. Several primer pairs for detection of leptospiral DNA were tested: one set amplified a fragment of the 16S rRNA, another set was complementary to a portion of Lig protein gene (Lig 1/Lig 2) and a third one used G1/G2 and B64I/B64II. Then a protocol of real-time PCR using a Taqman Probe was standardized and compared with the three protocols previously described. In farm A the seroprevalence was of 60%, while in farm B it was about 40%. Leptospires were isolated from 8 and 3 urine samples in farms A and B, respectively. All strains were identified as Leptospira serovar pomona. Leptospires were not isolated from blood and fetuses samples. All the molecular methods used amplified leptospiral DNA from all 15 serovars of our Leptospires panel, and also from blood, urine and fetuses tissues. Primers derived from rRNA gene sequence were the least specific, and none of the molecular methods tested was 100% sensitive. Furthermore, the main limitation of these PCR-based assays was their inability to identify the infecting serovar. Real-time PCR assay, which successfully detected leptospiral DNA, is rapid and specific but more expensive than conventional PCR. A combination of two detection methods (PCR and culture) is the most sensitive approach for early diagnosis of Leptospirosis. Key words: leptospirosis, bovine, PCR 650 Serological Surveillance of Contagious Bovine Pleuoropneumonia between 2005 and 2007 Years in Romania R. Radulescu, G. Petriceanu, A. Ragalie, E. Gutu Institute for Diagnosis and Animal Health, Immunology, Bucharest, Romania The reference method recommended by OIE for serological surveillance of Contagious Bovine Pleuropneumonia (CBPP) is the Complement Fixation Test (CFT). This method had been used in the past for eradication of CBPP in many countries. However, CFT have some disadvantages, mainly the production and standardization of specific antigen, providing some false positive reaction due to the mycoplasmas cross-reactivity from M. mycoides cluster. In Romania between 2005 and 2007, years have been examined for CBPP by CFT 5306 samples taken from cattle, deer, wild sheep, reindeer and buffalo. The samples were analyzed with two CFT antigens Mycoplasma mycoides subsp. mycoides provided by CBPP OIE Reference Laboratories from LINV Lisbon-Portugal and ISZ G.Caporale Teramo-

Infectious and Zoonotic Deseases (Public Health) 87

Italy. After the sample examinations, 90 (1.69%) were positive, 146 (2.75%), doubtful, 77 (1.45%) anti-complementary and 4993 (94.13%) were negative. At the reanalyzing of those positive, doubtful and anticoplementary samples after a short period of time all of them were negative. The specificity of CFT had been calculated using both antigens and the following results were obtained: 95-99% with Portuguese antigen and 82-100% with Italian antigen. From the 649 CFT examined samples using the Italian antigen which revealed a specificity of 82%, we examined by competitive ELISA (cELISA) all the positive (20), doubtful (85), anticomplementary (6) and 59 negative samples. From 170 examined samples only 12 presented doubtful results and the others were negative. For the introducing in the routine work of cELISA we also evaluated the precision parameter of the method by calculation of the repeatability and reproducibility of the obtained results. The variation coefficient (CV) of the reproducibility of control sera obtained results had been between 6.3% and 6.8% in comparison with 8% CV reported by the producer. Taken into consideration that cELISA had revealed a very good specificity of 92% and a correlation index of 41.76% against CFT results it is recommended that any anticomplementary, doubtful or positive results in CFT to be also analyzed by cELISA. Key words: M. mycoides, CFT, ELISA, CPBB, antigen 651 Compatibility of Inactivated Bovine Viral Diarrhoea (BVD) and Leptospira Interrogans Serovar Hardjo Vaccines I. Mawhinney , B. Makoschey 1 ICM Broadlands, Bury St. Edmonds, United Kingdom 2 Intervet-Schering Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: The purpose of this study was to examine whether concurrent administration of a Leptospira vaccine affects the immunogenicity of an inactivated bovine viral diarrhoea (BVD) vaccine. Materials and Methods: Twenty beef breeding animals were included in the trial. At the beginning of the study, one animal had very low antibody titers against BVDV the remaining animals were seronegative. All animals were seronegative for Leptospira prior to the vaccinations. All animals were vaccinated intramuscularly with a commercially available BVD vaccine (Bovilis® BVD, Intervet). Ten of them (group A) were vaccinated subcutaneously at the same time but on the opposite site of the neck with an inactivated Leptospira interrogans serovar hardjo vaccine (Leptavoid® H, Schering Plough). The remaining 10 animals (Group B) were vaccinated two weeks later with the Leptospira vaccine following the manufacturer recommendation. The BVD vaccination was repeated after four weeks in both groups. The animals of group A received a second dose of the Leptospira vaccine at the same time. The animals of group B were revaccinated with the Leptospira vaccine 2 weeks after the BVD booster. Blood samples were taken at the first and second BVD vaccination (days 1 and 28) and two and four weeks after the second vaccination (days 42 for group A and 42 and 56 for group B). The samples were processed into serum for antibody testing. Antibodies specific for BVD virus were measured by serum neutralisation test. Leptospira serology was carried out against each of the two hardjo serovars, prajitno and bovis using a standard MAT test (VLA, Weybridge). Results: There was no statistically significant difference when the serological response to the BVD vaccine was compared between the two groups. Likewise, there was no statistically significant difference in the Leptospira serology, neither for prajitno, nor for bovis when the antibody responses two weeks after the second dose of the Leptospira vaccine was compared between the two groups. Conclusions: From these data it was concluded that there was no negative effect on the neutralising antibody response when the two vaccines were applied concurrently (at the same time but on opposite sides of the animal). 652 Simultaneous Infection of Calves with Maternal Antibodies with Bovine Rotavirus and Coronaviruses under Experimental Conditions U. Schmidt , M. Timmermans , P. Tonen , B. Makoschey 1 Intervet-Schering Plough, R&D, Boxmeer, The Netherlands 1 Intervet-Schering-Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: Newborn calf diarrhoea is generally recognized as multifactorial disease. The list of pathogens that can be involved is

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quite extensive, comprising bovine rotaviruses, bovine coronaviruses, E. coli and Cryptosporidium parvum as the most important ones. In the present study, clinical signs after simultaneous infection of newborn calves with two rotaviruses and a coronavirus under laboratory conditions should be compared to the infection with either of the three pathogens alone. Moreover, it should be investigated, whether all three viruses replicate after co-infection. Materials and Methods: The experiment was divided in four separate studies. On days 5, 6 and 7 after birth, the calves were infected orally with bovine rotavirus P5G6 (study 1), bovine rotavirus P11G10 (study 2) and bovine coronavirus DB-2 (study 3) or all three viruses together (study 4). After the infection, the calves were observed twice daily for clinical signs of diarrhoea. From the day of infection onwards, fecal samples were taken twice daily. Virus shedding was quantified by ELISA technique. Results: All 12 animals infected with a single virus survived until the end of the observation period and some animals developed only very mild or no diarrhoea. By contrast, all four animals infected simultaneously with all three viruses had liquid faeces during at least one observation moment and two of the four animals had to be killed on humane reasons after episodes of severe diarrhoea. Both, rotavirus and coronavirus was detected in the faeces of all four animals infected simultaneously with the three viruses during several days. Moreover, both rotavirus G-types were present in a faecal sample from a coinfected calf. Conclusions: From these data, it can be concluded, that the coinfection of calves with rota- and coronaviruses under laboratory conditions reflects the field situation, where co-infection of pathogens causes more severe disease than infection with single pathogens. Moreover, the results reported here confirmed, that colostrum from unvaccinated dams did only partly protect against diarrhoea. Experimental data and field experiences have demonstrated the benefit of cow vaccination against enteric pathogens. 653 Serum-free Produced Live Virus Vaccines are Efficacious and Safe B. Makoschey , P. Gelder 1 Intervet-Schering Plough, Int. Marketing, Boxmeer, The Netherlands 2 Intervet-Schering Plough, R&D, Boxmeer, The Netherlands Objectives: Most cell culture based manufacturing processes for animal and human vaccines use bovine fetal serum (FCS). In potential, FCS can be contaminated with a number of viruses that can infect the bovine foetus transplacentally, especially with the bovine viral diarrhoea virus (BVDV). Therefore, we have developed a novel production procedure for bovine herpesvirus type 1 (BoHV-1) live virus vaccines that does not make use of serum or fractions thereof during any cell or virus passage, thus completely serum-free. It should be determined whether serum-free produced BoHV-1 viruses are suitable as live vaccines for use in cattle. Materials and Methods: The safety of serum-free produced BoHV1 virus was established in two separate studies. In the first study, six colostrum deprived calves each were vaccinated intranasally (IN) or intramuscularly (IM) with an overdose and two weeks later again with a single dose. In a second study, ten seronegative calves at the age of five to six months were vaccinated four times at fortnightly interval with an overdose, both IN and IM. The efficacy was tested in a vaccination-challenge study in seronegative calves as laid down in the European Phamacopeia. For comparison, a vaccine batch with BoHV-1 grown in serum-containing cell culture medium was included in the study. The calves were monitored for clinical signs after challenge infection with BoHV-1 wild type virus and challenge virus excretion was measured. Finally, the stability of the virus was determined by titration of the freeze-dried vaccine after storage at temperatures of +4°C and -20°C for more than one year. Results: There were no local or systemic reactions after vaccination observed in either of the two safety studies. With regard to efficacy, the serum-free produced vaccine performed equally well as the conventionally produced one and both met the strict requirements as laid down in the European Phamacopeia. Finally, the virus titers of the freeze-dried vaccines were stable for more than one year at temperatures of +4°C and -20°C. Conclusions: These data demonstrate the safety and efficacy of serumfree produced live vaccines in cattle as well as the stability of these products.

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654 Complete Protection from Renal Infection with Leptospira Borgpetersenii Serovar Hardjo in Calves after Transfer of Specific Antibodies E. Klaasen , T. Mols , M. Smit , B. Makoschey 1 Intervet-Schering-Plough, R&D, Boxmeer, The Netherlands 2 Intervet-Schering-Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: The purpose of this study was to examine whether administration of serum antibodies can protect cattle from renal infection with Leptospira borgpetersenii serovar Hardjo, even if challenge is performed as long as 7 weeks after serum transfusion. Materials and Methods: Ten conventional 5-week-old calves free of serum antibodies against serovar Hardjo were randomly assigned to two groups of five animals each. Each calf of group 1 received a transfusion of a hyper-immune serum with a titer of 9.1 [log2/ml] of antibodies specific for serovar Hardjo. The calves of group 2 (control group) were left untreated. Seven weeks later all animals were challenged on three consecutive days by conjunctival instillation of Leptospira borgpetersenii serovar Hardjo genotype hardjobovis. Serum samples were taken from the calves of groups 1 and 2 immediately prior to the transfusion as well as 24 hours, 4 weeks and 7 weeks thereafter. Antibodies specific for serovar Hardjo, were measured by ELISA. Urine samples were taken pre-challenge and 2, 3, 4, 5 and 6 weeks post-challenge to assess the effect of the challenge. The calves were sacrified 6 weeks post-challenge and kidney tissue was taken. Urine samples and kidney tissue homogenates were cultured. The cultures were examined every 3 weeks with dark-field microscopy. Cultures were considered negative when no leptospires were detected after 18 weeks of incubation. Results: The titration of serum samples taken before the transfusion confirmed that the calves of both groups were free of serum antibodies against serovar Hardjo and the untreated animals in group 2 remained seronegative until the time of challenge. By contrast, mean titers of 5.8, 4.0, and < 3.3 were measured in the samples taken from animals of group 1 at 24 hours, 4 weeks and 7 weeks after transfusion respectively. Renal infection was demonstrated in four out of five control animals, whereas in none of the five pre-treated calves renal infection could be detected. Conclusion: In this study, complete protection of cattle from renal infection with serovar Hardjo was obtained with merely humoral immunity through passive immunization. In this study, the protection lasted for at least 7 weeks after transfusion. At this time point, the serum titre of antibodies had declined below the detection limit of the ELISA. 655 Compatibility of a Live IBR Marker Vaccine and an Inactivated BVDV Vaccine B. Makoschey , J. Munoz Bielsa , L. Santos , M. Alvarez 1 Intervet-Schering-Plough, Int. Marketing, Boxmeer, The Netherlands 2 Intervet-Schering-Plough, Laboratorios Intervet S.A., Salamanca, Spain 3 University of Leon, Animal Health Department, Leon, Spain Objectives: The target animals and vaccination regimes for vaccines against the bovine rhinotracheitis (IBR) and the bovine viral diarrhea virus (BVDV) are very similar. In order to simplify animal handling it would be desirable to combine the two vaccination procedures. Therefore, we have examined whether a live IBR marker vaccine (Bovilis® IBR marker live, Intervet) and an inactivated BVD vaccine (Bovilis® BVD, Intervet) can be applied at the same time. Materials and Methods: A total of 57 head of cattle were included in the study and seven groups of animals were established. The animals were vaccinated on days 0, 28 and 208 according to different (combined) vaccination schedules. The immune responses against the bovine herpesvirus type 1 (BoHV-1) and BVDV were determined and compared for the different vaccination schedules. Results: BoHV-1 antibody titers after simultaneous (together in the same syringe) or concurrent (two separate injections) application of the inactivated BVD vaccine were not lower than after vaccination with the BoHV-1 vaccine alone. To the contrary, BoHV-1 titers were even higher after simultaneous application at the first or second dose of the basic vaccination course of the BVD vaccine and at the booster vaccination. Likewise, BVDV antibody titers after simultaneous or

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concurrent application of the live BoHV-1 vaccine at the first or second dose of the basic vaccination course of the BVD vaccine and at the booster vaccination were not different from the titers after vaccination only with the BVD vaccine. Conclusions: There was no negative effect on the antibody response against either of the two vaccines when the live IBR marker vaccine was applied simultaneously (together in the same syringe) or concurrently (two separate injections) with the first or second dose of the basic vaccination course of the BVD vaccine and then at the booster vaccination (3rd dose). 656 Compatibility of an Inactivated IBR Marker Vaccine and an Inactivated BVDV Vaccine B. Makoschey , J. Donate , L. Santos , M. Alvarez 1 Intervet-Schering-Plough, Int. Marketing, Boxmeer, The Netherlands 2 Intervet-Schering-Plough, Laboratorios Intervet S.A., Salamanca, Spain 3 University of Leon, Animal Health Department, Leon, Spain Objectives: The target animals and vaccination regimes for infectious bovine rhinotracheitis (IBR) and the bovine viral diarrhoea virus (BVDV) vaccines are very similar. To simplify animal handling on the farm it would be desirable to combine both vaccination regimes. We have performed a study to assess the efficacy in terms of antibody response of the simultaneous (both vaccines in the same syringe) and concurrent (both vaccines at the same time, but different injection sites) use of Bovilis® BVD (Intervet, The Netherlands) and Bovilis® IBR marker inac (Intervet, The Netherlands) in dairy cattle under field conditions. Materials and Methods: The study was carried out as a randomised, partially blinded and controlled field trial on a commercial dairy farm in Spain. All animals included in the study were shown to be free of antibodies against the IBR and the BVD virus prior to the first vaccination. Forty animals were included in the study and assigned at random to one of the four groups of ten animals each. The vaccination protocols were as follows: group A received only inactivated IBR marker vaccine, group B received only BVD vaccine, group C were vaccinated simultaneously (in the same syringe) with both vaccines, group D were vaccinated concurrently (both vaccines at the same time, but in two separate injection sites) with the two vaccines. All animals were vaccinated twice (at admission (d0) and four weeks later (d 28). Results: The neutralizing antibody response against BVDV did not reveal any differences between the group vaccinated only with the BVD vaccine and the groups that were vaccinated simultaneously or concurrently with the IBR marker vaccine. Likewise, the neutralizing antibody titers against BoHV-1 did not exhibit any negative effect by the simultaneous or concurrent use of the two products as compared to the single IBR marker vaccination. Conclusions: These results indicate that the two vaccines can be applied at the same time either at the same or at different injection sites without any negative effect on the neutralising antibody response against the two vaccine viruses. 657 A Serological Study on Bovine Viral Diarrhea - Mucosal Disease (BVD-MD) in Kurdistan Province Sh. Fakur , Fa. Hemmatzadeh 1 Faculty of Veterinary Medicine Islamic Azad University Sanandaj, Large Animal Clinical Sciences, Sanandaj, Iran 2 Facutly of Veterinary Medicine, University of Tehran, Large Animal Clinical Sciences, Tehran, Iran Objectives of study: Bovine viral diarrhea disease is one of the viral diseases in cattle that was reported 10-90 percent in many countries. The causative RNA virus is a member of the family togaviridae and the genus pestivirus which is antigenically closely related with the virus of BD in sheep. Cattle are the most sensitive species to virus and are considered the principal reservoir of BVD viruses. A large proportion of BVD infections are subclinical and majority seropositive cattle. Since the virus has affinity for the lymphoreticular tissue this can result immonosuppression and abortion, produce calves with congenital abnormalities and persistant infectious (PI), So diagnostic laboratory procedures such as virus isolation and serologic methods are required. Materials and methods: In this study 410 serum samples were tested by serum neutralization test (SN) by using the NADL strain of BVD virus.

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Results: The cows sera were collected from Kurdistan province of Iran. By SN test the rate of infection in all samples was 27.7%. This result done with similar study was approximately like that one carried out on 1357 serum samples from other provinces in Iran. Conclusions: Being on the border and consequently animals entering the country explains the high rate infection in Kurdistan province. Key words: viral diarrhea, serum neutralization, Sanandaj 658 Treatment of Infectious Diseases in Sheep with Enrofloxacin 10% (Kinetomax) - Case Reports MI. Ferreira , EJ. Facury Filho , AU. Carvalho , I. Borges , 1 3 G. Macedo Junior , N. Martins 1 Escola de Veterinária da UFMG, Departamento de Zootecnia, Belo Horizonte, Brazil 2 Escola de Veterinária da UFMG, Departamento de Clinica e Cirurgia Veterinaria, Belo Horizonte, Brazil 3 Escola de Veterinária da UFMG, Departamento de Medicina Veterinária Preventiva, Belo Horizonte, Brazil Enrofloxacin 10% (Kinetomax) is a potent chemotherapist agent showing bactericidal action. It has a broad spectrum of antimicrobial action presenting rapid absorption, distribution and metabolism in the body. Thirty-two crossbred LacaunexSanta Ines sheep were allocated to individual stalls and kept in confinement in the Laboratory of Animal Calorimetry and Metabolism, School of Veterinary Medicine at UFMG, for 120 days in lactation for researching on production and composition of milk. Case 1: seven animals - five sheep and two lambs - had acute inflammatory reaction of the conjunctiva; sclera hyperemia; photophobia; blepharospasm; lacrimation; mucopurulent ocular discharges; and corneal opacity, without ulceration, resembling keratoconjunctivitis. Enrofloxacin 10% (Kinetomax) was used for treatment in accordance with the recommendation of 7,5 mg/Kg /b.w., in all affected animals. On the fourth day after application of the product, the reduction of clinical symptoms was observed, with complete recovery of the animals. Prior to the treatment onset, samples of ocular secretion were collected, with the help of sterile swabs, and sent to the Laboratory of Microbiology at the Department of Preventive Veterinary Medicine, where Staphylococcus caprae was isolated. Ten days after treatment, two animals showed clinical relapse and the occurrence of two new cases. The animals were subjected to treatment, following the fore-cited protocol. On the fourth day after the second treatment, there was complete reduction of clinical symptoms. Case 2: monitoring the daily consumption, a gradual decrease of food intake and reduced milk production of one animal was observed for five consecutive days. In the clinical examination, there was an increasing in volume, redness, and heat on the crown of the left forelimb hoof (coronary band), so that the animal could not stand, featuring an interdigital phlegmon (footrot), in initial state of development. For treatment, enrofloxacin 10% (Kinetomax) was used at a dose of 7,5 mg/Kg /b.w., along with flumexine meglumine at the dose of 2,2 mg/Kg. Hoof trimming was made in the hoofs where no deeper injuries were found. On the following days, there was an increase of food intake and the recovery of the clinical signs was complete on the third day after treatment. Key words: footrot, keratoconjunctivitis, Kinetomax, sheep 659 Efficacy Following a Flexible Dose Regimen of SpirovacR, a Leptospira Borgpetersenii Serovar Hardjo (Type Hardjobovis) Bacterin, in a Challenge Study in Cattle D. Cleary , S. Behan , J. Wong , M. Fisher , S. Saginala , L. Kopta , 1 1 1 1 1 2 M. Tory , A. Weber , G. Gallo , R. Leyh , J. Galvin , J. Salt 1 Pfizer Animal Health, Veterinary Medicine Research and Development, Kalamazoo, Michigan, United States 2 Pfizer Animal Health, Animal Health Research Division, Sandwich, United Kingdom A research study was conducted to demonstrate the protection in cattle afforded by a flexible dosing regimen using a Leptospira borgpetersenii serovar Hardjo (type hardjo-bovis) bacterin (Spirovac, Pfizer Animal Health). All animal care and procedures were approved by an ethical review board for compliance with animal welfare guidelines. Spirovac (n=20) or placebo (n=11) treatments were randomly assigned to 4-week-old L. borgpetersenii seronegative [microscopic agglutination test (MAT) antibody titers of [1:12.5] heifer calves. The Spirovac' group of calves received a second bacterin dose

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a minimum of four months later. Three weeks following administration of the booster dose, all animals, including the placebo-vaccinated control group, were challenged for three consecutive days via conjunctival instillation of a heterologous strain of L. borgpetersenii. Urine samples were collected for bacterial culture up to five weeks following challenge when the animals were humanely euthanized and kidney samples were collected. All animals were seronegative to L. borgpetersenii at the time of first vaccination and all placebo control animals were seronegative at the time of the challenge. The primary variable utilized for the evaluation of protection was bacterial isolation (presence or absence) of L. borgpetersenii in the urine following challenge and in the kidneys at necropsy. Ninety-one percent (10/11) of the placebo control animals were positive for L. borgpetersenii isolation in both urine and kidney samples, confirming a valid challenge. No L. borgpetersenii was isolated from the urine or kidneys in the group receiving two doses of bacterin (0/20). These results confirm that two doses of Spirovac administered approximately four months apart can protect calves against virulent challenge. Key words: Leptospira hardjo, Spirovac, vaccines, immunology, cattle 660 Survey on Dairy Cows Infective Abortions in the North-East of Italy A. Barberio , N. Pozzato , L. Ceglie , S. Nardelli , M. Badan , 1 G. Vicenzoni 1 Istituto Zoop. Sper. Venezie, Sezione Verona e Vicenza, Vicenza, Italy 2 Istituto Zoop. Sper. Venezie, Dipartimento Sanita' Animale, Legnaro (PD), Italy Objectives of study: The purpose of this study is to describe the results of the abortion surveillance program in Veneto region (North-East of Italy) over a period of two years. Materials and Methods: Aborted fetuses accompanied by the dam's blood sample were delivered to the Regional Veterinary Laboratory (IZS), and submitted to a panel of laboratory tests. The cows sera were tested for antibodies against Brucella, Neospora, IBR, Leptospira, and BVD non structural protein NS2-3. ELISA tests were used for Neospora (VMRD), BVD (Pourquier) and IBR (Bommeli) antibodies detection, while microagglutination with the method of Martin and Petit was used for Leptospira. Brucella antibodies were detected following the EU regulation mandatory method. Necroscopy and routine bacteriology were performed on all fetuses, while Brucella spp isolation was carried out only if the abortion occurred after the 5th pregnancy month; Campylobacter fetus isolation was performed on abortions occurred before the 5th month. In case of positive results for BVD or N. caninum antibodies in mother serum, identification of these microorganisms in fetus was performed with RT-PCR for BVDV and PCR for Neospora. Results: During year 2005 and 2006, 577 abortions were delivered to IZS laboratories: 30% of abortions (170) were due to infective microorganisms and among these N. caninum was the most frequent (63%). Among the other microorganisms found, BVDV was isolated in 14% of positive fetuses, Arcanobacterium pyogenes in 9%, and Listeria monocytogenes in 4%. Considering that only fetuses aborted by serological positive cows were tested with PCR for N. caninum, the percentage of fetuses found infected by this parasite was very high (47%), and is in agreement with the reports indicating that cows with N. caninum antibodies are more likely to abort than seronegative cows. Serological tests showed a high prevalence of antibodies against BVD virus (54%), Neospora caninum, (39%), and IBR virus (32%). On the contrary there was a low frequency (3%) of positive sera for Leptospira, and among the 16 positive samples, only two cows had a serological titer high enough to be considered the consequence of a recent infection. Conclusions: In our opinion the abortion surveillance program provided many useful and interesting information about the health status of dairy farms and the activities to be performed to prevent abortion diseases in the future, besides guaranteeing a mandatory requirement for the declaration of officially brucellosis-free region. 661 Severe Outbreak of Malignant Catarrhal Fever in a Dairy Herd T. Autio , P. Syrjälä , M. Hautaniemi , U. Rikula , S. Pelkonen 1 Finnish Food Safety Authority Evira, Production Animal Health, Kuopio, Finland

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Finnish Food Safety Authority Evira, Veterinary Bacteriology, Kuopio, Finland 3 Finnish Food Safety Authority Evira, Veterinary Virology, Helsinki, Finland Objectives of the study: Sheep-associated malignant catarrhal fever (SA-MCF) is a severe and mostly fatal disease of cattle and other ruminants. Severe outbreaks have been described in bisons that are the most susceptible animals. Sheep-associated MCF has been linked to ovine herpesvirus-2 (OHV-2) infection. We describe here an outbreak of SA-MCF with high mortality in a Finnish dairy farm. Materials and Methods: The farm. The affected herd consisted of appr. 40 dairy cows and heifers, calves and young cattle. All animals were housed in the same insulated barn with shared air conditioning, operated with forced exhaust ventilators. Free stall was used for cows, and young cattle were raised in group pens. The farmer brought two lambs, born in spring 2005, into the dairy barn in November 2005 and the lambs stayed in the barn until early April 2006. Examinations: Pathological examinations were carried out and diagnoses were verified by OHV-2 specific PCR. Serum and EDTA blood samples were collected from all living animals in early April 2006 for analyses of OHV-2 (PCR) and MCF-antibodies (c-ELISA). Results: In early January 2006 the exhaust ventilators were off for eight hours due to a power failure. Thus the air quality in the barn was extremely poor with high humidity and severe condensation. First signs of MCF were seen in February and the last case was in November 2006. The peak of the cases was at spring time. The sings including high fever, milk drop in cows, hyperaemic oral and nasal membranes, depression, oculo-nasal discharge and diarrhoea were seen in a total of 31 animals including cows, calves, heifers and young cattle. One cow with typical clinical signs, high titre of specific antibodies and positive OHV-2 PCR result recovered. The remaining 30 animals with signs of MCF died or were euthanized. The diagnosis of MCF was based on gross and histopathology and presence of both OHV-2 DNA and specific antibodies. The animals were also tested for other agents of upper respiratory tract and intestinal infections including respiratory syncytial virus, infectious bovine rhinotracheitis, bovine coronavirus, bovine viral diarrhoeal virus, mycoplasmas and salmonella, with negative results. Conclusions: The adolescent sheep between 6 and 9 months of age have been shown to shed OHV-2 virus more often and in greater amounts than adults. Here keeping of two adolescent sheep in a dairy barn caused a severe outbreak of MCF in a dairy herd resulting in death of 30 animals. 662 Evaluation of Nasopharyngeal Swabs in Sampling for Respiratory Pathogens in Calves T. Autio , T. Pohjanvirta , A. Huovilainen , L. Sihvonen , 2 S. Pelkonen 1 Finnish Food Safety Authority Evira, Production Animal Health, Kuopio, Finland 2 Finnish Food Safety Authority Evira, Veterinary Bacteriology, Kuopio, Finland 3 Finnish Food Safety Authority Evira, Veterinary Virology, Helsinki, Finland Objectives of the study: The aim of this study was to compare nasopharyngeal swab (NP) and bronchoalveolar lavage (BAL) samples in sampling for respiratory pathogens in calves. The samples were examined for presence of Pasteurella multocida, bovine coronavirus (BCV) and respiratory syncytial virus (RSV), which are the most common pathogens associated with calf respiratory disease in Finland. The similarity of Pasteurella multocida isolates from upper and lower respiratory tract was analysed. Materials and Methods: NP and BAL samples were taken in parallel from 42 calves. NP was taken with a guarded 27-cm swab and BAL with a guarded nasal tube. The samples were examined bacteriologically by standard laboratory methods. Pulsed-field gel electrophoresis (SalI) was utilized for genetic typing of P. multocida isolates. BCV and RSV were detected with RT-PCR. Results: Five animals were negative for the studied pathogens. Both P. multocida and BCV were detected in 14 animals. A combination of P. multocida, RSV and BCV was found in two animals, and three were infected with both P. multocida and RSV. A total of five animals harboured RSV, of which two in both samples and two only in NP. BCV was detected in nine animals by both sampling techniques and in

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eight and three animals only in NP or BAL, respectively. P. multocida was detected in 33 animals. Out of these, 23 calves harboured P. multocida both in NP and BAL. All BAL samples positive for P. multocida were also NP positive, and 10 only in NP. Paired P. multocida isolates obtained from 23 animals with both sampling methods were characterized by PFGE. Majority of animals (20/23) harboured isolates showing similar macrorestriction patterns (MRP) or had minor differences in their MRPs. Conclusions: Nasopharyngeal swabbing was found to be a suitable method for sampling for BCV, RSV and P. multocida. All pathogens were recovered more often in nasopharyngeal swab than in BAL samples. PFGE typing indicated that in most Pasteurella cases the same strain was isolated by both sampling methods. Thus the P. multocida culture obtained by nasopharyngeal swabbing represents reasonable well for the microbes in the lung. 663 First Case of Immunohistochemical Detection of Mycoplasma bovis in Natural Cases of Calf Pneumonia in Hungary L. Szeredi , T. Labossa , M. Tenk , V. Pálfi , D. Rigó , T. Molnár 1 Central Agricultural Office, Budapest, Hungary 2 Veterinary Practitioner, Lajoskomárom, Hungary 3 CEVA, Budapest, Hungary

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Objectives of the study: Bovine respiratory disease complex (BRDC) is a major cause of morbidity, mortality and economic losses in cattle production systems. The cause of this disease is rather complex involving both managing factors and infectious agents (bacteria and viruses). In our study, we demonstrate the primary role of Mycoplasma (M.) bovis in a severe respiratory disease in feedlot cattle. Materials and methods: Five feedlot cattle died after a few days of illness without any specific symptom in a herd. Out of these 2 were treated with antibiotics whereas 3 suddenly died without any treatment. All the animals were imported from Poland, vaccinated previously against infectious bovine rhinotracheitis (IBR) and bovine respiratory syntitial virus (BRSV). After gross pathological investigation swabs were collected for bacterial culture. From the lungs, mycoplasma culture was also made in 1 case and virus isolation (VI) was attempted in 4 cases. Tissue samples were collected from the lungs and occasionally also from the trachea and the mediastinal lymph nodes for histological examination. Immunohistochemical (IHC) method was used to detect viruses (bovine herpesvirus 1 (BHV-1), BRSV, bovine respiratory coronavirus, influenza A virus) and bacteria (M. bovis and Chlamydiaceae) in formalin-fixed paraffin-embedded tissue samples. Results: The cranioventral part or the whole lungs were affected by dark discoloration in all cases. Histologically, fibrinopurulent pleuropneumonia, suppurative bronchiolitis and focal to coalescing coagulative necrosis were observed in the lungs in all cases. M. bovis was detected in large numbers in the lung tissue samples with IHC, mostly in the area of bronchiolitis and coagulative necrosis, and in smaller number in the area of the fibrinopurulent inflammation. M. bovis was isolated from one of the cases. Other pathogens were detected only infrequently: parainfluenza 3 virus (1 case with VI), RSV (1 case with IHC), Pasteurella multocida (2 cases with culture), Mannheimia haemolytica (1 case with culture). Conclusions: M. bovis was the most important pathogenic agent that induced a severe respiratory disease in the feedlot cattle examined in this study. Other bacteria and viruses were detected only in a few cases. In spite of the sudden death in 3 cases, the lung lesions were not acute in any of the cases. 664 Short-duration Antimicrobial Treatment of Bovine Respiratory Disease with Enrofloxacin, Florfenicol and Cefquinome R. Froyman , C. Boda , C. Rizet , A. Valognes , N. Brunner , 2 3 P. Liege , H. Navetat 1 Bayer HealthCare, Animal Health Division, 51368 Leverkusen, Germany 2 Anisteme Biosciences, Animal Health Division, 34400 SaintChristol, France 3 Centre d'Information Veterinaire Clinique, 03130 Le Donjon, France 4 LDV03, 03017 Moulins, France Mannheimia haemolytica (Mh) is the most prevalent causative bacterium in bovine respiratory disease (BRD) but Mycoplasma bovis

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(Mb) as co-infecting agent is becoming increasingly important. In spite of this, the possible involvement of Mb is mostly overlooked whilst choosing the appropriate antimicrobial treatment of BRD-affected cattle. This trial, applying short-duration treatments, compared compounds (florfenicol [FFC], cefquinome [CFQ]) that are specifically directed against Mh, with enrofloxacin (ENR) that targets both Mh and Mb. Following Mb and subsequent Mh challenge, performed 4 days apart, groups of young calves (n=10/group) were treated 3 h later with ENR (Baytril Max 10%, 7.5 mg/kg, one SC injection), FFC (Nuflor 30%, 20 mg/kg, 2 IM injections 48 h apart) or CFQ (Cobactan LA 7.5%, 2.5 mg/kg, 2 SC injections 48 h apart). All treated calves survived the study but 2/10 untreated control calves died on day (d) 5 after Mh challenge. Whereas the surviving untreated controls continued to have fever or high body temperature until the end of the study (d7 after Mh challenge), treatment with ENR and FFC controlled fever within 15 h. It took 72 h to bring down body temperature to baseline level in calves treated with CFQ. None of the calves treated with ENR, FFC or CFQ showed respiratory disease signs or morbidity compared with 4 control calves in which such symptoms persisted until the study end. At necropsy pneumonia lesions were seen in all control calves and in 2/10, 4/10 and 8/10 calves treated with ENR, FFC and CFQ, resp. The median pulmonary lesion scores (scale 0-4) were 1.56, 0, 0, and 0.44 in untreated, ENR, FFC and CFQ groups, resp. Mean Mh counts in lung tissue of ENR and FFC treated calves were very low (<100 cfu) 3.9 and were lower than in CFQ treated calves (10 cfu, P= 0.002); the mean Mh count of the control group did not differ from the CFQ group 5.1 (10 cfu, P= 0.21). Lungs of control calves were heavily Mb infected 9.0 (mean count 10 cfu). In 5/10 and 3/10 lungs of ENR and FFC treated calves Mb counts were very low (<100 cfu) and mean counts in these 3.4 5.1 groups amounted to 10 and 10 cfu, resp. Short-duration treatments with ENR (1 injection) and FFC (2 injections) cured BRD better than CFQ (2 injections). The fewest calves with pneumonia and the best inhibition of Mb multiplication were seen in ENR treated calves. 665 The Seasonal Changes of Hemogram with Tick-borne Pathogens in Cattle in Jeju Island, Korea YH. Li , SS. Lee , TY. Kang , J. Park , JS. Chae 1 Chonbuk National University, College of Veterinary Medicine, Jeonju, Korea, South 2 National Institute of Subtropical Agriculture, Livestock Division, Jeju, Korea, South 3 Seoul National University, College of Veterinary Medicine, Seoul, Korea, South The most common tick species found on the cattle ranches in Jeju Island, Korea is Hamaphysalis longicornis. Ticks transmitted hemoparasites and/or rickettsial pathogens affect the patterns of blood composition in the grazing cattle. Because these changes are seasonal, during the spring to late autumn we analyzed the complete blood profiles of 228 grazing Korean indigenous cattle (KIC) and 58 Holstein cattle ranches in Jeju Island. The complete blood profiles were sorted into anemic, leukocytic, and/or normal groups according to the results of red and white blood cell panels. The high percentages of Holstein cattle examined in the month of July (100%), September (65%) and November (40%) showed anemia. However, the numbers of KIC showing anemia were 62% in May, 35% in July, 5% in September, and 2% in November. DNA extracts from all of the blood samples were tested, using PCR, for Theileria buffeli/orientalis/sergenti group, Anaplasma phagocytophilum and Ehrlichia chaffeensis. PCR screening showed that 98% and 93% of Holstein cattle and 76% and 85% of KIC tested positive for T. buffeli/orientalis/sergenti group and A. phagocytophilum, respectively. One KIC tested PCR positive for E. chaffeensis in one of KIC. Mixed infections with more than one tick-borne pathogen were common. These results provide the PCR evidence for T. buffeli/orientalis/sergenti group, A. phagocytophilum, and E. chaffeensis infection in KIC and Holstein cattle in Jeju Island, Korea. According to the results, anemia in grazing cattle is associated with ticks and/or tick-borne pathogens. Key words: cattle, anemia, tick-borne pathogens, hemoparasite, Rickettsia 666 Frequency of Toxoplasma gondii Infection in Sheep from a Tropical Zone of Mexico and Analysis of the Humoral Response Changes in Ten Months H. Caballero-Ortega , H. Quiroz-Romero , S. Olazarán-Jenkins , 1 D. Correa

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Instituto Nacional de Pediatría, Laboratorio de Inmunología Experimental, México DF, Mexico 1 Facultad de Medicina Veterinaria y Zootecnia-UNAM, Departamento de Parasitologia, México DF, Mexico 3 Centro Experimental Pecuario del Estado de Puebla (CIPEP)INIFAP, Hueytamalco, Puebla, Mexico In this study an indirect ELISA and an immunoblot were standardized to detect anti-T. gondii IgG antibodies in sheep, and to compare them to a commercial ELISA, used as reference test. Sensitivity was 92.0 and 96.0%, and specificity 88.0 and 75.0%, respectively. Then, the serum samples of 103 sheep of a ranch located in the Eastern tropical region of Mexico were analyzed. A frequency ranging from 77 to 84% was observed, depending on the assay used. At least eight bands ranging from 10 to 250 kDa were detected in the positive sera by immunoblotting, with a heterogeneous pattern among animals. Fifty six of the original sheep were sampled ten months later and tested again. Six animals became negative in the second sample, while one case was negative at the first time and strongly positive ten months later. Considering this a new case, we calculated an incidence rate of 2.1%. IgG avidity ELISA was performed to 36 positive sample pairs, 33 being of highavidity at both times; slight increases in avidity were found in two samples and conservation of low-avidity was observed in one sheep. Higher prevalence rates of toxoplasmosis in moist warm compared to cold atmosphere is attributed to the long viability of T. gondii oocysts. This may explain the high T. gondii frequency in this region, which apparently has favorable climatic conditions for the transmission of this protozoan, besides the presence of both domestic and wild cats. Key words: frequency, immunoassay, Mexico, sheep, Toxoplasma gondii 667 Serological Prevalence of Encephalitozoon intestinalis Antibodies in Dairy Cows in Eastern Slovakia V. Hisira , G. Kovac , V. Petrovic , Z. Farkasova , F. Novotny , 3 3 3 A. Valencakova , P. Balent , L. Luptakova 1 Universitiy for Veterinary Medicine, Department of Cattle and Sheep Diseases, Kosice, Slovakia 2 University of Veterinary Medicine, Animal Reproduction, Kosice, Slovakia 3 University of Veterinary Medicine, Biology, Kosice, Slovakia In our study, we used immunological methods to detect antimicrosporidial antibodies in blood serum from dairy cows. We examinated the prevalence of antibodies against E. intestinalis in the blood serum of cows from four farms in Eastern Slovakia by ELISA method. Spontaneus E. intestinalis infections have been documented in human population, cattle, pigs, dogs, goats, and donkeys. E. intestinalis causes enteritis with diarrhoea, weight loss, and malabsorption dissemination to ocular, genitourinary and respiratory tracts. As a positive, we evaluated sera, which reacted in a titre 1:200 and the average absorbance of sera was minimum of 2.1 times the absorbance of negative control. The presence of specific anti-E. intestinalis antibodies was detected in 9 (13,8%) from 65 examinated sera. Serological positivity to E. intestinalis was detected in 2 (10%) from 20 cows on the first farm, in 1 (11,1%) from 9 cows on the second farm, in 6 (28,6%) from 21 cows on the third farm and in the last farm we found no positive cows. The results of the present study demonstrated that the presence of E. intestinalis infection in cows is manifested by increase in antibodies against the pathogens. In next experiments we plan to study the presence of E. intestinalis spores in biological samples (faeces and urine) from serological positive domestic animals (cattle and pigs) with polymerase chain reaction with the use of species specific primer pairs. These biological materials can tend as the potentional sources of infection for other animals and humans on farms although the possibility did not confirm that E. intestinalis infection is zoonotic in origin. Key words: microsporidia, Encephalitozoon intestinalis, dairy cow, ELISA 668 A Combined Approach to the Control of Bovine Viral Diarrhoea Virus (BVDV) Infection under Field Conditions G. Valla , C. Piancastelli , M. Bussacchini , J. Munoz Bielsa , 3 2 B. Makoschey , S. Cavirani 1 Intervet-Schering Plough, Intervet Italy, Peschiera Borromeo, Italy

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University of Parma, Department of Animal Health, Parma, Italy Intervet-Schering Plough, Int. Marketing, Boxmeer, The Netherlands

Objectives: The control of the Bovine Viral Diarrhea Virus infection includes the identification and removal of persistently infected (PI) animals and/or vaccination. Some inactivated vaccines, as Bovilis® BVD (Intervet International), have proven foetal protection, preventing the birth of new PI animals. This study had two objectives: (i) to monitor transplacental infection and (ii) to assess the efficacy of vaccination under field conditions. Materials and Methods: Six farms with a total of 1332 animals were included in the study. Three of them were not vaccinated, one was previously vaccinated with a modified live ts BVD vaccine and two with an inactivated BVD vaccine applied in a vaccination program for which foetal protection was not demonstrated. At the beginning of the study 76.5% of the animals were positive for BVDV NS2-3 antibodies. For the identification of PI animals and newborn calves two different methods were used: an antigen ELISA and PCR. All animals included in the study were vaccinated with a commercial inactivated BVD vaccine (Bovilis BVD) administered as two doses, 28 days apart (priming) and a booster every 6 months. The monitoring for detection of PI in newborns continued for 1 to 3 years. Results: At the beginning of the study, a total of 19 PI animals, was detected in 4 of the 6 farms. In one of these farms the PI animals were not immediately removed and remained in close contact with the other animals for about 10 months. The monitoring of the newborns showed that one new PI animal (0.12% of the 831 tested) was identified during the study. The mother of this calf, had been tested negative at the beginning of the study with ELISA antigen test, but was later on identified as PI when retested by PCR after the birth of her PI calf. This newborn PI can therefore not be related to a vaccine failure. Moreover, the PI mother was for at least 9 months in close contact with the other animals. Conclusions: The combined approach based on the identification and removal of PI and vaccination is an effective method of BVD infection control and the vaccination with the inactivated vaccine used in the study induced a high level of foetal protection even in presence of PI animals. 669 Influence of BVDV Vaccination on Reproductive Performances in Field Conditions G. Valla , F. Cammi , J. Munoz Bielsa , B. Makoschey , F. Petrera , 1 M. Bussacchini 1 Intervet-Schering Plough, Intervet Italy, Peschiera Borromeo, Italy 2 Azienda Sperimentale Vittorio Tadini, Intervet Italy, Gariga di Podenzano (PC), Italy 3 Intervet-Schering Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: BVDV infection can cause transplacental infection, heat returns and abortions. When the infection occurs during oestrus it can induce ovarian lesions and hormonal disorders, and therefore reducing the pregnancy rate. A study was set up to evaluate the benefits of vaccination with a commercial inactivated BVD vaccine (Bovilis BVD, Intervet) on the reproductive performance. Materials and Methods: The trial was conducted in a dairy farm of about 120 lactating cows (Azienda Sperimentale Vittorio Tadini). A number of parameters, including days open (DO value), Average Service Per Pregnancy (ASPP), Heat Detection Rate for Repeat Insemination (POSTHDR), Conception Rate (CR) and Pregnancy Rate (PR), were recorded during 9 years (1998-2006) using a specific software (Fertility Factor Farmer, Italy). Three different vaccination regimens were implemented during that period. Between May 1998 and April 2001 (Period A) all the animals were vaccinated, every 6 months, with an IBR marker modified live vaccine; in period B (May 2001 - April 2003), the farm was unvaccinated. From May 2003 to April 2006 (C) all the animals were vaccinated concurrently, every six months, with a modified live IBR marker vaccine and an inactivated BVD vaccine. Before starting BVD vaccination in April 2003, the herd was tested to identify PI animals. Results: Three PI were detected but not immediately removed. They remained in close contact with the other animals for about ten months. Despite the fact that no new PI animals were detected between 2003 and 2006, BVDV circulation was noticed through the seroconversion against NS2-3 antibodies. During the same period the IBR percentage of IBR gE positive animals decreased from 23% to 8%. The DO value was 152 days in period A, 154 in B and 146 in period C. The ASPP decreased from 1.87 in A and 1.55 in B to 1.52 in C. The POSTHDR

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increased from 40.0% in A and 37.4% in B to 45.1% in C. The CR and the PR were 46.1% and 14.3 in A, 51.3% and 13.3 in B and 53.1% and 14.8 in C, respectively. The profitability index, calculated for a lactation of 305 days and an average milk production of 9200 litres, was higher in the period C showing a difference in the revenue of 8 /cow/year versus period A, and 25 versus the unvaccinated period B. Conclusion: The introduction of BVD vaccination improved reproductive such as DO value and POSTHDR, resulting in an economical benefit for the herd. 670 Frequency of Isolation and Combitation of Pathogens Involved in Bovine Respiratory Disease in France: a Retrospective Analysis F. Payot , E. LeDrean , M. Bonnier , B. Makoschey , C. Dore 1 Intervet-Schering Plough, Farm and Production Animals Business Unit, Angers, France 1 LDA35, Rennes, France 3 Intervet-Schering Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: Bovine respiratory disease (BRD) is generally recognized as a multifactorial disease. Primary causative pathogens, such as the bovine respiratory syncytial virus (BRSV), and secondary pathogens (enhancing the impact of primary pathogens), such as bovine parainfluenza type virus (PI3), are commonly discerned. We have studied the frequency of isolation of pathogens from calves with BRD and the frequency of combination of these pathogens (i.e. found alone or combined). Material and Methods: The analysis was performed on data from the Departmental Veterinary Laboratory 35 (Rennes, France) from November 2006 to October 2007. The samples originated from recently infected animals with BRD, mostly coming from the western part of France. The presence of the following pathogens was investigated: bacteria: Pasteurellaceae (Mannheimia haemolytica, Pasteurella multocida and Histophilus somni), Arcanobacter pyogenes and Mycoplasma bovis, viruses: Bovine viral diarrhoea virus (BVDV), BRSV, bovine herpesvirus type 1 (BoHV-1) and PI3. Only samples that were tested for all these pathogens were included in this retrospective analysis. Results: 180 samples were included. They were obtained from respectively 65%, 8% and 27% of transtracheal lavage, nasal swabs and lung tissue at necropsy. 52 samples were found negative and 128 were positive for at least one pathogen. A combination of two or more pathogens was found in 56% (72) of cases whereas the presence of a single pathogen was observed in 44% (56) of cases. Globally, Pasteurellaceae, BRSV, M. bovis were isolated respectively from 73%, 35% and 47% of samples. The other pathogens were detected in less than 15% of samples. Focused on single isolated pathogens (n=56), the following prevalences were observed: Pasteurellaceae 50%, BRSV 25%, M. bovis 17%, A. pyogenes 4%, PI3V 4%. The frequency of combination of Pasteurellaceae or BRSV with other pathogens was 67%-70%. In contrast, the frequency of combination of PI3V, M. bovis or A. pyogenes with other pathogens was higher: 83% for each. BVDV and BoHV-1 were always combined with other pathogens. Conclusion: This retrospective analysis emphasized the multifactorial aetiology of BRD. Pasteurellaceae and BRSV were frequently isolated combined or as single pathogens. M. bovis were also frequently isolated but mostly associated with other pathogens as observed for A. pyogenes and PI3. 671 Comparison of Different Vaccination Protocols against Bovine Respiratory Disease Complex in Fattening Units C. Tresse , K. Risson , F. Bendailh , B. Makoschey , L. Oliviero 1 Ter elevage, Villedieu la Blouere, France 2 Ecole Vétérinaire, Toulouse, France 3 Intervet-Schering Plough, Int. Marketing, Boxmeer, The Netherlands 4 Intervet-Schering Plough, Farm and Production Animals Business Unit, Angers, France Objectives: Respiratory disease outbreaks are common after mixing and transportation of calves. In a field trial the efficacy of vaccination of calves in the breeding farm prior to transport was compared to vaccination after transport and mixing with animals from different origins. Materials and Methods: In the study, 161 calves at the age of 7 to 8 months were included. They originated from 15 farms and were first sent to 2 trading centers for approximately 12 hours and from there

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distributed over 13 different fattening farms. In the farms of origin, the calves of group EV (early vaccination) were either vaccinated twice at 1 month interval with a trivalent vaccine (bovine respiratory syncytial virus (BRSV), bovine parainfluenza type 1 (PI3), Mannheimia haemolytica), or twice at 1 month interval with a bivalent vaccine (BRSV, bovine viral diarrhoea virus (BVDV)). The vaccination of group EV was completed 15 days before transport to the trading center. A second group LV (late vaccination), of calves, that arrived at the trading centers at the same time as the EV group, was left unvaccinated until the arrival at the trading center. The animals of the group LV were vaccinated with the bivalent vaccine at the trading center (first dose) and the fattening farm (second dose, 1 month later). The two groups were raised under the same conditions in each fattening unit. The following parameters were recorded: morbidity and mortality due to respiratory disease, number of treatments due to respiratory disease during the first two months at the fattening unit. The observations were performed by persons that were not aware of the grouping. Results: The results showed a significant decrease in morbidity (5% versus 12.7%) and number of individual treatments (5.4% versus 18%) for the animals that received their vaccination at the breeding farm (group EV) as compared to the calves that were vaccinated at the trading center and the fattening unit (group LV). Conclusions: Timing of vaccination has a significant effect on the prevention of respiratory disease under the conditions described above. 672 The Use of ELISA for the Diagnosis of Goat Tuberculosis in Brazil C. Marassi , C. Almeida , S. Pinheiro S. Vasconcellos , 1 W. Lilenbaum 1 Universidade Federal Fluminense, Microbiology, Niterói - RJ Brasil, Brazil 2 Universidade de Sao Paulo, Preventive Medicine and Public Health, Sao Paulo, Brazil Bovine tuberculosis remains an endemic infection in several countries, despite several efforts to control the disease. In the past, it was believed that goats were naturally resistant to tuberculosis; however it is now well established that M. bovis and M. caprae could be responsible for this disease in goats. After one clinical case that was presented to the veterinary service of the University of Sao Paulo, Brazil, a complete screening by intradermal tuberculin test was performed in the herd. The goat herd was composed by 500 animals and 83 (16.6%) of them showed to be reactive to the comparative double cervical intradermal test. Four months after the test, all the reactive animals were slaughtered and 33 of them were randomly chosen for histopathological, serological and bacteriological procedures. Histopathology (Ziehl-Neelsen and Hematoxyllin-eosin) evidenced at least one characteristic lesion of tuberculosis in each animal, with typical granulommas where BAAR could be observed. Bacteriology (inoculation in Stonebrink-Lesslie media) was positive for M. bovis in 25 samples (75.8%), therefore confirming the etiology of the outbreak. Sera of these 33 animals were tested in an ELISA using the recombinant M. bovis protein MPB70 as capture antigens. From those, 31 were reactive to the test, with high ODs results, considering a cut-off point established by ROC curve analyzing results of M. bovis culture from tuberculous cattle (cut-off = 0.06; mean= 0.55; range: 0.157-1.357). These preliminary results suggested that MPB70-ELISA can be considered as a reliable tool to diagnose tuberculosis in goat herds, since this assay was capable to demonstrate the disease in 93.9% of the animals here examined. Key words: tuberculosis, goats, ELISA, diagnosis, M. bovis 673 Surveillance of Bluetongue Virus in Domestic Ruminants in Territory of the Slovakia- Draft of Research Program J. Bires, A. Kocisova, M. Mojzis, J. Pliesovsky, V. Petrovic University of Veterinary Medicine, Department of Cattle and Sheep Diseases, Kosice, Slovakia The research project is focused on acquirement of primary information about occurrence of health and economically serious orbivirus transmissive disease - bluetongue in cattle, sheep and goats in Slovakia. In content of project there will be the research of occurrence of vector Culicoides spp. and their position towards transmission of catarrhal fever on animals (entomological study), monitoring of potential occurrence of disease in cattle, sheep and goats (clinical and laboratory examinations) and verification and introduction virological diagnosis

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of aetiological factor of bluetongue. Diagnostic procedures of bluetongue on the level of orbivirus, vector and animals are in accordance with Manual of diagnostic tests and vaccines for terrestrial animals (OIE) and legislative issues of European Commission. The intensity of surveillance bluetongue will be realised on selected areas of Slovakia during whole year on all the levels of observation depending on climatic conditions and persistency of vectors in nature. Solution of project expects collaboration of experts from veterinary university, state veterinary institute, state veterinary administration, veterinary institute, veterinary surgeon and farmers. Key words: bluetongue, surveillance, domestic ruminants 674 Prevalence of BVDV Infection in Large and Small Cattle Herds in Southern Backa and SREM District in Serbia T. Petrovic , S. Lazic , D. Bugarski , B. Djuricic 1 Scientific Veterinary Institute Novi Sad, Virology, Novi Sad, Serbia 2 Scientific Veterinary Institute Novi Sad, Department of Epizootiology and Health Protection of Ruminants, Novi Sad, Serbia 3 Faculty of Veterinary Medicine Belgrade, Department for Infectious Diseases- Epidemiology, Novi Sad, Serbia In order to determine the prevalence of BVDV infection in small and large cattle herds in Southern Backa (S. Backa) and Srem district, the examination was carried out on presence of VN antibodies in blood sera in breeding cattle that were older than 6 months during 2004. The examined 3457 samples were from small cattle herds in S. Ba_ka (9 communities) and 4120 samples in Srem district (8 communities). The examination included animals from all the settlements (156) in this area. The examined 2721 samples were from 18 large cattle herds from S. Backa, and 298 samples were from 4 large cattle herds from Srem district. The examination included animals from all large herds (22) in this area. Virus neutralization test (VN) was performed according to standard procedure with small modifications. In the test NADL strain of BVD virus and cell culture MDBK were used. The sera were examined in double solution from 1:2 to 1:512.The presence of VN antibodies was discovered in 1833 (24.19%) samples from small cattle herds. Out of this number in S. Backa district there were 1082 (31.30%), and in Srem district there were 751 (18.23%) seropositive animals. The largest number of seropositive animals in S. Backa district was determined in Titel municipality (68.07%) and the smallest in Backi Petrovac municipality (6.63%). In Srem district the highest percent of seropositive animals was found in Beocin municipality (34.16%) and the lowest in Irig municipality (7.62%). The presence of VN antibodies was discovered in 1667 (55.22%) samples from large cattle herds. Out of this number in S. Backa district large herds there were 1500 (55.13%), and in Srem district large herds there were 167 (56.04%) seropositive animals. The prevalence of seropositive animals in big cattle herds from S. Backa district ranged from 0% (5 large herds) to 96.61%. High prevalence (>70%) was determined in 7 out of 18 herds. The prevalence of seropositive animals in large cattle herds from Srem district ranged from 0% (1 big herd) to 96.96%. High prevalence (>70%) was determined in 2 out of 4 herds.The obtained results point out that BVD infection is spread on the examined area. At the same time, prevalence is not equal, what means that there are areas where it is not present, or is only sporadically present. Key words: BVDV, VN test, prevalence 675 Antimicrobial Resistance in Escherichia coli Isolated from Calves in Slovakia V. Kmet , D. Bujnakova , D. Demankova , S. Novak , R. Bobcek , 2 2 T. Szakal , M. Huska 1 Slovak Academy of Sciences, Institute of Animal Physiology, Kosice, Slovakia 2 University of Veterinary Medicine, Department of Cattle and Sheep Diseases, Kosice, Slovakia A total of 159 faecal Escherichia coli isolates obtained from calves from seven regions in Slovakia during one year were examined for their quantitative susceptibility. MIC values to 20 antimicrobial agents were determined by a colorimetric microdilution test. The underlying resistance mechanism were tentatively identified from antibiogram profiles using interpretative reading method. The most frequently found resistance was to tetracycline (76 %), streptomycin (77 %) and ampicillin (78 %). Among tested E . coli isolates also high resitance rates to neomycin (51 %), chloramphenicol (30 %), cotrimoxazole (28

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94 XXV. Jubilee World Buiatrics Congress 2008

%) and spectinomycine (25%) could be recorded. Resistance to ampicillin+sulbactam (8,1 %), ceftiofur (4,4%), apramycin (0.6 %), gentamicin (7 %) and florfenicol (10 %) was lower. All of the tested E. coli isolates were sensitive to cefquinome and colistin. Resistance to ciprofloxacin (17 %) and enrofloxacin (16,9%) was intermediate. However, overall quinolone resistance detected by means of nalidix acid was as high as 29 %. The high level fluoroquinolone resistance (CIP: 4mg/L and ENR: 16 mg/L) (18%) as well as penicillinases ((TEM1,2/SHV1)-low (51%) and (TEM1,2/SHV1)-high (25%) was detected in E. coli isolates. This is the first report of a quantitative antibiotic susceptibility testing and phenotype resistance mechanisms determination in E. coli isolated from calves in Slovakia. Key words: E. coli, minimal inhibitory concentration, interpretative reading, quinolone resistance, calves 676 Deciphering the Value of Chlamydia-Serology in Cattle J. Boettcher, A. Vossen, A. Gangl, N. Meier, G. Wittkowski Bavarian Animal Health Service, Poing, Germany Chlamydiae are often suspected in reproductive and respiratory diseases of cattle. Because of frequently used ELISA-testing, we tried to assess its value. Based on 21683 sera a seroprevalence (SerP%) of 37.0%±0.3% was determined for Bavaria (Idexx-ELISA). Positive sera were detected in 92.8%±0.9% of the farms (n=765). Average within-herd prevalence was 37.0%±1.4%. Age-related effects (n=3522): SerP% was highest in calves less than 9 month of age (mona) (48%±6%) and lowest between 21-24 mona (41%±2%)(n.s.). On 5 clinically unaffected farms swabs were analysed for Chlamydia by PCR (357 animals). Detection rates for vaginal, nasal and conjunctival swabs were 2.5%±0.8%, 2.0%+0.7% and 1.1%±0.6%, respectively. Detection rate was 22% for calves <6 months, followed by animals 6-12 mona (12%), rates were higher than in animals >12 months (p=0.014). SerP% in ELISA were 36% and 10% for Idexx and Pourquier, respectively. While SerP% (Idexx) increased from 22% in calves <6 months to a maximum of 50% in animals 13-18 mona, a decrease was observed for Pourquier (22% to 5%). 682 cows were sampled 4x (Autumn 2003, Spring, Summer and Autumn 2004). Stored sera (-20°C) were tested at one time (Idexx). Differences of two samplings were statistically analysed by McNemar-test (MedCalc). Between Aut.03/04 no significant difference explained by seroconversion was detected (P=0.33), but remarkably 145 cows showed decreased and 163 increased reactivity. Results for Aut.03/Spr.04 were similar, but significant changes were detected for Spr./Sum.2004 (7%; CI95%:3.310.5%; P=0.0002) and Sum./Aut.04 (7.8%; CI95%:4.3-10.9%; P<0.0001). Difference for Spr./Sum.04 was due to loss of reactivity, in contrast for Sum./Aut.04 it could be explained by seroconversion. In contrast and as a control fluctuation of results except for seroconversion was not observed for Q-Fever. Chlamydial infection in cattle is widespread. Calves get infected early in life. Fluctuation of ELISA-results might be explained by periods of chlamydial-reactivation and short-lived antibodies. An effect of seasons was observed. Thus even seroconversion might be missleading. Quantitative PCR might be a promising tool. Financially supported by Free State Bavaria and the Bavarian Joint Founding Scheme for the Control and Eradication of Contagious Livestock Diseases. Key words: Chlamydia, serology, cattle 677 The Efficacy of Turkish Propolis and Whitfield's Ointments in the Treatment of Ringworm in Young Cattle Y. Cam , N. Koc , S. Silici , V. Gunes , H. Buldu , A. Onmaz , 2 F. Kasap 1 University of Erciyes, Faculty of Veterinary, Internal Medicine, Kayseri, Turkey 2 University of Erciyes, Faculty of Medicine, Microbiology, Kayseri, Turkey 3 University of Erciyes, Safiye Cikrikcioglu Vocational College, Animal Sciences Group, Kayseri, Turkey This study was conducted to evaluate the effect of Turkish propolis and Whitfield's ointments used as seperate or together in the treatment of ringworm in young cattle. In this study, a total of 35 young cattle aged between 3-8 months showing clinical signs of ringworm were used. Animals were divided into four groups as group I (untreated animals), group II (animals administered with Turkish propolis ointment), group III (animals administered with Whitfield's ointment) and group IV (animals administered with Turkish propolis and Whitfield's ointments). The group

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I consisted of 5 animals, while the other groups included 10 animals each. The total period of the study was 6 weeks. The animals in group I was kept as control throughout the study. The Turkish propolis and Whitfield's ointments, each alone or together were topically applied to animals every day in the treatment groups for two weeks. Clinical findings were observed during the study. Prior to treatment and at the end of the study, hair and crust samples from the lesions were collected for mycological examinations from all of the animals. Prior to treatment, Trichophyton verrucosum was isolated from all of the animals. In the group I, the lesions of ringworm were existed throughout the study. It was found that 3 animals of the group II, 4 of animals of the group III and 9 of animals of the group IV clinically healed. At the end of the study, T. verrucosum was isolated from 7 animals in group II, from 6 animals in group III, and from 2 animals in group IV. In conclusion, Turkish propolis and Whitfield's ointments were more effective when used together then that of each alone in the treatment of ringworm in young cattle. Key words: cattle, ringworm, treatment, Turkish propolis 678 Bovine Leukemia Virus (BLV) Infection in Experimental Animals P. Dimitrov , K. Gencheva , L. Yossifova , E. Gardeva , 2 1 1 R. Valtchovski , E. Shikova , R. Russev 1 Institute of Experimental Pathology and Parasitology, Bulgarian Academy of Sciences, Sofia, Bulgaria 2 National Diagnostic and Research Veterinary Institute, National Veterinary Service, Sofia, Bulgaria Studies on BLV are important not only because of the chronic disease the virus cause in cattle, incurring considerable economical losses, but for its significant homology with virus causing T-leukemia in human. We tried to create smaller, easily manipulated model of the disease. For that reason rats and rabbits were inoculated with BLV. According to the literature they are the most susceptible laboratory species. The aim of our investigation is to obtain additional clinical and immunological data and to formulate detailed pathomorphological characteristics of the disease in these animals. Materials and methods: -Inoculation - as inoculation materials white blood cells from leukemic cows with manifested leukocytosis (to 30000) and cells from FLK culture, producing BLV (proved by EM) were used. Rabbits and rats were inoculated twofold at the age of 30 and 40 days. -Radial immunodiffusion: In 1% agarose gel, using standard protocols -Pathohistology: routine -Electron microscopy (EM): materials were fixed in 2,5% glutaraldehyde, postfixed in osmium tetroxide (1% in PBS), embedded in Epon, stained with uranil acetate Results: By radial immunodiffusion test was found that 34% of the inoculated animals react positively. Among them 60% showed increased number of white blood cells to values of 17000 to 26000 per ml. Most of BLV-infected animals developed conjunctivitis, rhinitis, enteritis, pneumonia and weight loss. Histologicaly the alterations in lungs were manifested by hyperemia of the organ and the presence of focal serous and purulent pneumonia. In the liver lymphocyte foci in the periportal spaces and sometimes fresh necrotic patches were found. Perivascular lymphocyte clusters in the kidneys were also observed. In the spleen the white pulp though spatially reduced, was comparatively rich in cells and in some cases displayed well-shaped germinal centers. Conclusions: The data obtained show that treatment of rats and rabbits with BLV-producing cells of the FLK line and with white blood cells from cows with enzootic leucosis leads to their infection. A proof of this is also the presence of anti-viral antibodies in the serum of treated animals. The clinical, hematological and histological findings reveal some kind of immunosuppressive action of the virus. Key words: BLV, retroviruses, laboratory model 679 Bovine and Chiropters Rabies Laboratorial Diagnosis: an Epidemiological Approach F. Guimaraes, L. Ullmann, L. Camossi, H. Greca, B. Menozzi, H. Langoni Universidade Estadual Paulista Júlio de Mesquita Filho, Veterinary Hygiene and Public Health, Botucatu, Brazil Rabies is one of the most important zoonosis, characterized by the severity of the symptoms and high level of fatality. It can compromise all the mammals and human beings. The dogs are the

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main infection sources to the urban rabies, while the chiropters are important in rabies transmission to the production animals. Due to the relevance of this zoonosis to Public Health some measures of control are usually implemented: vaccination of dogs and cats, laboratorial diagnosis surveys, control of the chiropters, sanitary educational programs. The purpose of this study was to refer the positive rabies diagnosis from 2006 to 2007, conducted by the Zoonosis Diagnostic Center- FMVZ/UNESP-Botucatu/SP-Brazil staff. A total of 1061 brain samples from various animal species were analyzed during this period, from different counties sent by the epidemiological surveillance groups, GVEs 15 and 16. Direct immunofluorescence technique and the biological test were performed to rabies diagnosis. During the studied period 13 (1.23%) were positive, from these positives samples 38.46% (5/13) were from bovines and 61.54% (8/13) were from chiropter. When the total of bovine and chiropters samples was considered it was verified that the rabies positivity among the bovines were higher than among chiropters and the difference was statistically significant (P = 0.0032, Fisher's Test, confidence interval of 95%). In the same period and region it was verified negative results on 300 dog and 95 cat samples. Recently, January, 2008, an outbreak occurred in two cattle farms. In one of them, 10 percent of 340 bovines showed signs of chiropters bites, five presented rabies symptoms and died. In the other, 10 bovines from a total of 149, showed chiropters bites signs, six of them presented the classical rabies symptoms and died. In both herds anti-rabies vaccination was not used. The rabies diagnosis from these animals was confirmed by the laboratory tests. The results of this study demonstrated that the rural rabies transmitted by haematophagus chiropters still being a problem in Brazil. As a conclusion, it can be inferred that in spite of the adopted measures to control the urban rabies were effective in the studied region the problem of rural rabies was still prevalent. It was closely linked to the chiropters as rabies Lyssavirus reservoir, therefore, they represent a real risk to the persistence of the rabies in bovines. Key words: rabies, zoonosis, public health, chiropter, bovine 680 Prevalence and Risk Factors for Campylobacter sp. Colonization in Cattle in Poland M. Bednarski, A. Wieliczko Wroclaw University of Environmental and Life Sciences, Department of Epizootiology and Veterinary Administration with Clinic of Infectious Disease, Wroclaw, Poland In developed countries campylobacteriosis is now one of the most frequent reasons of foodborne diseases and usually it is associated with gastroenteritis. Animals and animal food products are the main source of the infection of Campylobacter for human. Because of the potential linkage between Campylobacter spp. harbored by cattle and human disease, in this study we investigated the prevalence and risk factors for colonization Campylobacter spp. in cattle. Objectives of study: The objectives of the study were to estimate prevalence and risk factors for colonization Campylobacter spp. in cattle in Poland. Materials and Methods: The study was performed on 896 cows from 84 farms located in South-Western Poland. Fecal samples were checked in two seasons: summer and winter. All fecal samples were analyzed by microbiological and PCR methods. Isolated Campylobacter sp. strains were characterized by fenotypic and genotypic methods. Prevalence data analysis was performed using Pearson chi-square test. Results: From all fecal samples tested by microbiological method 151 (16,85%) were positive for Campylobacter sp. C. jejuni was the most commonly isolated species with 143 positive fecal samples (94,7% of all isolated Campylobacter strains) whereas C. coli were present only in 7 samples (4,6% of isolated Campylobacter strains). Using PCR method 29,4% of samples were Campylobacter positive. Campylobacter spp. prevalence in cattle were correlated with: extensive type of herds, outdoor and grazing cattle, cows grouped by-products, type of cows housing and husbandry, TMR (total mixed ration), not chlorinated drinking water, presence of poultry and wild birds on farm. Conclusions: The study showed relatively high prevalence of Campylobacter spp. in cows. Therefore, Campylobacter spp. have to be regarded as an important hazard of zoonotic disease in cow farms. In most cases this pathogen was isolated in fecal samples taken from clinically normal animals. Key words: Campylobacter, foodborne diseases, risk factors

681 Antimicrobial Resistance of Thermophilic Campylobacter Spp. from Cattle Farms in Poland M. Bednarski, A. Wieliczko Wroclaw University of Environmental and Life Sciences, Department of Epizootiology and Veterinary Administration with Clinic of Infectious Disease, Wroclaw, Poland Campylobacter species are among the most frequently identified bacterial causes of human gastroenteritis. Because Campylobacter spp. harbored by cattle can be transmitted to human, in this study we investigated antimicrobial resistance of thermophilic Campylobacter isolated from cows. Objectives of study: The objectives of the study were to estimate antibiotic resistance of thermophilic Campylobacter isolated from cows in Poland. Materials and Methods: Our study included 150 strains of thermophilic Campylobacter (143 strains of C. jejuni and 7 strains of C. coli) collected from cows in South-Western Poland. Isolated strains of Campylobacter sp. were characterized by fenotypic and genotypic methods. The MICs to ciprofloxacin, erythromycin, gentamicin, and tetracycline hydrochloride were determined using the agar dilution methodology according to the CLSI. C. jejuni ATCC 33560 and C. coli ATCC 33559 were used as a control in susceptibility testing and also as a growth control strains. The MIC breakpoint used for the resistance to antibiotics was recommended by the National Committee for Clinical Laboratory Standards (NCCLS) for non-Enterobacteriaceae. Results: All strains of C. coli were susceptible to all four of the antimicrobial drugs studied. The most frequently detected resistance of C. jejuni was to ciprofloxacin (26 strains 18,2%). Resistance to tetracycline was observed in 5 (3,5%). All strains of C. jejuni were susceptible to erythromycin and gentamicin. Conclusions: Ciprofloxacin, erythromycin, gentamicin, and tetracycline were chosen for antimicrobial susceptibility testing in this study, because of their importance as a front-line therapeutic drug in humans and animals and the relatively frequent occurrence of resistance to these drugs. In conclusion, we demonstrated relatively high resistance to ciprofloxacin of C. jejuni. These data support the concept of continuous selection pressure for the emergence and spread of fluoroquinolone resistance in many parts of the world and in Poland as well. Key words: antibiotic resistance, Campylobacter, foodborne diseases 682 Frequencies of PRNP Gene Polymorphisms in Cattle in Vietnam and Thailand for Potential Association with BSE Y. Muramatsu , M. Horiuchi , T. Ogawa , K. Suzuki , 5 6 7 7 M. Kanameda , T. Hanh , C. Tongchai , I. Kuramontong , 7 1 P. Kriengsak , Y. Tamura 1 Rakuno Gakuen University, School of Veterinary Medicine, Ebetsu, Japan 2 Hokkaido University, School of Veterinary Medicine, Sapporo, Japan 3 Nippon Veterinary and Life Science University, Faculty of Veterinary Science, Musashino, Japan 4 Universidad National de La Plata, Facultad Ciencias Veterinarias, La Plata, Argentina 5 Japan International Cooperation Agency, Tokyo, Japan 6 National Institute of Veterinary Research, Department of Veterinary Hygiene, Hanoi, Vietnam 7 Chulalongkorn University, Department of Veterinary Microbiology, Bangkok, Thailand Objectives of study: Since 2004, significant associations between bovine spongiform encephalopathy (BSE) susceptibility in cattle and frequencies of insertion/deletion (ins/del; indel) polymorphisms within the bovine prion protein gene (PRNP) have been reported. To know the genetic background in the PRNP gene in cattle in Asian countries, we investigated the frequencies of indel polymorphisms within two variable sites, a 23-bp indel polymorphism in the promoter region (23indel) and a 12-bp indel polymorphism in intron 1 region (12indel), in the bovine PRNP in cattle in Vietnam and Thailand. Materials and Methods: A genomic DNA was extracted from 278 serum samples of Vietnamese cattle or 113 spleen samples of Thai cattle with a commercial kit in accordance with the manufacturer's instructions (QIAamp DNA Mini Kit, QIAGEN). The extracted DNA was subjected to PCR methods for detection of 23indel and 12indel polymorphisms in the bovine PRNP. Both PCR methods were performed as previously

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96 XXV. Jubilee World Buiatrics Congress 2008

described with slight modification (Sander et al., 2004). Briefly, a total of 50µl volume of reaction mixture containing 10 to 20µl volume of the extracted DNA was used for the first stages of both PCR methods. In case that no amplified DNA fragment was detected, 10µl each volume of the first PCR product was used as a template of the second PCR. The second PCR was performed in the same manner as the first PCR. Results: We could detect polymorphic DNA fragments in both of the 23indel site and the 12indel site of 206 Vietnamese cattle and 102 Thai cattle. As for the 23indel in Vietnamese cattle, the frequency distributions of the del polymorphisms, which are thought to be associated with BSE susceptibility, were significantly high. However, no significant difference was shown in the frequencies of both allelic and genotypic polymorphisms in the 23indel site of Thai cattle. On the other hand, the frequencies of del allelic and del/del genotypic polymorphisms in the 12indel site, which have been reported to confer BSE susceptibility, were significantly low in cattle of both countries. In particular, the frequencies of del polymorphisms in the 12indel site were extremely low in Thai cattle. Conclusion: We have provided evidence that cattle in Vietnam and Thailand have a unique genetic background in the PRNP gene in comparison with cattle or sires previously reported in other countries. Key words: bovine PRNP, BSE, Vietnam, Thailand, indel polymorphisms 683 Bluetongue Disease in Cattle in Northrhine-Westfalia Clinical Symptoms and Economic Relevance P. Heimberg, W. Adams, W. Hollberg, M. Holsteg, J. Winkelmann Chamber of Agriculture NRW, Cattle Health Service, Münster, Germany Background: In August 2006 the first outbreak of Bluetongue Disease (BT) in Middle Europe by Serotype 8 was confirmed: Most of the infected farms in Germany where located in Northrhine-Westfalia (NRW, about 800 farms). In 2007 we observed a rapid spread of BT virus in NRW, ending up with a total amount of nearly 10.000 positive tested farms, dairy cattle as well as sheep farms. The economic losses in sheep are quite easy to calculate because most of the infected animals died or have been culled (assumed mortality in NRW of about 2%).The losses are compensated by the compulsory insurance for epizootic diseases and the government. In contrast dairy cattle did not show so many fatalities, but we have seen many symptoms with economic importance on the affected dairy farms. A questionnaire was sent to cattle farmers organized by the chamber of agriculture NRW to find out which BT symptoms are most important for the farmers. This was done to show if vaccination against BT is not only necessary for sheep but also for cattle to reduce casualities. Questionnaire: During the last week of November and first week of December 2007 we got answers from 565 farms. 527 of those were dairy farms. While 429 dairy farms where tested positive for BT Serotype 8, there were 98 which had no positive test result for BT. The farmers who answered to our questionnaire had in total 93.248 animals in their holdings. Results: We found out that two of three cattle farmers had direct losses. Answers resulted to an average morbidity of 10,5 %; the total amount of 725 losses lead to a letalitiy of 7,4 % and a mortality of 0,8 % by BT Serotype 8 infection in NRW. Abortion was detected up to first week of December 2007 concerned about 3 % of all dairy cows of BT-positive farms. The third important reason for economic losses by BT was seen in reduced amount of milk. Additionally 30 % of the farmers reported a significant increase in costs for veterinary treatments since summer 2007. Conclusions: Although fatalities are not that much seen in cattle compared to sheep in NRW high remontation costs caused by fatalities and abortion, together with reduced milk delivery and increasing costs for veterinary treatment implemented that vaccination for cattle is as necessary as for sheep from point of economic view. At the end of the financial year 2007/08 in June 2008, casualities will be calculated for the average affected farm in Euro per cow. Key words: bluetongue, serotype 8, cattle, symptoms, economic relevance 684 Comparative Field Clinical Efficacy of Florfenicol+Flunixine Meglumine in the Treatment of Bovine Respiratory Disease PR. Cloet , O. Roy , F. Pillet , V. De Haas , C. Maisonneuve 1 Schering Plough Vétérinaire, Levallois Perret, France

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CEBIPHAR, CRO, Fondettes, France Schering Plough Animal Health, Levallois Perret, France Schering Plough Animal Health, Summit, United States

The efficacy and safety of Resflor® (florfenicol + flunixin meglumine) was evaluated in a GCP field trial as treatment of bovine respiratory disease in comparison with Draxxin® (tulathromycin). 233 animals (260.5 98.21 kg) from 8 different farms were included with signs of bovine respiratory disease and allocated to 43 pens: 114 calves (21 pens) were given Resflor® at a dose of 2 ml/15 kg and 119 calves (22 pens) were given Draxxin® at a dose of 1 ml/40 kg. Treatments were administered on D0 by one subcutaneous injection. The nasopharynx was swabbed for bacteriological examination on D0 and upon failure. All animals included on D0 exhibited abnormal respiration, rectal temperature 40.3 °C, and depression. Rectal temperatures were measured, clinical signs scored, and injection sites observed on D0, D0+6h, and from D1 to D4. From D5 to D14, the animals were observed for clinical signs indicative of progressive disease (depression, abnormal respiration). The main pathogens isolated on D0 were Pasteurella multocida and Mannheimia haemolytica (6 clinical sites out of 8); Mycoplasma bovis and Histophilus somni were isolated in 2 farms. Rectal temperatures were lowest in the Resflor® group from D0+6h to D4. The mean rectal temperature was statistically lowest on D1 in the Resflor® group (P=0.0196, Mixed Linear model). Depression scores were statistically lowest in the Resflor® group 6 hours after treatment, on D4, and on D8. Respiratory pattern was not different throughout the study between both groups. Success rate was highest in the Resflor® group at each time point (from D5 to D14). Animals in both groups displayed injection site reactions throughout the study. One animal in the Resflor® group died on D4 after having been removed on D3 (worsening); gross pathology revealed lesions of severe acute bronchopneumonia and marked pulmonary emphysema related to BRSV infection. This clinical field study shows that both the test product and the reference product were efficient as treatment of severe bovine respiratory disease in cattle. Statistical analysis of the data shows a significantly higher efficacy of Resflor® compared to Draxxin® regarding the success rate of treatment and in abating high temperatures. 685 Comparative Field Clinical Efficacy of a Single Intramuscular Injection of Marbofloxacine in the Treatment of Bovine Respiratory Disease O. Roy , F. Pillet , A. Heurtin-Vallé , N. Frontczak 1 CEBIPHAR, CRO, Fondettes, France 2 Vétoquinol, Lure, France

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This multicentric field clinical trial aimed to assess the efficacy and the safety of Marbocyl® S (marbofloxacine) as treatment of bovine respiratory disease in comparison with Draxxin® (tulathromycin) and Nuflor® (florfenicol). Two hundred cattle showing respiratory signs, depression, and rectal temperatures > 39.6 °C were included on 21 farms. They were randomly allocated to three treatment groups and given one injection on D0 accordingly (8 mg marbofloxacine/kg, 2.5 mg tulathromycine/kg, or 40 mg florfenicol/kg). Clinical signs were scored and rectal temperatures measured on D0, D1, D2, and D7. Pasteurella multocida, Mannheimia haemolytica, and Mycoplasma bovis were isolated on D0 from the animals in 14, 6 and 3 farms, respectively. Serological conversions related to natural infections with BVD and RSV viruses were detected in 1 and 5 farms, respectively. Rectal temperatures sharply decreased after treatment, mean values being lowest in the Marbocyl® S group (P = 0,0096) on D1. Respiratory signs, general behaviour and appetite scores progressively decreased in all groups, with no significant difference between them. On D1, the success rate was significantly (P=0.0299 and P=0.0540) higher in the Marbocyl® S (62.7%) group than in the other groups (< 50%). Success rates on D2 with Marbocyl® S, Nuflor®, and Draxxin® were respectively at 83.1%, 82.6%, and 76.4% (no significant differences). Relapses were observed on D7 in 1.5%, 6.9%, and 9.2% of cases treated with Nuflor®, Marbocyl® S, and Draxxin®, respectively, with no significant differences. Two animals died in the Draxxin® group due to respiratory disease, and none in the Marbocyl® S and the Nuflor® groups.The local tolerance was good in 96.5% of all the treated animals; 5, 1, and 1 animals displayed moderate signs of intolerance to Nuflor®, Draxxin®, and Marbocyl® S, respectively. This clinical field study showed that the 3 tested products were efficient as treatment of respiratory disease in cattle. Marbocyl® S induces cure significantly faster than the 2 other products tested products.

Infectious and Zoonotic Deseases (Public Health) 97

686 Characterisation of Cattle Originated Escherichia coli O157:H7 Isolates by Randomly Amplified Polymorphic Dna(RAPD) and Antibiotic Resistance O. Aslantas , Z. Cantekin , B. Sareyyupoglu , M. Akan 1 Mustafa Kemal University, Microbiology, Antakya, Turkey 2 Ankara Univercity Veterinary Faculty, Microbiology, Ankara, Turkey In this study, it was aimed to characterize 50 cattle originated Escherichia coli O157 isolates by randomly amplified polymorphic DNA (RAPD) analysis and their antibiotic resistance. RAPD analysis separated the E. coli O157 isolates into two clusters and 35 single isolates. All the strains were found to be resistant two or more of the antimicrobial agents tested. All the resistant strains were resistant to erytromycin and were suscaptible to amikacin, cefaperazone, imipenem and amoxycillin/clavulanic acid. Key words: E. coli O157, RAPD, Turkey 687 In vitro Antimicrobial Susceptibility of Brucella abortus Isolated in Korea H. Eun Jeong, K. Jong Wan, H. Moon, C. Dong Hee, K. Sung Il, C. Yun Sang, H. In Yeong, J. Suk Chan National Veterinary Research & Quarantine Service, Animal Disease Research, Anyang-city, Korea, South Objectives of study: Brucellosis is the most common zoonotic infection worldwide. Recently, Brucella spp. have been recognized as a group of microorganisms that could be used as biological weapons of terrorism. Brucellae are intracellular bacteria, therefore treatments are very difficult. In Korea, isolated bovine Brucella strains are not tested for antimicrobial susceptibility. This study deals with the drug susceptibility of naturally occurring Brucella abortus strains. Materials and Methods: During 1998 to 2006, B. abortus strains were isolated from dairy and beef cows (Hanwoo). The districts from which the strains were isolated are eight areas. The colonies of eighty four strains were tested for biological characteristics. Then, these strains were evaluated by means of inhibition study, minimal inhibitory concentrations (MICs) performed by method of broth microdilution susceptibility test. Results: Tetracyclines demonstrated the highest activity of any antimicrobial agent in this study (MIC range; 0.063~0.5 mg/mL). Among tetracyclines, minocycline had greater activity with MIC50 and MIC90 of 0.125 mg/mL. The MIC50 of tetracycline and doxycycline was same as 0.25mg/mL and the MIC90 of these two drugs are also same as the MIC50. Four quinolones, ofloxacin, norfloxacin, enrofloxacin and ciprofloxacin showed high activity against B. abortus. The MIC50 of ciprofloxacin was 0.25 mg/mL and MIC90 was 1 mg/mL. The MIC50 and MIC90 of ofloxacin and enrofloxacin were 0.5 mg/mL and 1 mg/mL, respectively. However, the MIC50 and MIC90 of norfloxacin was 4 mg/mL, which was less activity against B. abortus than other quinolones. Streptomycin and erythromycin had effective activities with MIC range 0.5~2 mg/mL and 1~4 mg/mL respectively. Rifampin had a good effect with MIC range from 0.5 to 4 µg/mL. The combination of tetracycline and rifampin showed good effect with MIC range from 0.063 to 0.5 mg/mL. Conclusions: The choice of antibiotics in this study focused on in vivo application to humans. We observed variation in sensitivity among B. abortus isolates in Korea. Tetracyclines are the most effective drugs of all other drugs measured. And aminoglycosides and quinolones showed active effect against B. abortus strains isolated in Korea. The combination of tetracycline and rifampin was used in some clinical reasearch. This study also demonstrated active effect of combination of tetracycline and rifampin. Key words: Brucella abortus, antimicrobial susceptibility, minimal inhibitory concentrations (MICs) 688 Persistent Infection of Foot and Mouth Disease Virus in some Slaughtered Cattle and Buffalos in Ahwaz Abattoir by RTPCR M. Ghane , A. Rasoli , M. Seyfiabad Shapouri , M. Haji Hajikolaei , 4 A. Komeilian 1 School of Veterinary Medicine, Shiraz Univrsity, Shiraz, Iran, Department of Clinical Science, Shiraz, Iran 2 Faculty of Veterinary Medicine, Shahid Chamran University, Department of Clinical Science, Ahwaz, Iran

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Faculty of Veterinary Medicine, Shahid Chamran University, Department of Pathobiology, Ahwaz, Iran 4 Islamic Azad University, Science and Research Branch, Department of Clinical Science, Tehran, Iran Foot and Mouth Disease (FMD) is a highly contagious disease of cloven-hoofed animals and has great potential for causing severe economic losses. An important feature of FMD in ruminants is persistent infection that also called carrier state.This may occur in FMD convalescent period as well as in vaccinated ruminants following exposure to infectious virus. In order to evaluate prevalence of persistent infection of FMDV in cattle and buffalos slaughtered in Ahwaz abattoir, 200 head of mature female cattle and buffalos were studied. After slaughter, samples were collected from oropharyngeal fluid. RT- PCR was carried out with general primers for all major virus serotypes. Out of 100 cattle samples, none of them were positive for virus but out of 100 buffalos samples, two positive samples were observed. Since buffalos are in close contact with other sensitive animals such as cattle and sheep, they may be of great importance in spreading the disease in the area. 689 Prevalence of Bovine Herpesvirus Type 1 in Southern Backa and Srem District in the Republic of Serbia S. Lazic , T. Petrovic , D. Lupulovic , M. Maljkovic , D. Bugarski 1 Scientific Veterinary Institute Novi Sad, Virology, Novi Sad, Serbia 2 Scientific Veterinary Institute Novi Sad, Department of Epizootiology, Novi Sad, Serbia Determination of the prevalence of bovine herpesvirus type 1 (BHV-1) in Southern Backa and Srem district was carried out by determining specific antibodies against BHV-1 using commercial ELISA test (Test Line Ltd, Check Republic). The total number of cattle in these two districts was 123911, but the examination comprised 22139 animals (17.87%). The estimation of prevalence was done using software program Win Epscope version 2. The prevalence of infection was calculated for every municipality, and then for the district. In Southern Backa district from the total number of 52383 the examination comprised 11911 animals from all 12 municipalities. The BHV-1 seropositive animals were detected in all the municipalities, except in Sremski Karlovci. In this municipality out of 114 animals, 35 were examined and all were seronegative. In other municipalities the percent of seropositive animals varied and ranged from 11.84% (Temerin municipality) to 33.46% (Becej municipality). In most municipalities (5) the percent of BHV-1 seropositive animals ranged from 11.84% to 20%. In 3 municipalities the percent of seropositive animals was 25%, and it was above 30% in 3 municipalities. In Srem district the examination was carried out in all 7 municipalities, so out of 71,548 animals 10,228 were examined. The smallest number of seropositive animals was discovered in Ruma municipality (14.57%), and the largest in Pecinci municipality (23.33%). In two municipalities (Sid and Sremska Mitrovica) the percent of seropositive animals was lower than 20%, while in other municipalities (Indija, Irig and Stara Pazova) the percent of BHV1 seropositive animals ranged from 21.60% to 22.20%. Based on these results it may be concluded that in Southern Backa district out of 52363 cattle there were 11911 (22.75%) examined on BHV-1. The specific antibodies against BHV-1 were detected in 3123, what means that the prevalence was 26.22%. In Srem district out of 71548 animals there were 10228 examined animals (14.29%), but specific antibodies against BHV1 were detected in 2045 animals what means that the prevalence was 19.99%. Key words: BHV-1 prevalence, ELISA, Serbia 690 Importance of BHV-1 in Etiology of Bovine Respiratory Disease Complex in Calves in Province Vojvodina (Republic of Serbia) D. Bugarski , T. Petrovic , I. Pusic , D. Lupulovic , D. Milanov , 2 S. Lazic 1 Scientific Veterinary Institute Novi Sad, Department of Epizootiology and Health Protection of Ruminants, Novi Sad, Serbia 2 Scientific Veterinary Institute Novi Sad, Department of Virology, Novi Sad, Serbia 3 Scientific Veterinary Institute Novi Sad, Department of Microbiology, Novi Sad, Serbia In Serbia eradication of BHV-1 infection has been voluntarily carried out, so prevalence ranged from 5% to more than 50% depending on the

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area. The importance of BHV-1 in etiology of respiratory disease complex in beef calves was conducted in the years 2005-2007. It was carried out on 12 fattening farms (250-500 animals per herd) by determining seroconversion after enzootic pneumonia outbreak, examining the presence of antibody titer before slaughtering and the virus isolation from nasal swabs was also done at the beginning of an acute respiratory infection. A characteristic feature common for all the farms is that new animals are introduced several times a year and therefore it is impossible to avoid simultaneous presence of different aged animals, what gives a possibility for long retention of the virus in a herd. Beef farms in Vojvodina purchase young calves from mountain regions, where the prevalence of BHV-1 in adult animals is even 45%. On all the farms the animals were clinically examined; out of total number the blood was sampled from 3-10%, most of them were sick, chronically sick or recovered from the respiratory syndrome. The second sampling, for determining seroconversion, was carried out 3-4 weeks after the first one. In all the fattening farms enzooty of bronchopneumonia was present. Clinical symptoms were: increased body temperature, weak serous nasal discharge, coughing and dyspnoea. BHV-1 seroconversion in totally recovered and animals chronically ill from bronchopneumonia was not determined in all the examined herds. In newly arrived calves seroprevalence on BHV-1 ranged from 0-10.56%. At the end of fattening period antibodies against BHV-1 ranged from 0-100%, depending on a herd. However, BHV-1 was not isolated during acute infection. From the obtained results it may be concluded that, currently, BHV-1 is not of great importance for developing respiratory syndrome in the herds of beef calves, what was not the case in past decades, so the infection is subclinically present. In fact, in all the beef farms, after the recovery from the disease, immuno-response towards BRSV and Pi-3 was determined. In the lungs of dead animals only mycoplasma was determined, or mycoplasma with bacteria. Key words: BHV-1, beef, respiratory syndrome, Vojvodina 691 Effect of Vaccination against Leptospira on Dairy Farm Reproduction Indexes D. Ruiz Di Genova COVAP, Bovine Technical Services, Pozoblanco, Spain Leptospirosis is a worldwide occurring disease, that when caused by serovars adapted to dairy cows (serovar Hardjo) produce chronic infections, where the signs are less evident. Objective: To assess the effect of vaccination against Leptospira interrogans serovar Hardjo on the reproduction indexes. Materials and Methods: The study includes 9 dairy farms (mean 70 cows/farm), in southern Spain with complete reproductive records comprising a year before the first leptospira vaccination and a year afterwards. In seven farms, serological diagnosis of leptospirosis was performed by MAT (microagglutination test) for serovar hardjo, prior to vaccination. A positive farm was considered when presenting one animal with a titter > 1/10. Another two farms were vaccinated due to the clinical suspicion, after ruling out other reproductive diseases.The monthly conception rates and first service conception rates were used as fertility indexes, comparing results obtained one year before vaccination with results obtained on the year afterwards.Cumulative data were analyzed using logistic regression, which allows to establish a model for dicotomic results (pregnant/not pregnant). The result is expressed as an odds ratio indicating the different occurrence probability of one or the other event, that is, the probability of pregnancy after one service, before and after vaccination. Results: The probability of pregnancy in vaccinated cows was 1,25 times higher than in not vaccinated ones. (OR=1,25399, EE=0,0673; P=0,0008), with the "vaccination" variable accounting for 21% of this variation in fertility. In the first-calving cows the probability of conception increased 1,70 times after vaccination (P<0,001), with a 40% variation explained by the model. No differences were found for first service fertility in cows which had calved more than once, but in first-calving cows the odds for first service pregnancy increased more than 90% (OR=1,93; EE=0,21; P=0,0015), with vaccination accounting for 26,74% of the fertility variation between both periods. Conclusions: Vaccination against leptospirosis with monovalent vaccines (Leptavoid®) has a significant effect on cow fertility in infected farms, especially in first-calving cows. Due to the difficulties to obtain a precise diagnosis of leptospirosis, vaccination is a useful tool in case of a clinical suspicion with chronic reproductive failure, especially marked in young animals.

692 Evaluation of the Immune Response in Inoculated Calves with a Recombinant Vaccine of Mycobacterium Vaccae against Bovine Tuberculosis M. Jaramillo , C. Martínez , O. Díaz , A. Espinoza , S. González , 2 1 1 3 1 G. Pérez , F. Santillán , T. Romero , P. Espitia , D. Arriaga 1 INIFAP, CENID Microbiologia, México DF, Mexico 2 Universidad Nacional Autonoma de México, Facultad De Medicina Veterinaria, Mexico State, Mexico 3 Universidad Nacional Autonoma de México, Immunology, México DF, Mexico The aim of this study was to evaluate the immune response induced in calves inoculated with a recombinant vaccine constructed in Mycobacterium vaccae to the prevention of the bovine tuberculosis in field conditions. The recombinant vaccine was obtained by transformation of the Stanford strain of M. vaccae with the pCAP3 plasmid containing the gene encoding the M tuberculosis phosphatebinding protein PstS-1. The study was carried out in a dairy herd with a prevalence of bovine tuberculosis of 35 % determined by reactivity to tuberculin. Forty calves of one month of age and negatives to diagnostic test to disease were divided in two groups of 20 animals each. One of them was subcutaneously inoculated with 104 CFU of the vaccine, while the other group was a control. The animals were periodically sampled to evaluate their humoral immune response by a comparative ELISA, employed as antigens the culture filtrate protein extracted (CFPE) of M. bovis and M. avium. Whole-blood culture gamma interferon (IFN-g) production in response to mycobacterial antigens was measured employing a commercially-available sandwich ELISA kit (BOVIGAM). The tuberculin test was applied eight months after the vaccination to both groups. Samples periodic of nasal exudates of the animals were cultured in Stonebrink media to intent the isolation of mycobacteria. The results showed a low but significant level of IFNg production in the vaccinated group during the time evaluated, while in the control group significant variations were observed with the different mycobacterials antigens assayed, included the ESAT-6 (P<0.05). On the other hand, the response of antibodies in the vaccinated group was important to M. bovis until third month after vaccination (P<0.05). Although, high levels of antibodies in the control group were observed since the first month of evaluation. None of the animals of both groups showed reactivity to the tuberculin and none was positive to isolation M. bovis. The results indicate that in spite of not being observed a significant immune response in the vaccinated group, this response conferred some grade of resistance to infection such as indicate their low response to ESAT-6 antigen during experimental period, regards to control group which response was higher. This antigen has been considered of utility to difference animals infected of vaccinated, so it can be assumed that in vaccinated animals a protective response against the disease was induced. Key words: bovine tuberculosis, vaccine, Mycobacterium bovis, immune respose, Mycobacterium vaccae 693 Evaluation of BCG Vaccine and M. bovis Culture Filtrate Protein Extract against Bovine Tuberculosis in Field Conditions O. Díaz , A. Espinosa , M. Jaramillo , S. González , G. Pérez , 1 4 5 3 D. Arriaga , N. Bojórquez , C. Soto , P. Espitia 1 INIFAP, CENID Microbiologia, México DF, Mexico 2 Universidad Nacional Autonoma de México, Facultad de Medicina Veterinaria y Zootecnia, Mexico State, Mexico 3 Universidad Nacional Autonoma de México, Immunology, México DF, Mexico 4 Institute for Diagnosis and Animal Health, Laboratório de Produçio de Imunobiológicos, México DF, Mexico 5 private, Animal Health, Mexico State, Mexico Efficacy of protein extract culture filtrate (EPFC) of Mycobacterium bovis and BCG vaccine was evaluated using 6-40 days old non infected Holstein Frieisian heifers, from a herd that had 50% TB prevalence. Sixty six heifers were vaccinated subcutaneusly with EFPC (300 µg), 82 were vaccinated with M. bovis BCG (104 UFC) and 28 animals remained as controls. The animals were then mixed with the rest of the herd. Ten animals from each group were sampled every 30 days for a year. The production of IFN was evaluated using bovine PPD, avian PPD and ESAT-6 antigens (Bovigam Tests CSL). Antibodies production was also evaluated by a comparative ELISA

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using EPFC. The intradermal tuberculin comparative test was applied to all animals at 12 and 24 months postvaccination. IFN production of cell cultures stimulated with bovine PPD showed significant increase (1.4, 0.90, 1.4 OD) at days 15, 90 and 180 postvaccination, in the group innoculated with EFPC. In the group vaccinated with BCG the increase was observed at 60 and 150 days postvaccination (0.90, 1.7 OD) while the control group showed increment at day 90 (1.35 OD). Animals from the three groups showed an increse (1.4 OD) at day 60 when stimulated with avian PPD. Stimulation with ESAT 6 did not give a clear response in any of the groups. With respect to the humoral response, all groups showed an increase in OD at day 90 but was significantly higher (P<0.05) in the group vaccinated with EPFC. The tuberculin skin test of the group vaccinated with EPFC showed that after 12 and 24 months 6/66 (13%) and 7/66 (11 %) were reactors. In the group vaccinated with BCG no reactors were found after 12 months but 4/88 (5%) were reactors after 24 months. In the control group 5/28 (23%) and 8/28 (29%) were reactors at 12 and 24 months, respectively. Considering that reactivity can be attributed to infection with M. bovis field strains we could conclude that the best protection was shown by BCG, followed by CFPE. On the other hand, the skin test showed a global reduction of the prevalence of the disease in the herd from 50 to 11% which indicates a real benefit in the use of these vaccines since they may increase the resistance to the disease at herd level. Key words: bovine tuberculosis, vaccine BCG, immune response, cattle 694 Detection of Mycoplasma bovis in Calves and Young Cattle in the Czech Republic D. Zendulková , K. Rosenbergová , Z. Pospísil , D. Haas , 2 2 R. Dvorák , L. Pavlata 1 University of Veterinary and Pharmaceutical Sciences Brno, Department for Infectious Diseases- Epidemiology, Brno, Czech Republic 2 University of Veterinary and Pharmaceutical Sciences Brno, Clinic of Ruminants Diseases, Brno, Czech Republic Mycoplasma bovis is at present a wide-spread microorganism, posing an important veterinary problem for cattle herds in the USA, Canada and some European countries. In the Czech Republic Mycoplasma bovis was isolated for the first time and only occasionally in 1975. Since that time till 2007 it was not detected here. However, in the neighbouring countries its prevalence has recently increased, though an exact prevalence is difficult to present because there are no reliable data on the actual situation. Between January and December 2007, a survey for the presence of mycoplasmas was performed in selected herds of calves and young cattle from various parts of the Czech Republic. Animals suffering from respiratory disease, in which nasal discharge, distressed breathing and respiratory rales were observed, were chosen for our investigation. A total of 126 samples (63 nasal swabs and 63 transtracheal aspirates) were collected from them and then processed for culture and identification of potential Mycoplasma spp. Transtracheal aspirates and isolates, in which Mycoplasma bovis was suspected, were analyzed by a nested-PCR. Mycoplasma spp. were isolated from nasal swabs in 76.8%, and the nested-PCR identified 23.7% of them as Mycoplasma bovis. The examination of transtracheal aspirates revealed that 69.6% of them were positive for Mycoplasma spp. The nested-PCR identified Mycoplasma bovis in 53.9% of these samples. It can be concluded that, in the Czech Republic, the isolation and identification of Mycoplasma bovis was again positive after more than 30 years from the first occasional detection. Our present results show that the number of animals infected with this pathogen increased considerably in comparison with 1975. One of the possible explanations for the spread of Mycoplasma bovis in cattle populations can probably be found in free transport of animals amongst farms and countries of the European Union. Supported by the project of MSMT No. 6215712403 Key words: Mycoplasma bovis, cattle, isolation, PCR 695 Determination of Lymphocytes Subpopulations in Cattle Vaccinated with BCG Vaccine and Mycobacterium bovis Culture Filtrate Proteins O. Díaz , M. Jaramillo , J. Jimenez , P. Espitia , L. Lascuraín 1 INIFAP, CENID Microbiologia, México DF, Mexico

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Universidad Autónoma Metroopolitana Unidad Xochimilco, Facultad de Medicina Veterinaria y Zootecnia, México DF, Mexico 3 Universidad Nacional Autonoma de México, Immunology, México DF, Mexico 4 Universidad Nacional Autonoma de México, Faculty of Medicine, México DF, Mexico One of the strategies to develop new and better vaccines against bovine tuberculosis is based on identification of those antigens capable of activating T lymphocytes with a Th1 cytokine secretion profile, which is characterized for production of gamma interferon (IFN-g) and Interleukine-2 (IL-2), cytokines associated with resistance to mycobacteria in experimental models. The aim of this work was to characterize lymphocyte subpopulations CD4, CD8 and WC responsive to mycobacterial antigens and determine the expression of the activation markers CD25 and CD45 in bovine immunized against bovine tuberculosis. For that eighteen Holstein heifers of eight months old and negatives to diagnostic test to disease. were employed They were divided in three groups of six animals each, which were designed as groups A, B, C and D. The group A was subcutaneously inoculated with 300 µg of culture filtrate protein extracted of Mycobacterium bovis; group B, received CFPE + IFN-g; group C was vaccinated with 6 10 CFU of BCG and the group D was control. All animals were challenged intratracheally, 5 months after vaccination with 105 CFU of a field M. bovis strain. PBMC from the blood of the animals were separated by gradient and in vitro stimulated with different antigens: bovine PPD, avian PPD, recombinant proteins Dip-Z and complex 45/47. Dip-Z is a lipoprotein of 27 kDa express in M. bovis, while the complex 45/47 is constituted by 45-47 kDa glycoproteins that show homology to 50-55 kDa glycoproteins of M. tuberculosis. Cells were collected after stimulation and analyzed by FACS using respective monoclonal antibodies. Production of IFN-g was determined in the supernatans using the BOVIGAM commercial kit. Stimulation of CD4 and gd (WC1) lymphocytes, rather than CD8, was observed with different antigens in vaccinated groups. However the group C showed the highest percentage of gd and CD8 cells and higher expression of IL2 receptor which correlates too with a significant IFN-g production. An important production of this cytokine was observed in all groups by complex 45/47 after challenge. The high reactivity observed in the BCG vaccinated group indicate the development of significant immune response in this group more than in the groups immunized with CFPE. Key words: lymphocytes subpopulation, cattle, bovine tuberculois, BCG 696 Tilmicosin Penetration in Aqueous Humor after its Subcutaneous Administration to Healthy Cattle and Cattle with Infectious and Traumatic Keratoconjunctivitis N. Mestorino , L. Marchetti , A. Lambertini , E. Turic , G. Rojas , 1 1 1 1 B. Huber , M. Lucas , M. Daniele , J. Errecalde 1 Faculty of Veterinary, UNLP, Pharmacology, La Plata, Argentina 2 BIOGENESIS BAGO S.A., Reserch & Development, GARIN, Argentina The objective of this study was to evaluate tilmicosin (TMS) penetration in aqueous humor (AH) of healthy cattle and cattle with infectious (IBK) and traumatic keratoconjunctivitis.TMS 30% (Biogénesis-Bago, Argentina) was administered by the SC route to 4 different animals groups, as follows: G1: 10 mg/kg, 10 Aberdeen Angus (AA) steers with aseptic keratitis G2: 5 mg/kg, 40 indian cross calves with Moraxella bovis IBK G3: 10 mg/kg, 40 AA calves with M. bovis IBK G4: 10 mg/kg, 40 healthy AA calves. The AH extraction was performed in the external iridocorneal angle. G1: AH samples were obtained of each of 10 animals at different post administration times until 120 h. A traumatic injury with development of an aseptic keratitis was caused by the successive extractions. G2-G4: Groups of 4 animals were alocated to each sampling point to perform only one extraction per animals, avoiding tissue lesion during the repetitive AH extraction. The extraction times were the same as for G1. AH TMS concentrations were measured by microbiological assay. Pharmacokinetic parameters were determined by non-compartmental analysis by WinNonlin 5.2.TMS AH concentrations were low, which could be explained by the fact that the AH is a fluid with low cellular and protein content, that suffer constant refill, being its volume replaced completely every 90min. With similar dose, sick animals exhibited TMS concentrations 2 times above those found in G4. The inflammatory

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response in the eye produces accumulation of polymorphonuclear neutrophils (PMNs) in tissue and ocular fluids, increase of the vascular permeability and therefore an increase of AH proteins. The increase of PMNs explains the larger concentrations of TMS reached in AH of the animals with keratoconjunctivitis. Although TMS levels in AH (either at 5 or 10 mg/kg) persisted during 72-96 h, they never surpassed the M. bovis MIC (< 1 µg/mL). Nevertheless a longer TMS persistence was evident in sick animals, being documented through longer mean residence time (41.2, 45.7 and 22.7 vs 15.9 h for the G1, 2, 3 and 4 respectively) and a smaller clearance from AH (222.8, 469.5, 773.3 vs 1613.5 mL/h/kg for the G 1, 2, 3 and 4, respectively). These information suggest that TMS penetrate to the ocular balloon and concentrates in AH depending on the degree of inflammation. Concentrations in vitreous humors and ocular tissue, should be evaluated in the future to consider the use of the TMS for the treatment of certain ocular infections. Key words: tilmicosin, aqueous humor, keratoconjunctivitis, pharmacokinetic, cattle 697 The use of PCR for Identification of Brucella abortus in Samples from Aborted Fetuses in a Dairy Farm in the State of Aguascalientes, Mexico D. Limón , G. Martínez , G. Zenteno , B. Arellano , E. Diaz , 1 1 R. Flores , F. Morales 1 INIFAP, CENID Microbiologia, MEXICO, D.F., Mexico 2 Universidad Nacional Autonoma de México, Facultad De Medicina Veterinaria, MEXICO, D.F., Mexico Abomasal fluid and tissues conserved in 10 % bufferated formalin solution from 13 aborted fetuses, as well as sera from 55 aborted cattle were collected in a dairy farm located in Aguascalientes, Mexico and were transported to the Diagnostic Laboratory of CENIDMicrobiologia, INIFAP. The abomasal fluid samples were inoculated in enriched Farrel's medium in order to attempt the Brucella abortus isolation. The tissues samples were processed for histophatological analysis, while the sera were used for detection of BVD, IBR, Neospora, Leptospira and Brucella antibodies. Those Brucella spp suggested isolations were examined by using a PCR procedure. Depending of the amplified fragments (361, 456, 1063 and 1298 bp) it is posible the identification and differentiation of field and vaccinal Brucella abortus strains. Six isolated strains were cultured from the abomasal fluid and all of them were identificated as Brucella abortus field strains by PCR. Brucellosis typical lesions (multifocal necrotical hepatitis and exudative pneumonia) were observed in 4/13 studied fetuses. The result of serological examination of 55 samples was as follows: Brucellosis, 27 %; Neosporosis, 40 %; Leptospirosis, 65 % (titers > 1:400) and IBR, 100% and BVD 80 %. During the last few years Leptospirosis and Neosporosis have shown an increase in their importance as a cause of abortion in cattle in Mexico, our results in the present study confirm this affirmation. The fact that a specific vaccination program to prevent neosporosis does not exist in the dairy farm under investigation, makes it important to consider this agent as important causal of the abortion. In regards to Brucellosis, it is a disease with high prevalence in the dairy cattle farms despite the fact that there is an intensive official control campaign in Mexico and excellent vaccines such as S19 and SRB51 exist; for this reason it is important to have diagnosis techniques with high sensivity and specificity as it is the case of PCR technique. It is important to promote the use of integral etiological diagnosis of abortions in order to be in conditions to apply adequate and effective control measurements. It often occurs that in some laboratories in Mexico the samples are tested only for diagnosis of brucellosis and do not consider the use of techniques for determinate the possible presence of other microorganisms associated with reproductive problems. Financial support SAGARPA-CONACyT. (2004/COI/23) Key words: PCR, Brucella abortus, aborted fetuses 698 Long-Term Excretion of Brucella abortus, Strain B19 in Milk and Urine by Vaccinated Adult Female Bovines M. Genovez , W. Pacheco , C. Pozzi , L. Paulin , M. Gamberini , 4 S. Azevedo 1 Instituto Biológico, Animal Health Center, Sâo Paulo, Brazil 2 Instituto de Zootecnia, Animal Health Center, Nova Odessa, Brazil 3 Universidade Federal de Goiás, Animal production, Goiania, Brazil 4 Universidade Federal De Campina Grande, Department for Infectious diseases- Epidemiology, Patos, Brazil

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Abortion caused by Brucella abortus is a common bovine reproductive disease in many countries and may be currently prevented by vaccination with live B19 vaccine, administered to 3-8 month old heifers. The length of excretion, its relation to the hormonal cycle and its effect on contactant and susceptible animals, including man, are not completely known. The persistence of B19 excretion in milk and urine by adult dairy cows vaccinated at 3-8 months old was investigated in this study. Samples were collected during 12 months (15 collections), beginning at estrus until immediate post-partum, from 14 females in three groups: G1(3-4 years old), G2 (5-6 years old) and G3 (7-9 years old). Milk and urine samples were monitored by bacteriologic analyses and Multiplex PCR differential for B.abortus and B19 strain; blood serum samples were analyzed by Rose Bengal test for brucellosis diagnosis and direct estradiol ELISA KIT and progesterone ELISA KIT (Diagnostics Biochem., Canada) were used for hormone monitoring. All milk samples were negative in serology and bacteriologic examination. B19 was detected by PCR in 74/210 (35.2%) and 12/210 (5.7%) from urine and milk samples, respectively (p=0.001). Excretion mainly occurred from the beginning of estrus to 150 days of pregnancy and for 30 days after parturition. Hence, excretion was intermittent, no matter the age of the animal and the time after vaccination. Hence, excretion was intermittent, no matter the age of the animal, the time after vaccination, and occurred in cows up to nine-year old. Hormonal stage did not seem to affect B19 excretion, and may be associated with erythritol scarcity. The impact of longterm B19 excretion requires more attention. B19 may remain in the environment with a real possibility of transmission to negative cattle herds, mainly to bulls, or even to other susceptible animal species, including man. Urine was the best sample for B19 detection. This paper was supported by FAPESP: Fundaçao de Amparo a Pesquisa do Estado de Sao Paulo, Brazil Key words: bovine brucellosis, B19 excretion, hormonal cycle 699 Main Causes of Abortion in Cattle at Different Areas of Mexico G. Zenteno , F. Manrriquez , I. Vitela , L. Diaz , R. Flores , 3 1 D. Martinez , F. Morales 1 INIFAP, CENID Microbiologia, México DF, Mexico 2 Instituto Tecnológico el Llano, Microbiología, Aguascalientes, Mexico 3 Universidad Veracruzana, Facultad de Medicina Veterinaria y Zootecnia, Facultad De Medicina Veterinaria, Veracruz, Mexico 4 Gobierno del Estado de Aguascalentes, Comité de Fomento y Protección Pecuaria, Aguascalientes, Mexico Data related to the incidence and prevalence of etiological agents associated with reproductive failures in cattle of Mexico are the result of isolated studies, only a few cases include results of integrated diagnosis. Some papers describe the results of serological profiles, but in most of them the interpretation of the results does not permit the diagnosis of the etiological agent. For example, previous studies performed by our research group demonstrated different prevalences of brucellosis, BVD, IBR, neosporosis and leptospirosis as follows: The brucellosis prevalence was always high (12-21 %) except in Veracruz (3%); Leptospirosis was similar to brucella with the highest prevalence in Aguascalientes (27%) and the lowest in Veracruz (3%). The BVD and IBR highest prevalences were observed in the same states (56-86%) while in Veracruz there were lower (47 and 38 % respectively). The prevalence of neosporosis varied between 30 and 45 % in most states, but in Veracruz was only 15 %. With these results it was impossible to relate the abortions with a particular infectious agent. The purpose of this study was to perform an integral, comparative serological profile, in sera collected from aborted and non aborted animals from 7 dairy cattle in different areas of the state of Aguascalientes, Mexico. Blood samples were collected from cattle aborted during a five month period previous to the collection and from non aborted animals. All samples were tested for the identification of Brucelosis, BVD, IBR, Neosporosis and Leptospirosis antibody titers. The results of this serological study, were then used in order to compare the prevalences of each etiological agent with either, aborted and non aborted populations. The results of this comparative study demonstrated that in some farms more than one etiological agent was found to be important: in 4 cattle farms brucellosis was a serious problem; in one farm the main cause of abortion was IBR, while BVD and neospora appeared to be the most important causes of abortion in

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three cattle farms each. Leptospirosis was also present. It is notorious the fact that in dairy cattle different etiological agents could be involved in reproductive problems, therefore the diagnosis procedures must be integral, including a set of serologic test that allows to determinate their presence and to be able to apply adequate and effective control measures. Financial support: SAGARPA-CONACyT. (2004/COI/23) Key Words: abortion, cattle, serology 700 Viability of Mycobacterium Avium Subsp. Paratuberculosis (MAP) in Baled Grass Silage J. Khol , V. Beran , P. Kralik , M. Trckova , I. Pavlik , 1 W. Baumgartner 1 Clinic for Ruminants, Department for Farm Animals and Herd Health, Vienna, Austria 1 Veterinary Research Institute, Brno, Czech Republic Objective of the study: Ensiling of grass from areas where livestock or wild animals infected with paratuberculosis are grazing can be a possible source of Mycobacterium avium subsp. paratuberculosis (MAP) infection in domestic ruminants. The results of a preliminary study about the viability of MAP, the causative agens of paratuberculosis, in baled grass silage should be presented in this poster. Materials and Methods: Seventeen bales of grass silage were spiked with a suspension containing MAP. Silage samples were collected periodically for four months to observe MAP viability over time. Collected samples were tested for MAP by bacterial culture and Real Time-PCR (Polymerase Chain Reaction). Results: Viable MAP could not be detected at any sampling date during the trial by culture, more than 60 % of the analysed samples were tested positive by Real Time-PCR. Conclusions: Even though the results of the presented work suggest that grass silage seems to be an unlikely source of paratuberculosis infection for livestock, further investigations are needed to elucidate the reaction of MAP to unsuitable environmental conditions and their influence on the infectivity of the bacterium. 701 Phylogenetic Position of an Unreported BPV Type Detected From a Cutaneous Lesion of a Brazilian Cattle Herd M. Lunardi, M. Claus, A. Alfieri, A. Alfieri Universidade Estadual de Londrina, Departamento de Medicina Veterinária Preventiva, Londrina, Brazil In Brazil, Bovine Papillomavirus (BPV) infections are endemic in beef and mainly in dairy cattle herds. Despite the high frequency of BPV infection, the identification of BPV types in Brazilian cattle is still sporadic. In a prior study, through the analysis of a partial segment of L1 gene, we could identify the presence of previously described BPV types and four putative new BPV types associated with skin warts. The aim of this study was to determine the entire L1 nt sequence of a putative novel BPV type (BPV/BR-UEL2) detected in Brazil, and thus state its phylogenetic position. As a phylogenetic analysis employing the FAP amplicon had revealed our isolate as closest related to BPV-4 (Xi genus), two pair of degenerate primers were designed by using alignments of L1, L2, LCR regions of genome of Xi genus representatives. In addition, aiming to obtain the full L1 gene sequence, the previously described FAP primer pair was also employed both in the original form as in combination with designed primers. The PCR amplicons were purified from agarose gel and submitted to cloning. Plasmid DNA from 2 clones was sequenced in both directions using M13 forward/ reverse primers. Sequences were examined with the software PHRED for quality analysis and the consensus sequence was determined using the software CAP3. The alignment was obtained with the software BioEdit. A neighbour-joining phylogenetic tree was constructed with complete L1 ORF sequences of 38 PVs classified in 18 genera, and with the entire L1 ORF sequence of BPV/BR-UEL2 isolate, using the MEGA program v.3.1. A consensus sequence, representing the terminal portion of L2 gene, the complete sequence of L1 gene and an initial segment of LCR, could be achieved from three PCR amplicons. By ORF analysis, it was possible to determine that the L1 ORF encoded protein of the Brazilian isolate consists of 532 aa. The phylogenetic analysis revealed that the BPV/BR-UEL2 is related with BPV types held in Xi genus. Besides, this isolate displayed the

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highest L1 nt sequence similarity with BPV type 4 (74%), suggesting its classification in the Xi genus. The realization of further studies involving the molecular epidemiology of BPV infections, in Brazilian cattle herds as much in diverse geographical areas around the world, become necessary to verify the prevalence of this new viral type and to check its association with cutaneous lesions. Financial support: CNPq, CAPES, FINEP and FAP/PR Key words: BPV; L1 ORF; phylogeny 702 Monitor pProject : Evidence of Some Respiratory Virus Isolation from Marchigiana Breeding Farms with Respiratory Disorders, Note 2. S. Petrin , M. Panicci , S. Briscolini , L. Cucco , M. Ferrari , 1 1 G. Filippini , G. Pezzotti 1 Istituto Zooprofilattico Sperimentale dell'Umbria e delle Marche, Perugia, Italy 2 Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna Brescia, Brescia, Italy Objectives of study: In the framework of the monitor project, necropsy has been carried out on 52 dead calves originating from herds with bovine respiratory disease (Monitor project note 1). This report describes the results of the bacteriological and virological investigations carried out on subjects died for respiratory disorders. Materials and methods: Organs (trachea, lung, bronchial lymph nodes), were collected from 52 dead animals for bacteriological examination and viral isolation. The bacteriological examination was performed according to Quinn (1999); briefly, specimens were streaked on blood agar (BA), Mc Conkey agar (MC), Mannitol Salt agar (MSA); and incubated for 1224 h at 37 °C. Isolates were characterized according to Quinn (1999), and further identified using the API Systems (Biomerieux). Testing for Mycoplasma spp. was carried out inoculating PPLO agar and broth, incubated for 12 days at 37 °C under 5% CO2, and checked daily. For virus isolation the samples were diluted, centrifuged and the supernatants were inoculated into BEK cells. In absence of citopathic effect (CPE), 3 subpassages were made. In positive cases, the identification of the isolated virus was evidenced with serum neutralization test (SN) using reference immune serums against IBR and with nested-PCR methods according to Vilcek S. et al. (1994) to identify BRSV. For the SN tests virus isolates were diluted serially and mixed with the reference immune serum in 96well microtiter plates which were held for 90 min at 22 °C. After the incubation 20,000 BEK cells in E-MEM were added. The plates were checked for 7 days, and the CPE was evaluated. Virus dilution mixed with E-MEM represented the control. The virus was identified if titre in the presence of immune serum was at least 2 log lower than the virus titre in E-MEM. Results: The results evidenced the presence of the following bacteria: M. haemolytica (15 %); P. Multocida (26 %); Mycoplasma spp (4 %); Negative cases (55 %). From trachea and lung samples of three animals, CPE was detected in BEK cell cultures whose features were typical of IBR or BRSV, respectively. They were detected at the first or second serial passage. IBR and BRSV were identified by SN and nested PCR tests, respectively. Conclusions: The results of the investigation demostrated that viruses, isolated from Marchigiana breeding farms with respiratory disorders are members of the Herpesviridae and Paramyxoviridae family. 703 Previously Described BPV Types and Putative New Types in Cutaneous Papillomatosis from Brazilian Cattle Herds M. Claus, M. Lunardi, A. Alfieri, A. Alfieri Universidade Estadual de Londrina, Departamento de Medicina Veterinária Preventiva, Londrina, Brazil The aim of the current study was to report the identification of putative new BPV types in skin warts in cattle herds from Paraná state of Brazil. Papilloma specimens (n=27) were taken individually from diverse body sites of adult and young bovines, from dairy (n=2) and beef (n=2) cattle herds from Paraná state, South region of Brazil. The PCR assay was carried out using the primer pair FAP59 and FAP64. All PCR products were purified and a direct sequencing was performed with FAP primers. For the amplicons which the prior analysis revealed them as a putative new BPV type, a cloning and a further sequencing, in both directions, was performed employing the plasmid DNA from two selected clones of each sample. For quality analysis of chromatogram readings and determination of the

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consensus sequences, the PHRED and CAP3 softwares were used, respectively. The sequence identity was verified with all sequences deposited in the GenBank by using the BLAST software. The alignment was obtained with the software BioEdit, and a neighbourjoining phylogenetic tree was constructed using the MEGA program v.3.1. A new PV type is defined as a complete genome, whose L1 gene sequence is at least 10% dissimilar from the closest known PV type. As the FAP59/64 products represent only part of the L1 gene, the obtained sequences that had differed from BPV-1 to -10, were named putative new PV types. PV DNA was detected in all DNA samples analysed, resulting in amplicons with 478 bp. The BPV-1, 2, -6 and ­8, were identified in 4, 8, 9, and 2 papilloma specimens, respectively. In addition, four putative new BPV types were identified in other six skin warts. Four Brazilian putative new BPV types were designated as BPV/BR-UEL2 to -5 (accession numbers: EU293538, EU293539, EU293540 and EU293541, respectively). As our putative new BPV types were detected from skin warts, their potential for causing cutaneous lesions could be suggested. Besides, the BPV/BR-UEL4 and -5 isolates represent the first putative BPV types closely related with BPV-2 and BPV-8, respectively. The detection of these BPVs types in skin wart specimens supports the existence of virus diversity throughout the Brazilian livestock. Finally, the identification of four putative new BPV types is the first report of the presence of different BPV types in American continent. Financial support: CNPq, CAPES, FINEP and FAP/PR Key words: BPV, skin wart, phylogeny 704 Monitor Project : Serological Survey in Marchigiana Breeding Farms, Note 1. M. Paniccia', G. Filippini, A. Duranti, F. Ciuti, A. Dorinzi, P. Papa, P. Mangili, S. Petrini Istituto Zooprofilattico Sperimentale dell'Umbria e delle Marche, Perugia, Italy Objectives of study: In order to evaluate the prevalence of the major infections causing respiratory, enteric or reproductive syndromes in the Marchigiana breeding farms of the Marche region (Italy), a surveillance programme, named Monitor, was carried out through serological investigations. The following agents were considered: BHV-1, BRSV, PI-3 and BVDV. Moreover, the presence of antibodies against Mycobacterium avium subspecies paratuberculosis (MAP) was evaluated. Within the reproductive infections, Chlamydia abortus (CA), Leptospira pomona (LP), Leptospira hardjo (LH) and Neospora caninum (NC) were considered. Materials and methods: A cross sectional study was carried from March to December 2006. The target population consisted of bovine farms for meat production present in Marche region (49.737 heads and 2.802 herds). A one-stage cluster sampling was performed and 22 herds were selected. In these farms, 1662 bovine serum samples from adult cattles (>12 months of age) were collected and tested against BHV-1, PI-3, BRSV, BVDV, MAP, NC using commercial ELISA tests, CA through complement fixation test (Donn A et al., 1997) and LP-LH through microagglutination test (O.I.E. Methods). The results were evaluated using Epi-info 3.4.2 to estimate the prevalence of the infections considered. Results: The result demonstrated that all the agents considered were present in the Marche region. In particular, the estimate prevalences were 34,0 % (I.C.95%: 6,7 -61,3) for BHV-1, 91,9 % (I.C.95%: 83,6 -100 %) for PI3 and 54,9% (I.C.95% 37,0 - 72,8%) for BRSV. Moreover, 3,5 % (I.C. 95%: 0,1 - 6,4%) of the animals resulted positive for antibodies against MAP, 3,8 % (I.C.95%: 2,8-6,1%) for CA, 7,4% (I.C. 95%: 2,3 - 12,6%) for NC and 1,5% (I.C. 95%: 1,3-4,3%) for LP-LH. Finally, 1,1% (I.C. 95%: 0,4-2,6%) resulted positive for the presence of BVDV in blood. Conclusions: The results demonstrated that the agents considered largely occur in cattle population of the Marchigiana breed and must be taken in consideration approaching the diagnosis of respiratory, enteric or reproductive syndromes in these kind of farms. In addition, the results have to be assumed as a baseline for further investigations or for the definition of future intervention measures in this traditional Italian bovine breed, such as vaccination or control or eradication programmes for selected infectious agents. 705 Botulism in Cattle: how the Laboratory can Help in Diagnosing the Disease? F. Gessler , H. Boehnel 1 Miprolab GmbH, Goettingen, Germany

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For almost centuries botulism is a disease complex known to the cattle farmer. It is caused by Clostridium botulinum, an anaerobic, sporeforming bacterium. It is not the bacterium itself, but the highly potent neurotoxins, which at the end are responsible for the clinical signs observed. According to their chemical structures and pathophysiological properties seven distinct toxin groups were identified, which were named A to G. Types C and D are considered to be those toxins predominant in cattle botulism leading to flaccid muscular paralysis. The classical form of botulism is feed-poisoning, an intoxication caused by the uptake of the neurotoxin pre-formed in the feed, eg silage. In Botulinum toxico-infections, spores and/or bacteria are ingested, colonize the intestine and produce the toxins in vivo. It is the toxico-infectious botulism, which has been re-emerging over the recent years. Besides clinical, pathological and epidemiological investigations, the detection of the toxins and/or the toxigenic bacteria in the laboratory contributes to a valid diagnosis of the disease. The presentation will cover: 1. basic aspects of botulinum neurotoxins, 2. selection of samples and matrices for the laboratory investigations (eg: feed, soil, clinical samples), 3. short overview on the detection of the toxins, specific antibodies and the bacteria in the small clinical or the specialized microbiological laboratory, and 4. interpretation of the results and recommendations 706 Application of the MTT Assay for Titration of Cytopathic Bovine Viruses A. Simsek, O. Bulut Faculty of Veterinary Medicine, The Selçuk University, Department of Virology, Konya, Turkey In the current study, a colorimetric method for titration of cytopathic bovine viruses was developed. MTT colorimetric assay is based on the ability of a mitochondrial dehydrogenase enzyme from viable cells in culture to reduce the tetrazolium salts of the yellow MTT and form a blue formazan crystal. Bovine Herpesvirus Type 1 (BHV-1), Parainfluenza 3 (PI-3), Bovine Rotavirus (BRV) and Bovine Adenovirus Type 1 (BAV-1) which are cytopathic bovine viruses were propagated and titrated in Madin Darby bovine kidney (MDBK) cell line in this study. All viruses were also assayed for titration with the MTT colorimetric method. The correlation between virus titers when two assays were determined by assessing the correlation coefficient (r) with Microsoft Excel program. The MTT colorimetric assay was showed a good correlation in terms of the viable cell number and the reduction of MTT. The titers determined by the two methods (microscopic insemination for CPE and MTT microassay) were found to be highly correlated (r=0.995). In conclusion, the results of this study confirmed that the MTT assay can be applied to titrating viral systems in which the virus is able to produce CPE in vitro. In particular, the MTT method proved to be useful to estimate cell densities in virus-infected cell cultures and was more effortless than classical methods. Concurrently, the method could be computerized with an ELISA reader and facilitated the titration of large numbers of virus samples in a short time. Key words: viral titration, colorimetric assay, cattle 707 Comparative Study of Rose Bengal and Competitive ELISA Tests in Double Purpose Herds in the State of Lara Venezuela J. Maldonado , A. Kowalski , C. Villasmil , O. Verde , M. Milla , 1 1 1 J. Alvarado , F. Escalona , A. Rodriguez 1 Universidad Centroccidental Lisandro Alvarado, Barquisimeto, Venezuela 2 CIEPE, San Felipe, Venezuela 3 UNEY, San Felipe, Venezuela Brucellosis is a zoonotic disease that negatively affects milk production in Latin America. Infections with Brucella abortus bacteria observed in human populations are fundamentally caused by ingestion of nonpasteurized milk by-products. In Venezuela, the most of the milk production is obtained from crossbred double purpose herds, which are generally under deficient sanitary conditions. A significant amount of milk produced is used for production of non-pasteurized cheese for human consumption. In Venezuela, the official screening test for brucellosis is the Rose Bengal test (RBT), leaving competitive ELISA

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(cELISA) only to definitively confirm results obtained in the aforementioned test. It performed a trial study to assess the status of brucellosis in the state of Lara, Venezuela. Initially, milk ring tests were performed in 320 farms, with 30% of them testing positive. Serum samples were collected from all animals of farms that tested positive to ring test. A total of 13,351 serum samples were collected and tested with RBT and cELISA assays. Positive index observed with RBT was lower when compared to cELISA (0.51 % vs 6.36 %, respectively). Furthermore, both tests confirm that positive index in the state of Lara is very low. However, relative sensitivity observed in RBT is also very low when compared to cELISA. This apparent low sensitivity could result in a high number of false negative animals that, consequently, will not be eliminated from the herds. Thus, this would constitute a constant source of infection to other animals and people. Key words: brucellosis, Rose Bengal test, competitive ELISA 708 In vitro Evaluation of Different Antibiotic Combinations used for Treatment of Pasteurellaceae from Bovine Respiratory Disease (BRD) J. Ganiere , L. Denuault , O. Bidaud , J. Swinkels , F. Payot 1 Nantes Veterinary School, Research Unit of Infectious Diseases, Nantes, France 2 Intervet Schering-Plough, Research Unit of Infectious Diseases, Angers, France 3 Intervet Schering-Plough, Int. Marketing, Boxmeer, The Netherlands Objectives: In the field, cattle with BRD may receive different antibiotics simultaneously or successively. The objective of this study was to describe the interaction of cefquinome with other antibiotics frequently used to treat BRD. Materials and Methods: The effect of combinations of cefquinome with oxytetracyclin, marbofloxacin, florfenicol, tilmicosin and tulathromycin against the Pasteurellaceae involved in BRD: M. haemolytica, P. multocida and H. somni was tested in vitro. One strain of each bacterium isolated from clinical cases was used to assess the interaction of the combinations, determined by the Fractional Inhibitory Concentration Index (FICI) and confirmed by time-kill kinetics. Four interactions were concerned: FICI<0,5: synergy; 0,5<FICI<1:addition; 1<FICI< 2: indifference; FICI>2: antagonism. Effect of the combination cefquinometulathromycin and cefquinome-marbofloxacin was confirmed for M. haemolytica on 4 additional strains. Results: No antagonism was observed for the different combinations for all Pasteurellaceae. For P. multocida, indifference was observed between 4/5 antibiotic combinations. Addition was observed for the same combinations for H. somni. For M. haemolytica, an additive effect was observed between 4/5 combinations. This effect was marked for the combination cefquinome-tulathromycin (mean FICI: 0.8-0.2), which was close to synergy for 2/5 strains (FICI: 0.56).This additive effect was also confirmed for the combination cefquinomemarbofloxacin (additive effect for 3/5 strains and slight indifference for 2/5 strains). These interactions were confirmed by time-kill kinetics. Bactericidal activity of cefquinome combined with marbofloxacin or tulathromycin was slightly higher compared to each of these antibiotics alone, indicating an additive effect. Conclusions: No antagonism of the different combinations of cefquinome with other antibiotics was observed against Pasteurellaceae. Hence, under field conditions, it can be assumed that the efficacy of cefquinome is not decreased by the presence of other antibiotics. Instead, an additive effect was frequently observed. 709 Susceptibility of Pasteurellaceae and A. pyogenes to Antibiotics Frequently Used in Treatment of Bovine Respiratory Disease (BRD) F. Payot , J. Swinkels , M. Bonnier 1 Intervet Schering-Plough, Farm and Production Animals Business Unit, Angers, France 2 Intervet Schering Plough, Int. Marketing, Boxmeer, The Netherlands 3 Laboratoire D'Analyses d'Ille-et-Vilaine, Microbiology, Rennes, France Objectives: The objective of this report is to describe the susceptibility of the main bacterial BRD pathogens to antibiotics commonly used in France. Materials and Methods: All bacterial strains were isolated from tracheal lavage, nasal swabs or lung tissue at necropsy of clinical

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cases from West-France in 2006-2007 and analysed in the Departmental Vet Laboratory 35 (Rennes). Inhibition zones for cefquinome and 17 other antibiotics commonly used in BRD (erythromycin, lincomycin, gentamicin, ceftiofur, amoxicillin, spectinomycin, tulathromycin, tilmicosin, florfenicol, colistin, oxolinic acid, flumequin, enrofloxacin, danofloxacin, marbofloxacin, TMPS and tetracycline) were measured by the disk diffusion method for P. multocida (n=122), M. haemolytica (n=76), A. pyogenes (n=18) and H. somni (n=11). Standard critical diameters defined by CA-SFM criteria were used. Similarly, the antibiotic susceptibility of M. haemolytica, was compared over the past 3 years using 19 (isolated in 2004) and 193 strains (isolated in 2005-2006). Results: M. haemolytica was less susceptible to antibiotics than P. multocida and H. somni. However, the strains were > 99% susceptible to cefquinome, ceftiofur, gentamicin, colistin and florfenicol and > 95% susceptible to tulathromycin, enrofloxacin and marbofloxacin. All strains of A. pyogenes were 100% susceptible to amoxicillin, cefquinome, ceftiofur, florfenicol, marbofloxacin and enrofloxacin. Over the past 3 years, the susceptibility of M. haemolytica remained stable with the strains 100 % susceptible to cefquinome, ceftiofur and colistin and highly susceptible to florfenicol (99%), marbofloxacin (9698.5%) and tulathromycin (95-96%). Low susceptibility (<80%) to amoxicillin, spectinomycin, licomycin, tilmicosin, tetracyclin, oxolinic acid and flumequin was found. Conclusions: The combined susceptibility of all Pasteurellaceae and A. pyogenes exceeded 95% only for cefquinome, ceftiofur, florfenicol and marbofloxacin. Ideally, the choice of an antibiotic to treat BRD is based on the aetiology and susceptibility of the bacterial pathogens in the herd. If not available, broad-spectrum antibiotics with proven high in vitro efficacy (> 95% susceptibility) such as cefquinome, ceftiofur, florfenicol and marbofloxacin should be used. 710 Comparative Analysis of Immune Responses in Cattle Vaccinated with Brucella RB51, Rev. 1 and S19 Vaccines M. Aldubaib Qassim University, Veterinary Medicine, Buridah, Saudi Arabia Three groups of brucellosis-free adult Holstein cattle were inoculated with RB51, Rev. 1 and S19 brucella vaccines, separately. Seum samples of vaccinated cattle except those vaccinated with RB51 reacted positively with conventional Rose Bengal antigen st from the 1 week postvaccination. The reaction increased in strength rd th and reached to maximum at 3 to 8 week postvaccination and then th declined to minimum at 18 week postvaccination. Serum samples of RB51-vaccinated cattle reacted negatively with the conventional Rose Bengal antigen along the course of vaccination. Meanwhile, those RB51-vaccinated cattle sera reacted positively in slide agglutination using Rose Bengal antigen originated from RB51 strain prepared in this study. Indirect ELISA adopted in this study was used to monitor serum samples of different groups for brucella antibodies. In case of vaccination of cattle with RB51, sera of all vaccinated animals had a significant brucella antibody response at 3, 6, 9, 12, 15 and 18 weeks postvaccination. In case of vaccination of cattle with S19 and Rev. 1, sera of vaccinated animals had a significant brucella antibody response at 3, 6, 9 and 15 weeks postvaccination. The decline in the brucella antibody response was more obvious with S19 and Rev. 1 than RB51 vaccination at the 18th week postvaccination. 711 Granulomatous Meningoencephalitis Caused by Mycobacterium bovis in a Calf C. Del Fava, E. Roxo, E. Cunha, A. Cabral Instituto Biológico, Centro de Pesquisa e Desenvolvimento de Sanidade Animal, Sâo Paulo, Brazil Bovine tuberculosis is a chronic disease caused by Mycobacterium bovis, characterized by respiratory disorders, emaciation and lymphadenomegaly. Brazil began a National Program for Control and Eradication in 2001 and the notification of the disease is compulsory. Rarely, neurological signs occur associated with granulomatous meningoencephalitis or other lesions in the Central Nervous System (CNS). Neurological lesions are categorized in four patterns: meningitis (the most common), tubercles in the brain or the spinal cord, compression of the spinal cord by tuberculous spondylitis and neuritis caused by

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lesions in other organs. Young animals most frequently become infected by ingestion of infected milk. We present a case report of tuberculous meningoencephalitis in a dairy calf that was less than one year old, who presented nervous signs (recumbency, irritability and aggression, weakness, dizziness), which progressed to death in three days. At necropsy, white nodules were found in the liver, lungs and diaphragm. The clinically suspected diagnosis was rabies, but results were negative for direct immunofluorescence and intracerebral inoculation in mice. Brain samples and spinal cord were preserved in 10% formalin for histopathological analysis (HE staining). There was severe granulomatous meningomyelitis with Langhans' giant cells. In cerebral neuropile there were some granulomas with a central area of necrosis, perivascular cuffing with mononuclear cells, gliosis and hyperemia. The lung had many caseated granulomas, a typical finding in bovine tuberculosis, with central necrosis, surrounded by epithelioid macrophages, lymphocytes, plasma cells, Langhans' giant cells and fibrosis at the periphery. There was mild calcification of the pulmonary granulomas. Samples of lung and brain were decontaminated by the Petroff method and inoculated into Petragnani and Stonebrink media and M. bovis was isolated. This case report shows that bovine tuberculosis should be included in the differential diagnosis of bovine neurological disorders. According to Brazilian regulation, sanitary measures in the heard infected (test-andslaughter) should be adopted. FAPESP Key words: tuberculosis, Mycobacterium bovis, meningoencephalitis, calf 712 A Study on Diagnosis, Pathogenesis of Pseudomonas aeruginosa Isolated from Different Cases of Human and Animals in Mosul City - Iraq D. Ali , B. Ismaeel 1 College of Veterinary Medicine, University of Mosul, Iraq, Department of Microbiology, Mosul, Iraq 2 College of Veterinary Medicine, University of Dohuk, Department of Microbiology, Duhok, Iraq The current study tackles the isolation and identification of Pseudomonas aeruginosa from different human infection cases (wounds, otitis media, urine, stool, and burns) in Al-Zahrawi, Al-Salam and Ibn-Al-Atheer hospitals at Mosul city. From animals (sheep, cows, goats) samples were also collected (wounds, otitis, urine, feces and mastitis) cases for P. aeruginosa isolation and identification. The morphological, cultural and biochemical tests confirmed 43 (17.4%) isolates as P. aeruginosa from 250 human samples, and 17 (10.3%) from 165 animal samples. P. aeruginosa human and animals' isolates virulence factors represented by enzymes (protease, lipease, lecithinase, b-lactamase gelatinase, coagulase, haemolysin, toxins) and capsule production were studied; 80% of human isolates showed capsule presence, while all animal isolates (100%) contained capsule. The pigments production especially pyocyanin, which differentiates P. aeruginosa from other microorganisms, was detected in 80% of the human isolates and in 100% of the animal isolates. The study showed similarity between P. aeruginosa of human and animal isolates in their virulence factors production ability. The study results revealed that 15 pathogenic isolates from 43 tested human isolates and 10 pathogenic isolates from 17 tested animal isolates produced positive results for an exotoxin A production in rabbit skin reaction test, through inducing hypersensitivity reactions of type I (early or immediate) and of delayed type (Type IV). Baby suckling mice test and rabbit ileal loop test confirmed that both human and animal P. aeruginosa isolates produced an enterotoxin. The histopathological study revealed different changes represented by elongation and blunted villi with necrosis. 713 Bovine Paralityc Rabies: Prevention and Control in Mexico M. Manzano , S. Vázquez-Morón , R. Ramirez , M. Orozco , 2 1 J. Echevarría , A. De Vega 1 Comite de Fomento y Salud Animal, Campana de Rabia, Puebla, México, Mexico 2 Instituto de Salud Carlos III, Servicio de Microbiologia Diagnostica, Madrid, Spain 3 Unión Ganadera Regional Sierra Oriente, Gerencia, Teziutlan, Mexico Bovine paralytic rabies is a contagious and fatal disease that affects a large number of productive animal species and also humans, creating a risk both

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to human and animal health. In America this involves a serious problem. One of the determining factors to prevent cases of rabies in animals and in humans is, without any doubt, the diffusion and training on measures of prevention and control of the disease. A DVD on this kind of measures was recorded in the City of Puebla, Mexico, by the Committee of Promotion and Animal Health and the Foundation "PRODUCE", Puebla. First, the DVD has been presented all over the country, and, on a subsequent occasion, it was presented in the RITA (The international meeting on research advances in rabies control in the Americas) meetings of Brazil and Mexico, in coordination with the Panamerican Health Organization. The DVD was distributed to all American countries through these meetings.The mentioned DVD shows two aspects of this problem: in the first part a case of an outbreak of paralytic rabies is narrated in the colloquial language of the cattle farmers from the northern mountains belonging to the Puebla State; while the second part includes recommendations about the cold chain, antirabies vaccination, sending specimens to the laboratory, classification of bats and control of hematophagous bats.Furthermore, molecular studies are currently being realized in the Instituto de Salud Carlos III (Madrid, Spain) to complement the field work made in Mexico. For all the reasons exposed it is of great interest to share this experience with European Buiatric groups. 714 Effect of Cow Vaccination against BRSV and PI3 on Immune Status ante Partum and the Transfer of Colostral Antibodies to Calves B. Makoschey , R. Brunner , M. Koenig , C. Gumpert , H. Heckert 1 Intervet Schering-Plough, Int. Marketing, Boxmeer, The Netherlands 2 Intervet Schering-Plough, Businessunit Cattle, Unterschleissheim, Germany 3 Justus-Liebig-Universität Gießen, Faculty of Veterinary Medicine, Institute of Virology, Giessen, Germany 4 Freie Universität Berlin, Clinic for Ruminants and Pigs, Berlin, Germany Objectives: The effect of immunization of pregnant cows with respiratory pathogens (Bovine Respiratory Syncytial Virus (BRSV) and Parainfluenza Type 3 (PI3)) on the antibody levels in the serum and colostrum as well the transfer of the colostral antibodies to the newborn calves should be determined. Material and Methods: The study was performed on a dairy farm where the dams and their calves are housed at two different locations. A commercial vaccine containing inactivated antigens of BRSV, PI3 and Mannheimia haemolytica (M. haemolytica) (Bovipast® RSP / Bovigrip RSP plus, Intervet) was used. Twenty-six dams (n=26) were vaccinated twice (6-8 and 2-4 weeks before the anticipated date of calving). A group of 19 cows was not vaccinated with the respiratory vaccine. In addition, all cows (n=45) were routinely immunized with a neonatal calf diarrhea containing inactivated bovine rotavirus, bovine coronavirus and E. coli at 6 weeks before calving. Colostrum was collected from each cow and fed to the respective calf at a volume of at least 4 liter within the first 12 hours of life. Blood samples of the dams were taken at the time of the first vaccination and around calving. Moreover, samples of the first milkings were collected. Blood samples of the calves were taken two days after birth. Neutralising antibody titers against BRSV and PI3 were measured in the serum and colostrum samples. Results: The interpretation of results regarding BRSV was troubled by high pre-vaccination antibody levels in a certain proportion of animals. Significantly higher antibody titres against BRSV were found in dams from the vaccinate group after adjustment of measured titres by subtracting individual pre-vaccination values. Moreover, there was a correlation between the BRSV antibody titer in the cows and the titers measured in the calves. With regard to PI3, vaccination of dams resulted in a booster of neutralizing antibodies and a transfer of the antibodies to their newborn calves via colostrum regardless the pre-vaccination titer against PI3. Moreover, the PI3 antibody titers in the cows correlated well with the titers measured in the two days old newborns. Conclusions: Cow vaccination against BRSV and PI3 could be a strategy to achieve more homogeneous antibody levels in the serum of the dams and their calves and can therefore complement the active immunisation program in the calves. 715 Detection of Prevalence of Rotaviral and Corona Viral Diarrhea in Newborn Calves B. Shojaee , F. Hemmatzadeh , H. Momtaz , M. Ahmadi bozorg , 2 2 B. Khoramian , M. Eslampour

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Islamic Azad University of Shahrekord, College of Veterinary Medicine, Shahrekord, Iran 2 Facutly of Veterinary Medicine, University of Tehran, Department of Clinical Science, Tehran, Iran This study performed in order to determine of rotaviral and coronaviral diarrhea prevalance in newborn calves in Iran, in the period of 12 months (2004-2005). Antigen detection ELISA kit was used for determination of rotaviral antigen and Coronaviral antigen in fecal samples. Among 200 fecal samples 37 of them (18.5 %) were positive to Rotavirus and all of them were negative with coronavirus infection. During different seasons, the lowest rate of infection was in summer (6%) and high prevalence was in winter (48%). Frequency of infection rate of male calves was 14.28 % and among female ones was 20.13%. In dairy farms with a proper hygienic management, frequency of Rotaviral diarrhea was 5.19% and among the calves of the farms without a proper health management this frequency was about 63.04%. In different age groups, the least Rotaviral frequency was among 1-7 days age group and it was reported to be 3.17% and the most one was among the 15-21 days age group with 66.66% of infection rate. Statistical analysis of the results showed a significant association between infection, season and health management in the farm. Infectious frequency among the calves with 15-21 days of ages had a significant statistical difference with calves in other age groups. But no linear statistical correlation was founded between increase in infection rate and age of the calves. 716 Seroepidemiological Study of BVD, IBR, RSV and PI3 in Morocco Z. Zouagui, S. Alali, A. Aiyar Institut Agronomique et Vétérinaire Hassan II, Morocco, Pathologie Médicale et Chirurgicale des Ruminants, Rabat, Morocco The objective of this study is to assess the seroprevalence of four economically important viral infections among Moroccan cattle herds. Theses infections are bovine virus diarrhoea (BVD), infectious bovine rhinotracheitis (IBR), bovine respiratory syncytial virus (RSV) and parainfluenza-3 virus (PI3) infections. Our work aimed also to reveal possible relationships between the prevalence of these viruses and some epidemiological factors. For this purpose, a large epidemiological investigation has been undertaken in six regions, known to be important regarding cattle rearing (Doukkala, Haouz, Gharb, Moulouya, Tadla and Souss-Massa). 1609 blood samples, collected from non vaccinated cattle, have been analyzed by the ELISA method to asses their status concerning the four viruses. The serological results showed that the general antibody prevalence was 39%, 23%, 79% and 68% for BVD, IBR, RSV and PI3, respectively. However, regional variations of the four infections have been recorded. Moreover, the majority of the investigated farms were significantly affected by the mentioned viruses. The survey revealed also that cattle born and raised in Morocco were more affected than those imported since several years. On the other hand, IBR prevalence has been partially influenced by the method of service used. Key words: BVD, IBR, RSV, PI3, Morocco 717 Polimerase Chain Reaction Standardization for Semen, Urine and Organs of Rams Experimentally Inoculated with B. ovis C. Nozaki, N. Lira, J. Megid UNESP- School of Veterinary Medicine and Animal Sciences, Higiene Veterinária e Saúde Pública, Botucatu-SP, Brazil The objective of the study was to adapt and evaluate PCR compared with microbiological culture in the detection of B. ovis in semen, urine and organs of experimentally inoculated rams. Thirty one rams, 1-2 years old, were challenged by intrapreputial and conjuntival route with a suspension containing 2x109 CFU/mL of B. ovis. All animals inoculated were weekly submitted to clinical evaluation, as well as collection of blood for serological diagnosis, and collection of semen and urine for culture and PCR. From the 31 experimentally inoculated animals, PCR detected bacterial DNA in the semen of 24 animals, whereas culture showed the agent in 13 rams. PCR showed greater sensitivity (77%) than culture (42%). In urine samples, sensitivity of the techniques was not significantly different (48% for culture and 58% for PCR). PCR detected the presence of the agent in 25% of the organs tested while culture detected it in only 2% of the samples. Although no statistical difference was observed between the

techniques, results suggest that PCR should be used instead of microbiological culture. Key words: Brucella ovis, PCR, culture, rams, epididymitis 718 Serological Profile of Rams Experimentally Inoculated with Brucella ovis in the Different Phases of the Illness: 1 Correlation with Clinical Signals 2 Evaluation of Agar Gel Immunodifusion Test and Serum Neutralization Test C. Nozaki , N. Lira , O. Augusto Filho , H. Azevedo , L. Rodello , 2 1 S. Bicudo , J. Megid 1 Universidade Estadual Paulista, Higiene Veterinária e Saúde Pública, Botucatu-SP, Brazil 2 Universidade Estadual Paulista, Reproduction Animal, BotucatuSP, Brazil The present research had as objective to determine the serological profile after experimental inoculation of rams with B. ovis using the Agar Gel Immunodifusion test and Serum aglutination test (SAR) using the commercial kit D-TecCB. The results were correlated to the reproductive clinical signals, evaluating the phases of higher effectiveness of the serological tests and the method of clinical evaluation in the different phases of the experimental illness. Thirty one rams, 1-2 years old, were challenged by intrapreputial and 9 conjuntival route with a suspension containing 2x10 CFU/mL of B. ovis REO 198, simultaneously. After inoculation, animals were weekly submitted to clinical evaluation and blood collection for serological analysis. SAR showed a decreasing percentage of positive results, from 80% to 11%, from the second to the seventh week after inoculation. IDGA showed increasing percentages of positive results, from 45% to 63%, from the first to the third week, and then a decrease to 4% in the th 11 week. Fluctuation in titers was observed in both tests. Clinical changes began in the first week after inoculation and were associated with positive serology in the acute phase of the disease. IDGA demonstrated variable sensitivity in the different phases of the disease, showing positive animals in the chronic phase, and was preferred in routine diagnosis. Negative serological results associated to clinical alterations were characteristics of the chronic phase of the disease demonstrating the inefficacy of serology in this phase. Key words: agar gel immunodifusion test, serum aglutination test, sheep brucellosis, clinical signs, serology, experimental inoculation 719 Seroprevalence of Six Serovars of Leptospira in Human and Bovine from Toca, Boyacá (Colombia) J. López, M. Pulido, R. Andrade, A. Caycedo Universidad Pedagógica y Tecnológica de Colombia, Grupo de investigación en medicina veterinaria y zootecnia, Tunja, Colombia Objectives: Determining the presence of serovars of Leptospira in human and bovine by means of the microscopic agglutination test (MAT), and; establishing the probable source of serovars as much in the humans as in the bovine ones. Materials and methods: Location: six properties milkmaids from the rural area of Toca, Boyacá (cold climate). 36 mature people (26 women and 11 men) donated 4 ml of blood of the radial vein. 116 samples from bovine were collected, corresponding to 8 ml of blood from the caudal vein. Samples were centrifuged (3.600 rpm during five minutes) and the resulting serum froze to -20° C until its analysis with MAT. Results: Serums from people had a positivity of 29.7% (11 of 36); being from Canicola 10,8% (4 of 36); Icterohaemorragiae, Pomona and Automnalis 5,4% each one, (2 of 36 people each serovar) Hardjo 2,7% (1 of 36). The serovar Sejroë was seronegative in human. In bovine there was a 48,1% of positivity (64 of 133); being from Icterohemorragiae 11,3% (15 of 133), Pomona 13.5% (18 of 133), Hardjo 6.0% (8 of 133), Canicola 8.3% (11 of 133), Sejroë 6.8% (1 of 133); Automnalis 8.3% (11 of 133). Conclusions: Prevalence obtained of different servars of Leptospira interrogans is adjusted to data obtained by other investigators in Colombia. The results don't offer a precise interpretation about the origin of the infection with different stumps of Leptospira interrogans so much in human as in bovine. There was not statistical significance in any association, in spite of this, infection probabilities exist between human and bovine: For serovar icterohaemorragiae 2,22 times in bovine that in human; serovar pomona (2,73 times more), hardjo (2,3 times more) and automnalis (1,57 times more). This would be due to the biggest contact among bovine and elements contaminated by urine

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or secretions of wild animals, rats or dogs. Serovar canicola seems to have a protective effect in bovine (0,74 times with regard to humans), being the infection probability by means of secretions of dogs that remain in the property. From serovar Sejroë any hypothesis could not settle down, because humans went negative to the serovar. Results demonstrate the circulation of different serotypes among the animal species and they highlight the importance of carrying out more advanced epidemic studies, with the purpose of understanding the epidemiology of Leptospira in the region. Key words: leptospira, human, cattle, Toca, epidemiology 720 Comparison of Gene Detection by IS6110 and IS1081 Polymerase Chain Reaction and Isolation by LJ and BM Media in Tuberculoid Tissues of Slaughtered Cattle YS. Cho , YH. Jean , IS. Roh , I. Hwang , D. Cho , H. Nam , 1 3 1 E. Heo , JM. Kim1, HR. Han , SC. Jung 1 National Veterinary Research & Quarantine Service, Bacteriology and Parasitology Division, Anyang, Korea, South 2 National Veterinary Research & Quarantine Service, Animal Disease Diagnostic Center, Anyang, Korea, South 3 Seoul National University, College of Veterinary Medicine, Seoul, Korea, South Objectives of study: Bovine tuberculosis has been diagnosed by the examination of tuberculoid tissue as well as tuberculin skin test. Bovine tuberculosis at slaughterhouse is diagnosed by histopathological finding, polymerase chain reaction, and isolation of Mycobacterium bovis. This study provides the isolation rate of Mycobacterium bovis from tuberculoid tissues of slaughtered cattle and more efficient laboratory methods to diagnose bovine tuberculosis at slaughterhouse. Materials and Methods: One part of tuberculoid tissues (n = 89) from 39 slaughtered cattle were macerated and treated with 0.75% hexadecylpyridium chloride as decontaminant. In addition, the other part of those were treated with 10% formalin and stained with H&E and acid-fast staining kit. The macerated tissues were incubated on LJ w/o glycerol and BM media at 37 °C for 16 weeks. LJ w/ glycerol media were also incubated at 30 °C for 16 weeks for the isolation of Mycobacterium avium. DNA was extracted by GuSCN-silica and IS6110 and IS1081 primers were used for PCR. Results: M. bovis was isolated in 82 per cent of slaughtered cattle with tuberculoid tissues (32/39). Sensitivity, specificity, positive predictive value, and negative predictive value of IS6110 direct PCR (dPCR) were 69, 71, 92, and 33, and those of IS1081 dPCR were 79, 71, 79, and 29, respectively. IS6110 dPCR showed the highest positive predictive value, while IS1081 dPCR showed the highest sensitivity. The result by dPCR accorded with that by H&E staining, but the tuberculoid tissues were not always detected acid-fast bacilli. M. bovis isolation rate by BM (77%; 57/74) was higher than that by LJ w/o glycerol (72%; 63/87). Eighty seven per cent of the isolates (72/83) were IS6110 positive, and 86 per cent of those (67/78) IS1081 positive. At 30 °C on LJ w/ glycerol, 16 mycobacteria-like bacilli were isolated from 90 samples, and 14 isolates of those were identified as Mycobacterium bovis by IS6110 and IS1081 PCR. Conclusions: The results from IS6110 and IS1081 dPCR were not always accorded each other. Therefore, the application of both IS6110 and IS1081 PCR could increase the diagnostic accuracy of bovine tuberculosis. The isolation rate of BM media showed higher than that of LJ media. The isolates with only IS6110 or IS1081 gene would be further analyzed to examine new genotype of M. bovis in the future. Key words: Mycobacterium bovis, bovine tuberculosis, IS6110, IS1081, LJ media, BM media 721 Outbreak of Acute BVD in Brazilian Beef Cattle: Clinicopathological Findings and Molecular Characterization of a BVDV Strain Subtype 1b M. Lunardi, S. Headley, J. Lisboa, R. Otonel, A. Alfieri, A. Alfieri Universidade Estadual de Londrina, Departamento de Medicina Veterinária Preventiva, Londrina, Brazil When first described, Bovine Viral Diarrhea (BVD) was characterized as an acute transmissible disease associated with severe leucopenia, high fever, depression, diarrhea, gastrointestinal erosions, and hemorrhages. Recently the severe acute form has been related only to some hypervirulent BVDV-2 strains. This article reports the detection of BVDV-1b associated with an acute and fatal outbreak of BVD in a

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Brazilian herd. Depression, anorexia, watery diarrhea, sialorrhea, and weakness were observed in six steers. The affected animals died 24 hours to 15 days after the first manifestation of infection was observed. One of these animals was evaluated for laboratorial, clinical, and pathological alterations. Blood samples were collected ante mortem by routine procedures. Selected tissues were fixed in 10% neutral buffered formalin solution and processed for histopathological evaluation. RNA extraction was performed following silica/guanidine isothiocyanate method. The RT-PCR assay was carried out using a set of primers (103/372) designed from the 5-UTR region of the BVDV genome. The identities of the RT-PCR products were confirmed by direct sequencing of the amplicon from one clinical specimen. Sequences were examined with the software PHRED for quality analysis and the consensus sequence was determined using the software CAP3. The alignment was obtained by BioEdit software. A neighbour-joining phylogenetic tree was constructed using the MEGA program. Laboratory findings were non-specific; clinically, the animal was weak, with dehydration and erosive oral lesions. Pathological alterations were predominant at the tongue, esophagus, and rumen. The RT-PCR assay identified BVDV in all clinical samples analyzed. The sequence obtained has been deposited in GenBank (accession number EF406123). Phylogenetic analysis of BVDV derived from lymph node revealed that this strain was clustered with other BVDV subtype 1b isolates, sharing high similarity (96.7%) with the representative subtype 1b isolate Osloss. The real role played by BVDV-1b in acute diseases and the frequency of this subtype in the epizootology of acute and severe BVD are not totally elucidated. Molecular characterization is the only definitive method to identify the BVDV genotype involved in acute outbreaks of severe BVD, since both clinical presentation and pathologic findings can be similar to BVDV-2 infection. Financial support: CNPq, CAPES, FINEP and FAP/PR Key words: cattle, BVD, BVDV, 5-UTR 722 Winter Dysentery by Bovine Coronavirus Infection in a Tropical Country E. Takiuchi, A. Barry, D. Stipp, R. Otonel, A. Alfieri, A. Alfieri Universidade Estadual de Londrina, Departamento de Medicina Veterinária Preventiva, Londrina, Brazil Winter Dysentery (WD) is an acute diarrhea caused by Bovine Coronavirus (BCoV) that affects adult cattle during the winter with high morbidity and low mortality rates. Low temperature and UV light intensity during the winter lead to high levels of environmental contamination by BCoV. The WD occurrence in dairy herds results in economic losses caused by severe reduction in milk production. Association between BCoV infection and WD was reported in the USA, Europe, Japan and Canada, being sporadic in tropical areas. This study reports, the detection of BCoV in an adult cow during an outbreak of acute diarrhea in Brazil. The outbreak occurred during June, in Ponta Grossa city, Paraná, Brazil and affect 138 from 154 lactating cows. Three animals died. Most cows had bloody and watery diarrhea (three to four days), and milk production dropped from 37 to 26 liters/cow/day. Fecal sample was collected directly from large intestine of a necropsied cow. The sample was negative to C. perfringens type C, Eimeria spp, Cryptosporidium spp, enteroxigenic E. coli, rotavirus and bovine viral diarrhea virus. Liquid feces suspension at 50% (v/v) was prepared in PBS. RNA extraction was performed with phenol/chloroform/isoamyl alcohol and silica/guanidinium isothiocyanate methods. PCR and semi-nested assay were performed according to Takiuchi et al. (2006) with primers BCoV1/BCoV2 and BCoV3/BCoV2, respectively, with a predicted 251 bp product (N gene). The products were analyzed by electrophoresis in ethidium bromide stained 2% agarose gel, under UV light. BCoV amplicons from semi-nested PCR were purified, sequenced, and similarity checked in BLAST software confirming that the 251 bp amplicon was from a BCoV strain. This result suggests the role of BCoV in the etiology of WD in Brazilian dairy herds. High incidence of WD in cold months is notified in regions with well-defined seasons. The present case indicates that WD also occurs in tropical areas where temperatures are higher. The location of city in the South of Brazil, with temperature normally lower, and the season (winter) in which the outbreak occurred, perhaps have created favorable conditions to virus stability facilitating dissemination among confined cattle. In conclusion, while there are some studies associating BCoV with outbreaks of WD in other countries, there are a few reports in South America, Brazil.

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Financial support: CNPq, CAPES, FINEP and FAP/PR Key words: dairy herd, winter dysentery, Bovine cCoronavirus 723 Frequency of BCoV in Neonatal Calf Diarrhea of Brazilian Herds D. Stipp, A. Barry, E. Takiuchi, R. Otonel, A. Alfieri, A. Alfieri Universidade Estadual de Londrina, Departamento de Medicina Veterinária Preventiva, Londrina, Brazil Neonatal diarrhea is one of the main causes of calf morbidity and mortality that determine economical losses to cattle herds worldwide. Several environmental, managemental, nutritional and physiological factors may occur either alone or in synergy with the different infectious agents. The rotavirus is considered the main etiological agent of viral gastroenteritis in mammals. However, other viruses have been identified in bovine neonatal enteric infections such as the Bovine Coronavirus (BCoV). Epidemiological data on the frequency of BCoV occurring in South America livestock farms is very limited. Besides, the applicability of a sensitive and specific diagnostic technique such as RT-PCR is fundamental to not underestimating the real prevalence of BCoV. We evaluate the BCoV frequency in diarrheic fecal samples from calves up to 60 days old from beef and dairy cattle herds by a semi nested-PCR assay (SN-PCR). The animals were from four Brazilian States: Sao Paulo (SP), Paraná (PR), Minas Gerais (MG) and Mato Grosso (MT). The samples were prepared as either as 10% (w/v) suspension of semi-solid feces or as a 50% (v/v) suspension of liquid feces in PBS pH 7.2. For RNA extraction a combination of phenol/ chloroform/ isoamyl alcohol and silica/isothiocyanate methods was performed. The primers were designed from the highly conserved region of the nucleocapside gene of the Mebus strain. For the first round of amplification the primers BCoV1 and BCoV2 were used and designed to amplify a fragment of 454 bp. For the second round the primers BCoV3 and BCoV2 were used that was expected to amplify a product of 251 bp. Among the 262 diarrheic fecal samples included in this study, BCoV was detected in 15.6% (41/262) of the samples of which 37.5% (6/16), 11.1% (6/54), 12.2% (22/181) and 63.6% (7/11) were from SP, PR, MT and MG, respectively. The BCoV infection was more frequent (24.1%) in diarrheic fecal samples from calves between the third and fourth weeks of life. The rate of positive results confirms that the BCoV is an important etiological agent of the acute neonatal diarrhea in Brazilian dairy and beef cattle herds. The SN-PCR used in this study demonstrated to be an accurate tool for the routine detection and diagnosis of BCoV in feces from naturally infected calves. Financial support: CNPq, CAPES, FINEP and FAP/PR Key words: dairy herd, winter dysentery, Bovine Coronavirus 724 Assessment of BoHV-5 A663 Growth Kinetic and Comparison with BoHV Reference Strains M. Ladelfa , F. Kotsias , M. Del Medico Zajac , S. Romera 1 CONICET, Buenos Aires, Argentina 2 Instituto Nacional de Tecnología Agropecuaria-INTA, Buenos Aires, Argentina Bovine Herpesviruses 1 and 5 (BoHV-1, BoHV-5) are two closely related alphaherpesvirus which co-circulate in Argentina. BoHV-5 causes fatal encephalitis while BoHV-1 is associated with abortions, respiratory and genital disease. BoHV-5 strains are divided into three subtypes. a (N569 strain), b (A663 strain) and non a/b. A663 was isolated in our country and is considered a reference strain. The aim of this work is to establish the growth kinetic of the BoHV-5 A663 and compare it with BoHV-5 N569 and BoHV-1 LA strains. For that purpose, one step and multiple steps kinetics were performed. MDBK cells were infected with the strains mentioned above at a MOI of 0.1 and 5, virus adsorption was synchronized and then incubated at 37°C. After that, extracellular virus was inactivated. At 0, 3, 6, 12, 24, 36 and 48 hours post infection (hpi) and 0, 3, 6, 12, 15, 18 and 24 hpi, for MOI 0.1 and 5, respectively, intra and extracellular fractions were obtained, as well as total fraction, and titrated twice on MDBK in duplicate.The growth kinetics curves showed similar profile among LA, N569 and A663 strains. Virus titre reached at the end of the assay 6.7 6.8 16.7 7.4 was 10 , 10 and 10 TCID50/ml at MOI 5 and 10 , 107 and 7.2 10 TCID50/ml at MOI 0.1 for LA, N569 and A663, respectively. Maximum viral titre was obtained at 18, 24 and 12 hpi at MOI 5 and 48, 36 and 48 hpi at MOI 0.1 for LA, N569 and A663, respectively. The A663 extracellular fraction showed a remarkable difference at

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MOI 0.1 in comparison with LA and N569, rising lower titres at all the times assayed. Virus release percentage was similar among the strains at both MOI and the maximum figures were in all cases near to 50 %. Finally, the eclipse period was analised. At MOI 5, it finished at 3, 6 and 12 hpi for LA, A663 and N569 respectively, and intracellular virus was detected at 3 hpi in all cases. At MOI 0.1, the eclipse period finished at 12 hpi for all three viruses. Intracellular virus was detected at 3 hpi for LA and at 6 hpi for A663 and N569. These results provide relevant information about virological aspects of BoHV-5 A663 strain, for which there is scarce information available, and allow its comparison with other related viruses. Concerning this, although A663 and N569 growth kinetics are similar, significant differences in the extracellular fraction titres were observed at low MOI, while LA strain behaves like N569. Besides, these results constitute an important tool for future in vitro experiment design. Key words: BoHV-5 A663, BoHV-5 N569, BoHV-1 LA, growth kinetic 725 A BoHV-1 gE Gene Deleted Live Vaccine is not Excreted and does not Establish a Latent Infection in Trigeminal Ganglia after Infection in Calves F. Kotsias , M. Del Médico Zajac , M. Ladelfa , S. Romera 1 CONICET, Buenos Aires, Argentina 2 Instituto Nacional de Tecnología Agropecuaria-INTA, Buenos Aires, Argentina A recombinant Bovine Herpesvirus 1 (BoHV-1) strain lacking the glycoprotein E gene (gE gene) (BoHV-1gE,gal) has been previously developed in our laboratory. Although its protective, immunogenic and safety properties have already been assessed when used as a live vaccine, some features concerning its pathogenesis, different from those reported for other gE gene deleted BoHV-1 strains, remained unstudied. In order to characterize the BoHV-1gE,gal strain in its natural host so as to understand its unique biological behavior, we now present the preliminary results concerning in vivo properties of this live vaccine candidate virus. Six calves were housed in an isolation stable, and intranasally inoculated with a high dose of the BoHV-1gE,gal virus. Animals were monitored until days 9, 15, 21 and 42 post-infection (pi) when they were euthanized. The two remaining calves were treated with dexametasone in order to reactivate latent virus, and two weeks after they were euthananized at 65 and 87 days pi. Nasal samples were tested for infectious virus, viral DNA and IgA antibodies while IgG antibodies were evaluated in serum samples. Trigeminal ganglia (TG) tissue samples were examined for viral DNA. Animals showed no clinical illness and rectal temperature remained normal throughout the experience. Infectious virus could not be isolated from nasal swabs at any day pi, however viral DNA was detected by PCR at days 4 and 7 pi. Serum IgG antibodies were found in all animals except for calf euthanized on day 9 pi. Moreover, presence of IgA antibodies in nasal secretions were detected on days 7, 9 and 12 pi in all six calves. After dexametasone treatment, no infectious virus could be detected in nasal swabs nor clinical manifestations observed. PCR from TG were negative for all the animals under study. The BoHV-1gE,gal strain used in this experience proved to be avirulent in calves, since no clinical signs were observed nor virus shedding detected. Results from serum antibodies taken together with the presence of IgA in nasal secretions indicate that calves elicited a detectable immune response. Absence of viral DNA in all TG and failure to reactivate both infected animals suggest that this recombinant virus does not establish a biologically important latent infection. This is the first report on a gE gene deleted BoHV-1 virus not excreted after infection nor detected in TG. For all this, BoHV-1gE,gal strain constitutes an epidemiologically safe vaccine candidate. Key words: BoHV-1, gE gene deleted, live vaccine, cattle, latency 726 Serological Survey of Coxiella burnetii Infection in Herds of Caltanissetta District A. Stancanelli , D. Macr , C. Di Bella , A. Torina , S. Agnello 1 IZS Sicilia, Area Caltanissetta, Caltanissetta, Italy 2 IZS Sicilia, Palermo, Italy

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Q fever is a worldwide zoonosis caused by a gram-negative obligate intracellular bacterium, Coxiella burnetii. Although Q fever is present virtually in all "animal kingdoms", including arthropods, the disease affects mostly humans, cattle, sheep and goats. The aim of this study

108 XXV. Jubilee World Buiatrics Congress 2008

was to evaluate, by serology, prevalence of Coxiella burnetii infection in herds of Caltanissetta district. 124 of 289 herds were selected, with stratified random sampling, distributed in four parts of Caltanissetta district. Particularly 71 herds were chosen from Caltanissetta territory, 9 from Gela, 29 from Mussomeli and 15 from San Cataldo. 1711 sera were collected to test the presence of specific antibodies to C. burnetii. Sera were tested by a commercial available ELISA (Chekit Q-fever; IDEXX Laboratoires). Results showed serological positivity in 60/124 (48,4%) herds with 118 positive sera/1711 (6,9%). 727 Apparent Prevalence of Antibodies to Coxiella burnetii (Q Fever) in Bulk Tank Milk of Dairy Herds in the Walloon Region of Belgium G. Czaplicki , J. Houtain , C. Manteca , C. Saegerman 1 ARSIA, Diagnostic Center for Animal Diseases, Loncin, Belgium 2 CEVA santé Animale, Diagnostic Center for Animal Diseases, Libourne, France 3 University of Liege - Faculty of Veterinary Medicine, Epidemiology and Risk Analysis applied to Veterinary sciences, Faculty of, Liege, Belgium Objectives: Q Fever is a zoonotic disease caused by Coxiella burnetii, an obligate intracellular bacteria. Ruminants (sheep, goats and cattle) remain the major natural reservoir for human infection. The aim of the present study was on the one hand, to evaluate the apparent prevalence in the Walloon dairy herds and on the other hand, to identify herd level risk factors and clinical signs statistically associated with seropositivity in the population of randomly selected Walloon dairy cattle. Material and methods: 566 herds were randomly selected according to a classical methodology (Jenicek and Cléroux, 1987). Herd level predictors data were collected from a questionnaire on farm demographics, management practices and observed clinical signs during the twelve last months. Two hundred and six farms responded on a voluntary basis to this questionnaire and submitted a sample of bulk tank milk sampled in February 2006. Bulk milks were centrifuged and stored at 20°C until testing for antibodies to Coxiella burnetii with a commercial Indirect Elisa test (LSI). Apparent prevalence was estimated with a ninety-five percent confidence intervals (95% CI) assuming a binomial exact distribution. Statistical analysis of the data was done using a chi square test and the tendency of each parameter to become a risk factor was evaluated by an odds ratio calculation with 95% confidence intervals (logarithmic approximation); a P value < 0.05 was considered significant. Results: Global apparent prevalence: 54.9 % (95% CI : 47.8-61.8%) of herds show serological evidence of Coxiella burnetii infection. Clinical signs as abortion in heifers and in cows, stillborn or weak calves are statistically associated with seropositivity. Risk factors associated with seropositivity are free stalling and watering with well water while blocked stalling, watering with tap water and disinfection of sheds act as protectors of infection. Conclusions: This first report on Q Fever infection prevalence in Walloon dairy herds indicates a quite high level of infection. Seropositivity at herd level does not mean shedding of Coxiella burnetii, but identifies herds where at least 10% of lactating animals are sero-positive to C. burnetii. A significant association exists between sero-positivity and typical clinical signs of Q Fever observed in those herds. Some risk factors and some protectors are identified on a statistical basis. Key words: bovine, Q Fever, serology, prevalence 728 Identification of Bovine Viral Diarrhoea Virus Mimotopes/Epitopes through Selection from a Peptide 12-mer Phage Display Library A. Zamit , M. Ostrowski , N. Fondevila , O. Lopez , A. Bratanich , 1 S. Romera 1 Instituto Nacional de Tecnología Agropecuaria-INTA, BUENOS AIRES, Argentina 2 Institut Curie, Paris, France 3 Northern Michigan University, Marquette, United States 4 Facultad de Cs Veterinarias-UBA, BUENOS AIRES, Argentina Bovine Viral Diarrhoea Virus (BVDV) is a worldwide distributed pestivirus that causes important economical losses to cattle industry. Argentine is no exception with a population of 50 million bovines and high seroprevalences up to 90%. The aim of our study is epitope mapping of immunodominant E2 protein of BVDV using a peptide 12mer phage display library.

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Materials and Methods: A peptide 12-mer phage display commercial library was depleted of mimotopes resembling BVDV proteins except those corresponding to E2, by using serum from a calf immunized against recombinant BVDV-E2BDV. Then, a positive selection was performed with affinity purified antibodies against BVDV strain NADL. Individual phage clones were amplified and displayed peptide sequences analyzed using CLUSTALX with Tudos matrix. Four peptides were designed for immunization purposes and sent to synthesize. Two bovines and 4 groups of guinea pigs were vaccinated with peptides coupled to KLH. Also, phages displaying chosen peptides were amplified and used to immunize mice. Sera were analyzed by peptide-ELISA, phageELISA, indirect ELISA and seroneutralization. Results: Enrichment of specific clones was achieved in 4 rounds of panning and 30 clones were individually amplified. Multiple sequence alignment analysis with modified parameters and substitution matrix allowed a better clustering of peptides. Considering clusters, peptide frequency and hydrophobicity and aminoacids representativity in library, 4 rationally designed peptides with immunogenic properties were obtained. Satisfactory antibody responses against peptides were observed in mice immunized with phages, and bovines and guinea pigs vaccinated with synthetic peptides. Also, antibodies produced in mice recognized synthetic peptides. Sera from immunized animals were either citotoxic or not neutralizing. Indirect ELISA showed high background and was not conclusive. Conclusions: It was possible to obtain immunogenic peptides resembling epitopes of BVDV using a peptide 12-mer phage display library as mapping strategy. Epitope mapping is imperative for improving local vaccines design and control. Key words: BVDV, mimotope, phage display, E2 729 Amplification of Inv Gene of Salmonalla Serotypes by Polymerase Chain Reaction (PCR) as a Specific Method of Detection of Salmonellae T. Zahraei Salehi , M. Mahzonieh , A. Ashrafi 1 Faculty of Veterinary Medicine, University of Tehran, Microbiology, Tehran, Iran 2 Faculty of Veterinary, Pathobiology, Shahrekord, Iran Background: Detection of inv gene in Salmonella serotypes by PCR. Materials and Methods: Sixty Salmonella strains were isolated from animals and human sources. In this research, at first 60 isolated Salmonella from animals and human were tested by biochemical tests such as carbohydrate utilization tests and urease test and then were serogrouped by Salmonella O anisera. DNA of isolated Salmonella were extracted by Holmes and Quigley method. Two primers (St139 and St141) and PCR reagents from fermantas company were used for amplication of inv gene. PCR reaction was carried out in Master cycle (Eppendorf). The PCR product were loaded into 1.2% agarose gel and electrophoresed for 60 minutes at 120 V. Results: All isolates showed biochemical properties of Salmonellae. In PCR assay, target gene (invA gene) with 284 bp size were observed in all the strains, which is corresponded with size of band of positive control and DNA marker. So, all of the strains in this survey had invA gene. Conclution: According to the results of this study PCR method based on inv gene is useful to rapid identification of Salmonella serotypes. Key words: Salmonella, invA gene, PCR 730 Toxoplasmosis in a Commercial Dairy Farm in Turkey S. Gazyagci , C. Babur , S. Kilic , B. Celebi , A. Gazyagci , 1 B. Yagci 1 Faculty of Veterinary in Kirikkale University, Department of Internal Medicine, Kirikkale, Turkey 2 National Research Center, Epidemiology and Public Health, Ankara, Turkey Blood samples were taken from 742 dairy cattles in a commercial farm. Antibody titers were determined against Toxoplasma gondii by SabineFeldman Dye method. Serological examination revealed 640 cattle to be seropositive. Among these sera the numbers of seropositive samples at certain dilution steps were as follows: 268 at 1/16, 184 at 1/64, 112 at 1/256 and 76 at 1/1024. Three cats could circulate in the warehouse in the farm. All sera were tested with Sabin-Feldman Dye test for Toxoplasma gondii spesific antibodies. Antibody titers were 1/256 all of them.This study investigates the seroprevalence of Toxoplasma gondii in dairy cattle from a commercial farm in Turkey. Key words: cattle, Sabin Feldman dye test, Toxoplasma gondii

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Infectious and Zoonotic Deseases (Public Health) 109

731 Detection and Investigation on the within-farm Transmission of Bovine Herpesvirus-1 in Farms Located in the Trace District of Turkey A. Unver , N. Turan , H. Yilmaz 1 Farmamag, Firuzkoy Yolu, Baglar Sokak, Avcilar, Istanbul, Turkey 2 University of Istanbul, Veterinary Faculty, Department of Virology, Avcilar, Istanbul, Turkey The aim of this study was to detect BHV-1 infection and to investigate within-farm transmission in farms located in the Trace district of Turkey. For this, 3 dairy farms having the animals over 20 were selected from the Marmara region. Two visits were made to these farms 6-7 months intervals. At each visit, the nasal swabs and blood from each animal and the milk sample from milking cows were taken. Blood sera were analysed by ELISA (IDEXX, gB blocking) for the presence of antibodies to BHV-1. Whereas the PCR was used to detect BHV-1 DNA in the nasal swabs and milk samples. In samples taken during the first visit, antibodies to BHV-1 were detected in 13 of 89 animals in 3 farms. 4 of 27 (14.8%) animals, 6 of 21 (28.7%) animals and 3 of 41 (7.3%) animals were found to be seropositive in farms-1, 2 and 3, respectively. BHV-1 DNA was detected in nasal swabs of 2 animals and milk sample of one animal in farm-1, milk sample of 1 animal in farm-2 and nasal swabs of 6 animals in farm-3. In samples taken during the second visit, antibodies to BHV-1 were detected in 18 of 71 animals in 3 farms. 6 of 24 (25%) animals, 6 of 17 (35.2%) animals and 6 of 30 (20%) animals were found to be seropositive in farms-1, 2 and 3, respectively. BHV-1 DNA was detected in nasal swabs of 3 animals in farm-1, nasal swabs of 2 animals and milk sample of 1 animal in farm2. No BHV-1 DNA was detected in the samples taken from the farm-3. In conclusion, the BHV-1 infection was detected by ELISA and PCR in all 3 farms. Some of the animals found to be seronegative by ELISA were positive on PCR and these animals might be either seronegative carriers or acutely infected. Therefore, it would be better to check the animals in the farms by both test ELISA and PCR. Also care must be taken to test new animals and separating the youngs from the adults. Key words: bovine, herpesvirus-1, transmission, PCR, ELISA, Turkey 733 Outbreak of Foot-and-Mouth Disease with High Fatality Rate in Young Lambs in Tabriz - Iran G. Noursaadat , M. Tooloei , F. Rezazadeh 1 Large Animal Clinician, Tabriz, Iran 2 University of Tabriz, Faculty of Veterinary Medicine, Department of Clinical Science, Tabriz, Iran Foot-and-mouth disease (FMD) is caused by an aphthovirus (family picornaviridae) and affects all cloven foot animals. It is endemic in Africa, Asia, South America and parts of Europe. The disease in adult sheep usually causes milder clinical signs than in cattle or pigs, and is often subtle enough to go undiagnosed. In contrast, FMD in lambs has been reported to cause high mortality during field outbreaks. From January to April 2007, five to fifty days-old lambs of the approximately all of the flocks pertaining to the east region of Tabriz affected with a highly fatal disease. The morbidity and mortality rates in lambs of each flock in this outbreak were about 10-50 and 70-80%, respectively. Adult sheep had also been affected, but the disease in sheep was very mild and with less frequent and extent than lambs. All affected lambs were clinically examined. Observed clinical signs include: severe tachycardia, high fever (up to 42°C), tachypnea, dispnea, salivation and drooling, development of vesicular lesions in interdigital and coronary region of all four foot and in mouth especially over the tongue and lips as well as lameness. Most of the affected sheep were also suffering from lameness. At the necropsy of at least one hundred died lambs, hyperemia and hepatization of lungs especially in the apical and ventral lobes, swelling and enlargement of mediastinal lymph nodes, petechiation of spleen and pericardial fats, pericarditis and pericardial effusion, epicardial and myocardial hemorrhages with pale and necrotic areas or the typical tiger heart appearance and hepatomegaly were observed. Histopathologic examination revealed severe hyaline degeneration and necrosis of myocardial fibers and marked interstitial infiltration by mononuclear cells as well as congestion and periacinar hepatocellular necrosis in the liver. The diagnosis of Foot-and-mouth disease was confirmed by the evaluation of the clinical and necropsy findings as well as histopathologic changes. Mortality rates in the affected flocks mainly decreased through vaccination of the all affected and unaffected lambs and sheep even one-day old lambs and also

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administration of some antibiotics such as Trimetoprim, Sulfadiazine and Gentamycin for 72 hours. Key words: foot-and-mouth disease, lambs, Tabriz 734 Macroscopic and Microscopic Studies on Pathological Lesions of Deleted Ovine Livers in Kermanshah Slaughterhouse Dr. Bahiraie , Dr. Pouyanmehr , M. Razmju 1 Razi University,Faculty of veterinary Medicine, Department of Animal Health and Anatomy, Kermanshah, Iran 2 Faculty of Veterinary Medicine, Meat Hygiene Department, Razi University Objective study: The pathological survey on liver which had been recorded from sheep carcasses in slaughterhouse of Kermanshah. The indication of lesions which causes liver deletion during slaughtering process. Material and Methods: The macroscopic and microscopic examination carried out on 160 sheep livers which were recorded in city slaughterhouse. The samples were collected randomly during 1 year (2006). Macroscopic survey was done on each recorded liver. Preparation for microscopic examinations carried out on macroscopic lesions samples by use of routine histological methods. Dimension of samples were 5x, 5x, 5x millimeters. The technique for preparation of tissue sections by tissue processor (Histokinettte) equipment was including: 1-Fixation (buffere formalin 10%, 2-3 days), Washing (water 4-5 hours), dehydration (using of ascending concentrations of ethylic alcohols70%, 80%, 90%, 100%, 100%), Clearing(Xylene), Impregnation (paraffin 56 °C-68 °C), Blocking (paraffin Merck), Sectioning (Microtome).The thickness of prepared sections for study was 5. Staining was Haematoxylin & Eosin (H&E). Statistical analysis was done by SPSS. Results: The highest macroscopic lesion was cirrhosis and then lesions due to hydatid cysts and Fascioulosis. In microscopic study, the highest diagnosed lesions was Bacterial hepatitis 20 cases (12.5 %) and then pathological lesions due to Fascioulosis 18 cases (11.35%), hepatic cirrhosis16 cases(%10), Chronic bill ducts inflammation 16 cases(10%), Telangiectasia16 cases (10%), Dicroselliousis 14 cases (8.75%), Chronic hepatitis 14 cases (8.75%), Hydatid cyst 10 cases (6.25%) , Fatty Liver 8 cases (5%),Hemorrhagic Pies hepatitis 8 cases (5%), Blood congestion in sinusoidal perihepatitis and Eosinophilic parasitical hepatitis each of them 6 cases (3.75%), Cholangitis and fibroma 4 cases (2.5%). In this research, centrilobular Necrosis and cholangiocarsinoma which reported in another spices was none found (0.0%). Conclusion: The effects of contamination to parasitical agents identified as the most causes of deletion of sheep liver directly or indirectly. Even thought the cirrhosis was major factor in macroscopic judgment for deletion, but the parasitical agents are the essential factors for deletion of lives. Key words: sheep health, meat hygiene, pathological lesions of liver, judgment of carcasses 736 Antibacterial Activity of Aqueous Methanol Extracts of Zingiber Officianale, Curcuma Longa, Vernonia Anthelmintica, Acacia Nilotica and Melia Azedarach Z. Abbas , M. Aslam , M. Arshad , Z. Iqba , M. Ashraf 1 University of Agriculture, Department of Parasitology, Faisalabad38040, Pakistan 2 University of Agriculture, Department of Microbiology, Faisalabad, Pakistan Introduction: The development of drug resistance as well as appearance of undesirable side effects of certain antibiotics has led to the search of new antibacterial agents in particular from medicinal plants. Therefore, the present study was conducted to assess the antibacterial potential of aqueous methanol extract of Zingiber officianale, Curcuma longa, Vernonia anthelmintica, Acacia nilotica and Melia azedarach against Staphylococcus aureus, Escherichia coli and Bacillus subtilis. Materials and Methods: The antibacterial activity of these extracts was tested using disc diffusion method. Filter paper discs were soaked with 15ul of plant extracts stock solution and were allowed to dry at room temperature for 10-15 min. A bacterial culture was uniformly spread on the surface of Mueller Hinton agar plates using a sterile swab. Each disc was pressed down uniformly against the surface of agar and then examined for any inhibition zone if present. The test was

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110 XXV. Jubilee World Buiatrics Congress 2008

repeated three times to ensure reliability.The MIC was also determined by using agar dilution method and agar well diffusion method. Results: Aqueous methnol extract of Acacia nilotica was found effective against Staphylococcus aureus and Bacillus subtilis. The other four extracts did not show any inhibitory effect against any of the three bacterial species. The Minimum Inhibitory Concentration (MIC) values of aqueous methanol extract of Acacia nilotica determined by agar dilution method and agar well diffusion method against Staphylococcus aureus were 0.78 mg/ml and 1.56 mg/ml, respectively. In case of Bacillus subtilis, MIC value of extract of Acacia nilotica as determined by agar dilution method and agar well dilution method were 1.56 mg/ml and 3.125 mg/ml, respectively.

Discussion: Traditional medical treatments in daily life are now being used with empiric methods. According to a study performed by the WHO based on publications on pharmacopoeias and medical plants in 91 countries, the number of medicinal plants is nearly 20,000. However, the botanical source, provenance of the plant, harvest time or development stage, extraction technique, plant material being fresh and dried, test micro-organisms, location, seasonality and antimicrobial methodology used, are the factors that influence the antimicrobial activity and, therefore must be taken into account whenever antimicrobial assays of the extracts of plants are performed. Key words: antibacterial activity: Zingiber officianale, Curcuma longa, Vernonia anthelmintica, Acacia nilotica and Melia azedarach

Sheep, Goat and Buffaloes

17 Effect of Vaccination against Enterotoxaemia of Pregnant Ewes on Antitoxin Alpha and Epsilon Antibodies in Colostrum, Milk and Lambs after Colostrum Intake M. Socié-Jacob , B. Bolkaerts , L. Wiggers , J. Van de Ven , 1 N. Kirschvink 1 University of Namur, Veterinary Department, Animal Physiology, Namur, Belgium 2 Schering-Plough, Animal Health, Brussels, Belgium Enterotoxaemia caused by C. perfringens belongs to the pathologies in lambs that often causes mortality. Protection against enterotoxaemia can be provided by vaccination, but the development of a specific immune response by newborn lambs lasts 4-6 wk. The aim of this study was to test whether vaccination of ewes against enterotoxaemia would provide specific colostral- and milkassociated anti-toxin and antibodies to suckling lambs. Ewes were divided in a unvaccinated control group (C:n=90) and in a vaccination group (V:n=130) that underwent vaccination with a 10valence vaccine 6 (T-6) and 2 wk prior to lambing. Blood of ewes (C:n=36;V:n=53) was sampled before vaccination and at lambing (T0). Colostrum was sampled at lambing (C:n=90;V:n=130) and blood was sampled in lambs 24h after birth (C:n=60;V:n=90). Ewes'milk (C:n=60;V:n=110) was sampled at weaning (9-12 wks after lambing). Samples were analysed for antitoxin and antibody (Ab) levels by use of an EIA. Results: Data are shown as mean SE and are expressed in arbitrary units of Ab. Ewes serum T-6 C (Ab) (Ab) V (Ab) (Ab) 20 12 7 13 51 12 37 11 Ewes serum T0 25 19 8 11 887 883* 1721 2929* Colostrum Milk at Lambs T0 weaning serum T24 22 16 9 15 583 49* 2075 305* 12 5 44 83 23* 50 10* 47 6 28 58 2935 603*A 10008 2381*A

a 1 1 1 2 3

Wageningen University and Research Centre, Quantitative Veterinary Epidemiology, Wageningen, Netherlands

Objective: To know whether FMDV can be transmitted in a susceptible sheep population and if a single vaccination can reduce FMDV transmission sufficiently to stop an epidemic (R < 1), the aim of this study was to quantify the effect of vaccination on the virus transmission within groups of lambs. Material and Methods: In 6 separate groups of 4 non-vaccinated, and in 6 groups of 4 single-vaccinated lambs were included. Two lambs per group were inoculated with FMDV and the other two were exposed by direct contact to the inoculated lambs. Daily observations of clinical signs, and serological and virological tests were performed. We estimated the transmission rate (ß) using a generalized linear mixed effect model, the reproduction ratio R using the final size method, and the duration of the infectious period using parametric (exponential) survival analysis. Results: In the non-vaccinated lambs, clinical signs were mild and rare. Virus was excreted before and after clinical signs were observed. Virus was isolated even from lambs without clinical signs. In the nonvaccinated groups, the transmission rate ß was 0.105 (0.044;0.253) per day, and the reproduction ratio Rnv 1.14 (0.3;3.3). The duration of the infectious period was estimated to be 21.11 (10.6;42.1) days. The ß and duration in the vaccinated lambs could not be determined due to limited observations of virus excretion. The reproduction ratio Rv was 0.22 (0.01;1.78). The non-vaccinated lambs had a significantly higher mean daily virus excretion and number of days virus is excreted compared to the vaccinated groups of lambs. Conclusion: Vaccination did have a significant effect on virus excretion and number of days virus was excreted. Although limited virus excretion was observed in the vaccinated lambs, transmission did not differ significantly between vaccinated and non-vaccinated groups. Nevertheless, the point-estimate of Rv tends to be lower than Rnv. Key words: foot and mouth disease, lambs, transmission, virus excretion 19 Definition of Uncommon Fatal Toxoplasmosis in Newborn Lambs H. Atmaca , O. Kul , N. Ocal , C. Babur , N. Kabakci 1 Faculty of Veterinary Medicine, Kirikkale University, Veterinary Pathology, Kirikkale, Turkey 2 Faculty of Veterinary Medicine, Kirikkale University, Veterinary Internal Medicine, Kirikkale, Turkey 3 Refik Saydam Hygiene Center, Parasitology, Ankara, Turkey Toxoplasmosis is a protozoer disease caused by an Apicomplexan parasite, T. gondii, and causes abortion, stillbirth and death almost in all species throughout the world. Although, transplacental infection is common in sheep and goats, systemic toxoplasmosis may also occur in newborn, young and immuncompromised animals. In natural infections, T. gondii causes necrotic placentitis and it may transplacentally pass to the fetus and finally resulted with abortion. In this study, we report an unusual clinical form of toxoplasmosis characterized by death of 20 lambs (5 of which twin) following severe neurological symptoms within 2 months of their life. Diseased lambs showed mild to severe clinical signs such as incoordination, head shaking, tremor and difficulty in walking when forced to walk. No abortion was observed in the pregnant ewes during the lambing season. Two lambs having typical signs of the disease were

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*Significantly different from respective C-value, P<0.05; significantly different from respective Ewes serum T0-value, P<0.01. These data show that vaccination of pregnant ewes against enterotoxaemia significantly increased Ab and in serum and colostrum. Lambs of vaccinated ewes showed significantly increased Ab 24h after birth, indicating that colostrum from vaccinated ewes provided significantly higher Ab levels than did vaccination of adult sheep. Ab levels in milk at weaning were low but a significant vaccination effect was still present and could provide a local gastro-intestinal protection in suckling lambs. Key words: enterotoxemia, lambs, vaccination of ewes, colostrum 18 Quantification of Foot and Mouth Disease Virus Excretion and Transmission within Groups of Lambs with and without Vaccination K. Orsel , A. Dekker , A. Bouma , J. Stegeman , M. De Jong 1 Utrecht University, Farm Animal Health, Utrecht, Netherlands 2 Central Institute for animal Disease control CIDC, Farm Animal Health, Lelystad, Netherlands

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Sheep, Goat and Buffaloes 111

necropsied, tissue samples were taken and processed for pathologic examinations. Blood samples of 15 ewes (dams of the 20 dead lambs) and 3 rams were collected and then Sabin Feldman dye test was used for the detection of T. gondii spesific antibodies and respectively 13 (86,6%) and 2 (66,6%) were found to be positive. Toxoplasmosis in pregnant ewes generally results with abortion by the way either following a new acute infection or reactivation of the chronic infection during pregnancy. Therefore, abortion can be used as an indicative of a transmitted infectious disease from dam to fetus. So it may be has a key role on differential diagnosis of such a herd problem e.g. toxoplasmosis. Postnatal lamb deaths and lacking abortion during pregnancy are not frequently seen in natural toxoplasmosis in sheep. However, death of a large number of newborn lambs in a herd should be investigated fortoxoplasmosis, serologically and etiologically. Key words: Toxoplasma gondii, Sabin Feldman dye test, immunohistochemistry, fatal toxoplasmosis, lamb 21 The Role of Estradiol and Somatotropin in the Pathogenesis of Ovine Pregnancy Ketosis R. Everts , R. Jorritsma , M. Houweling , L. Leengoed Van , 2 A. Tielens 1 Utrecht University, Department of Farm Animal Health, Utrecht, Netherlands 2 Utrecht University, Department of Biochemistry and Cell Biology, Utrecht, Netherlands Ovine pregnancy ketosis (pregnancy toxemia, twin lamb disease) is the most common metabolic disorder of ewes during the last weeks of pregnancy. Ewes with the disease normally carry two or more lambs. It is biochemically and endocrinologically characterized by hypoinsulinemia, ketosis, and an increase in the plasma concentrations of non-esterified fatty acids (NEFAs), cortisol, estrogens and somatotropin. While morbidity is low, the reported mortality in absence of therapy is 90%. It was thought that the main cause of pregnancy ketosis in sheep is a disturbance of carbohydrate metabolism by the high demands for glucose by de developing fetuses in the last trimester of pregnancy. For several reasons the appropriateness of this hypothesis can be questioned. First, the glucose turnover rate during lactation is two to three times the demand during pregnancy, without according increase in ketone bodies during lactation. Second, fasting of pregnant ewes does not induce all clinical and endocrinological aspects of pregnancy ketosis. In contrast to ewes with spontaneous pregnancy ketosis, ewes with induced ketosis start eating almost immediately upon refeeding. Third, the observation that immediately after (induced) delivery, a rapid recovery of the ewe occurs despite the increased energy demand for the production of milk, further questions the importance of a high demand for glucose as the sole inducer of ovine pregnancy ketosis. Therefore, we propose that the rise in 17,-estradiol and somatotropin, seen in affected ewes, hampers the normal metabolic regulation of the negative energy balance, resulting in pregnancy ketosis. The aim of this study was to investigate the influence of estradiol and somatotropin on the lipid metabolism in isolated sheep fat cells and to compare the lipid metabolism in sheep with pregnancy ketosis to that in healthy pregnant sheep. For that reason, subcutaneous fat biopsies were taken from sheep at the perineal area of the tail base. Lipolysis was assessed in freshly isolated fat cells and after 30h of preincubation. Furthermore, in the fat biopsies mRNA levels of several metabolic regulators such as PPAR, adiponectin and leptin were assayed. Key words: ketosis, ovine, lipid metabolism, estradiol, somatotropin 22 Individual Animal Risk Factors for Ovine Clinical Mastitis S. Waage , S. Vatn 1 Norwegian School of Veterinary Science, Production Animal Clinical Sciences, Oslo, Norway 2 Animalia, Oslo, Norway Clinical mastitis (CM) occurs frequently in ewes, causing great economic loss and a serious animal welfare problem in the sheep industry. The objectives of this study: were to examine the distribution of ovine CM cases in relation to the time of lambing and identify and quantify the effect of risk factors for the disease. Materials and methods: We conducted a case-control study, using individual animal data and disease records from a national sheep

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registry in Norway. Ewes that lambed in the spring of 2004 were included. Eligible cases (n=2857) and controls (n=76,716) from 1056 flocks of meat sheep were matched on flock (m:n matching) and multivariate conditional logistic regression was used for analysis of the data. Results: A large proportion of the CM cases occurred close to parturition; 32% were treated between days 0 and 4 after lambing, and 55% were treated within 2 weeks postpartum. Increasing age of the ewe and the need for assistance at lambing owing to dystocia were associated with increased CM risk. However, the effects of both these factors were modified by the number of lambs born and were most pronounced for ewes with 1 lamb. Increasing number of lambs born to the ewe, including stillborn, was associated with a marked increase in the odds of CM. For example, odds for 2-year old ewes without dystocia were 6.7 times greater for those with more than 3 lambs than for those with 1 lamb. Ewes of old Norwegian breeds were less likely to experience CM than were ewes of other breeds (OR=0.59; 95% CI: 0.46-0.74). Ewes treated for CM at least once during the preceding three years had greater odds of CM than ewes without a CM history (OR=4.0; 95% CI: 3.0-5.3). Conclusions: The fact that the risk of CM increases considerably with increasing number of lambs should be taken into account in sheep breeding programmes. Ewes that have experienced CM are at increased risk of suffering a new CM episode and should be slaughtered before the next breeding season, even if there is no sign of chronic udder changes. Our effect estimates for the risk factors, which are adjusted for breed and unaffected by inter-flock variations, are likely to be valid also for other meat sheep populations. Key words: ewe, mastitis, risk factor, case-control study 23 Relationship between Blue Tongue Virus Infection, Cyclicity and Individual Characteristics of Ewes before Autumnal Breeding in Texel Sheep Flocks in North of France C. Ponsart , N. Pozzi , D. Jeantils , JM. Camus , MC. Deloche , 5 6 1 2 S. Bareille , A. Bensmaili , P. Humblot , B. Guerin 1 UNCEIA, R&D, Maisons-Alfort, France 2 LNCR, R&D, Maisons-Alfort, France 3 CEIA des Ardennes, Villers Semeuse, France 4 Chambre d'Agriculture 08, Service Technique, Villers Semeuse, France 5 Intervet-Schering-Plough, Beaucouzé, France 6 GDSB, Villers Semeuse, France In 2007, the Bluetongue (BT) outbreak from North East to the centre of France may have affected breeding performances of ewes in these regions. This study aimed to describe the effects of BT virus infection in 149 Texel ewes on cyclicity and individual characteristics during breeding. Seven flocks from the Ardennes region having at least 10 ewes with clinical signs (CS) observed within 30 days before breeding and for which the breeding season started in October 2007 were included in the study. In each flock, two equal groups of 10-15 ewes (total with CS, n=77 or without CS, n=72) matched on age were bred with the same rams. Body condition (BC) scoring and blood sampling were performed in average 14±4 and 27±4 days after the start of the breeding period. Cyclicity was determined by progesterone assays: ewes for which the two progesterone measurements were <1.8 ng/ml were considered as anoestrous ewes. The others were considered as cyclic. BT status was diagnosed by using a combination of antibody detection (ELISA) and viral RNA detection (PCR) at the 2nd blood Table 1. Association between BT status and characteristics of ewes (p<0.05, by chi analysis) Characteristics Class (%, n) NEG (13, n=19) 40 9.3 0 0 NI (36, n=54) 14.3 24.1 62.1 44.1 A (29, n=43) 17.1 31.5 31.0 35.3 I (22, n=33) 28.60 35.2 6.9 20.6

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Age of ewes (year)

1 (35) 2-3 (54) 4 (58) < 2.5 (34)

Body condition score at first sampling

[2.5 ; 3.5] (53) >3.5 (62)

3.8 27.4

43.4 25.8

35.8 19.4

17.0 27.4

112 XXV. Jubilee World Buiatrics Congress 2008

sampling. The BT status was found negative in 19 ewes (NEG, 13%, ELISA-/PCR-), whereas 54 were newly infected (NI, 36%, ELISA/PCR+), 43 were at an acute stage (A, 29 %, ELISA+/PCR+) and 33 immune (I, 22 %, ELISA+/PCR- ; Table 1). All ewes were cyclic, with no difference found in mean progesterone levels in ewes with or without CS (11.3±0.5 vs 11.6±0.5 ng/ml, p>0.05) and when comparing negative and infected ewes (NEG : 11.1±0.5 vs NI+A+I : 11.4±0.5 ng/ml, p>0.05). The presence of CS within 30 d before breeding was associated with a lower BC score at first sampling (CS : 3.0±0.1 vs No CS : 3.5±0.1, p<0.01). In each flock, the BT status (NI, A, I) was significantly related to the age of ewes (more NEG females among young ewes) and the body condition score at first sampling (Table 1). To conclude, BT status seemed to have no effect on cyclicity rate in Texel ewes but was related to age and BC of ewes. Key words: Blue Tongue Virus, ewes, cyclicity 52 A Five-Year Longitudinal Study of Serum Antibody Responses in a Large Cohort of Dairy Sheep Infected with Mycobacterium Avium Subsp. Paratuberculosis (MAP) G. Foucras ,S. Lesceu ,S. Casu ,A. Carta ,P. Pourquier 1 Ecole Vétérinaire, Pathologie Médicale et Chirurgicale des Ruminants, Toulouse, France 2 Istituto Zootecnico e Caseario per la Sardegna, OLMEDO, Italy 3 ID-VET, Montpellier, France Objectives: The aim of the study was to follow within flock MAP infection using an absorbed ELISA, and determine the rate of serological conversion in a single birth cohort of dairy ewes during a 5 year-period. Materials and Methods: A flock of dairy ewes (n=965) endemically infected by Map, was blooded every year for 5 years, and serum samples were stored at -20 °C. Animals were slaughtered at 5 years of age, and several tissues and feces, in addition to a final serum were collected. Moreover, carcass weight, fattening, and macroscopic lesions (ileal thickness, mesenteric lymph node hypertrophy) were recorded. Lesion intensity was scored (scale 1 to 4) accordingly. Serum samples were tested retrospectively by ELISA (ID Screen Paratuberculosis Indirect, ID-VET, France) with a pre-absorption step on M. phlei. Samples were tested in duplicate wells, and data expressed as sample to positive control ratio (S/P values). Results: High reproducibility was obtained with ID Screen Paratuberculosis Indirect ELISA between wells and between days, with a correlation coefficient higher than 95%. At least 2/3 of the animals testing positive had S/P values above 100%, giving a large difference between positive and negative samples. During the course of the study, cohort size decreased progressively, main losses being caused by clinical paratuberculosis. When ELISA values were interpreted as compared to a previously defined threshold (S/P 60%), frequency of sheep ranking positive increases from 4.1% (18 month-old) to 14.7% (60 month-old), with the highest incidence at the age of 42 months that was 19.1%. After five years, estimated serological prevalence is 25.7%. To our surprise, individual values varied significantly during the course of the survey, with some animals changing from positive to negative status during the last two years. Most of these animals had paratuberculosis-evocated lesions, indicating that these results are not false positive. These observations suggest that, once arisen, MAP-specific antibody response may not be as stable as previously thought, with some responses falling below positive threshold during the course of the infection. Conclusions: These data improve our knowledge about the evolution of Map antibody response in sheep, and suggest that repeated screening twice a year from the age of 2 years might be necessary for optimal detection of MAP-infected sheep. Key words: sheep, paratuberculosis, serology, ELISA 53 European Field Clinical Studies with Cydectin 2 % LA for Sheep T. Blond ,D. Parker 1 Fort Dodge Animal Health, Regional Technical Office, Tours, France, France 2 Fort Dodge Animal Health, Regional Technical Office, Southampton, United Kingdom The objective of this study was to evaluate the prophylactic efficacy of Cydectin® 2 % LA injectable for sheep against nematode infections in sheep under field conditions in the EU. Studies were carried out in

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France, Greece and United Kingdom. A total of one hundred and sixty five sheep, either lambs or ewes naturally infected with gastrointestinal nematodes, were selected for these trials. One hundred and ten of them were treated with the tested product at the recommended dose while the others were injected with saline at the base of the ear. In all the studies the animals shared the same grazing throughout the trial. Individual animal fecal samples were collected pretreatment and on days 28, 42, 56, 70, 84, 98, 112, 126 (2 trial only), 140,168 (1 trial only) and 182 (1 trial only) days post treatment. Bulk larval cultures were carried out at each sampling time. Undifferentiated egg counts were partitioned into helminth genera based on the relative proportion of larvae identified by coproculture. Animals were either weighed or condition scored on a few occasions evenly spaced during the trial. Local and general tolerance of the treatment was assessed. Based on geometric means, Cydectin 2 % LA was effective in reducing the number of undifferentiated helminths eggs by more than 90 % from day 28 to day 126. In the UK trial, control lambs had to be treated with salvage treatment on day 28 because the infection level of the animals was too high, Day 28 control group egg counts were used to compute treatment efficacy at subsequent sampling times hence the genius/species efficacy could not be determined in this trial. In the other two trials the product showed efficacy superior to 97 % for Ostertaggia, Cooperia, Haemonchus, Nematodirus and Trichostronlgylus until day 126. There were no significant differences in the average weight gains of the lambs, the average weight gains were 16.7 and 17.9 kg, respectively, for treated and control groups in the UK trial, and 10.9 and 8.9 kg for the ewes in the Greek trial. Condition scores done on the ewes in the French study were 1.8 and 1.6, respectively, in the treated and control groups. No adverse effects were observed in any of the studies. Cydectin 2 % LA injectable for sheep has been shown to be effective under various field conditions in the EU, with an efficacy against gastrointestinal nematodes greater than 90 % for 126 days and a good tolerance at the injection site. 54 Effect of Detomidine-Butorphanol-Ketamine and Atipamezole on Clinical, Cardiorespiratory, Haematologial Parameters in Sheep G. Atalan , V. Gunes , Z. Okumus , F. Celebi , G. Atalan , 1 A. Onmaz 1 University of Erciyes, Faculty of Veterinary, Surgery, Kayseri, Turkey 2 University of Ataturk, Faculty of Veterinary, Surgery, Erzurum, Turkey 3 University of Ataturk, Faculty of Veterinary, Department of Physiology, Erzurum, Turkey 4 Sisli Etfal Hospital, Anaesthesiology, Istanbul, Turkey The aim of this study was to evaluate the sedative-anaesthetic effects of a combination of detomidine (D, 30 mg/kg, intramuscular (IM)), butorphanol (B, 0.2 mg/kg, IM), ketamine (K, 5 mg/kg, IM ) in sheeps. 9 sheeps (3 female and 6 male, 1 to 2 years old, mean 53,6±7,5) were used. The combination of D and B were administrated to produce sedation. Ten minutes after D and B administration, K was injected. The anaesthetic effects of the drugs were reversed by administration of Atipamazole (AT) at 30 minutes after K injection. The sedativeanaesthetic effect of D+B+K combination and alterations in cardiorespiratory, electrocardiographic (ECG- lead-II findings, 25 mm/sn) blood gas and haematological parameters were investigated before and 10 min after pre-medication with D+B, at 10, 20 and 30 min during the K anaesthesia and at 10 min following AT administration. The D+B+K drug combination produced a satisfactory sedation and anaesthesia for the all sheep. They recovered successfully. The sedative effect of the B+D appeared within 3 minutes and the anaesthetic effect started within 5 minutes after K administration as indicated by lack of pain stimuli. Mean heart rate decreased significantly until the administration of AT but increased following the AT injection. Mean respiratory rate increased significantly throughout the anaesthesia. Body temperature decreased gradually during the anaesthesia from 39.6 to 38.7. In ECG findings, amplitude of P values decreased at 40 minutes of anaesthesia but returned its baseline value after 10 min of AT injection. PR interval and R amplitude increased gradually during anaesthesia but decreased after the administration of AT. Q-T value gradually increased at 10 min after B+D, at 10, 20 after K administration and having the highest value at 30 minute of anaesthesia but decreased after 10 min of AT injection. The mean pH and pO2 values gradually decreased at 10 min after K injection, and increased after AT administration, approximating to their

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baseline values. The mean HCO3, PCO2 and ctCO2 increased during anaesthesia but decreased after AT injection. There were no significant alterations for haematological parameters. It is concluded that B+D+K anaesthesia effected the cardiorespiratoric parameters in normal ranges. It might be used for induction of anaesthesia. This anaesthesia is successfully reversed by administration of AT. Key words: sheep, anaesthesia, butorphanol, detomidine, Atipamezole 55 Haematological and Clinical Biochemical Parameters for Adult Sheep and Goat A. Tschuor, B. Riond, U. Braun, H. Lutz University of Zurich, Department for Farm Animals, Zurich, Switzerland Objectives: Sheep and goat husbandry has always played an important role in Swiss agriculture, but in recent years these animals are increasingly appreciated as hobby pets. This opens new diagnostic and therapeutic perspectives for veterinary surgeons, notably in the determination of haematological and clinical biochemical parameters. For the methods used nowadays there are no reference range values available for adult sheep and goats kept under central European conditions. Especially clinical biochemical parameters had to be compared with incomplete reference values, which were determined in the past with aged methods. Due to this, every interpretation of laboratory findings left incertitude. Material and methods: Therefore, the present study was designed to determine haematological (haematocrit, haemoglobin, total erythrocyte count, MCH, MCHC, MCV, white cell count and differentiation, fibrinogen, plasmatic proteins) and clinical biochemical parameters (bilirubin, gluc, urea, crea, albumin, cholesterol, triglycerides, AP, amylase, GLDH, ASAT, ALAT, GGT, CK, lipase, SDH, LDH, sodium, potassium, chloride, calcium, magnesium, phosphorus, iron, fructosamine, Alpha-1-Globuline, Alpha-2-Globuline, Beta-Globulins, Gamma-Globulins, A/G-ratio) in 102 goats and 102 sheep from Swiss flocks. The haematological examination was done using a Cell Dyn 3500® (Abbott AG, Diagnostics Division) and the examination of the biochemical parameters were done by a Cobas Integra (Roche Diagnostics). Results: Significant differences were found between the two species especially regarding erythrocyte count, mean erythrocyte volume, haemoglobin content as well as leukocyte count. There were also significant differences between this new obtained reference values compared to the aged ones from central Europe. Conclusion: It is therefore of utmost importance to discriminate between sheep and goats when interpreting laboratory findings by means of actual reference values. Additionally, the study provides a literature based overview explaining the significant differences between these two species in comparison to the bovine reference values. 56 Impact of Bluetongue Virus Serotype 8 on Rams' Semen Quality N. Kirschvink , M. Raes , C. Saegerman 1 University of Namur, Veterinary Department, Animal Physiology, Namur, Belgium 2 University of Liege, Department for Infectious Diseases Epidemiology; Faculty of Veterinary Medicine, Liege, Belgium Bluetongue virus (BTV) is reported to induce transient infertility in infected ruminants. The aim of this study was to assess the effects of BTV serotype 8 on semen quality of rams tested at several occasions after infection and by comparing them to healthy control rams. 11 healthy rams tested prior to the BTV outbreak in Belgium were used as controls (C) and 12 rams with natural infection by BTV serotype 8 (BT) were included in this study. BT rams were tested on 2-6 occasions (BT-S1 to BT-S6) 5 to 138 days after development of clinical signs of BTV. BTV infection was serologically confirmed. Semen was evaluated for motility (score ranging from 0-5), spermatozoid concentration and viability (% of normal living [NL]; normal death [ND]; abnormal death [AD] spermatozoa). Results are shown as mean ± SD. These results demonstrate the important impact of serotype 8 BTV on rams' semen quality. 85 days after development of clinical signs of BTV, the variables tested did no more significantly differ from control animals, suggesting that full recovery of semen quality occurred.

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Controls BT-S1 Motility 4.5±0.2 Conc (109 sperm/ml) 3.2±0.7 % NL 75±6 % ND 21±7 % AD 4±3

BT-S2

BT-S3

BT-S4

BT-S5

BT-S6

0.8±0.5* 1.3±1.9* 2.4±1.9* 3.3±1.6* 4.0±0.9 4.5±0.3 1±0.7* 9±10* 48±12* 43±11* 1.2±1.4* 1.2±0.9* 1.5±0.8* 2.2±0.9* 27±32* 46±32* 54±25* 63±16* 37±22* 33±21* 30±10* 29±11* 37±28* 21±25* 16±23* 9±13 2.6±0.6 76±8 20±7 4±4

* significantly different from respective control-value, p<0.05; Welch Test. Key words: BTV serotype 8, ram, semen quality, repeated semen analysis 57 Preliminary data to Suggest the Use of Colostrum and/or Milk Samples as Test Media for Johne's Disease in Sheep and Goats M. Ayers, B. Mamer, M. Bulgin University of Idaho, Caine Veterinary Teaching Hospital, Animal and Veterinary Science, Caldwell, Idaho, USA, United States Objectives: To investigate which test(s) and which sample(s) would provide the most accurate, convenient, and economical means of finding positive individuals in sheep flocks and goat herds that have had Johne's disease, caused by Mycobacterium avium subsp paratuberculosis (MAP), diagnosed in the flock or herd. Materials and Methods: Twenty-nine paired sera, milk, and fecal samples were collected from ewes and does of known Johne's positive flocks/herds. Milk samples were centrifuged with the skim used for antibody testing and the pellet for culture. Sera and skim milk were split for antibody testing using two serological ELISA tests (IDEXX HerdchekTM and Biocor ParachekTM). Our lab ran the milk undiluted and at 1:2 dilution and the sera per kit instructions using IDEXX HerdchekTM. A second lab ran the split samples using kit instructions for both the IDEXX and Biocor tests. Milk pellets and feces were cultured using an increased inoculum in two culture media: BACTECTM MGITTM(BD Diagnostics) para TB liquid medium with the fluorometric manual read method; and Herrold's egg yolk agar (HEYA) with and without mycobactin J. Cultures were incubated at 35 C for up to 12 months. Results: Twelve of 29 sera were positive with IDEXX HerdchekTM and 4 of 29 positive with Biocor ParachekTM. Eleven of 29 milk samples were positive undiluted and/or at 1:2 dilution with IDEXX HerdchekTM, 2 of 29 positive with Biocor ParachekTM, and 0 of 29 positive with IDEXX HerdchekTM at the standard sera dilution of 1:20. So far 6 of the 12 serology and/or milk positive animals have been fecal culture positive. An additional animal was milk pellet positive, fecal acid fast bacilli (AFB) positive, and tissue AFB positive on histopathology. Making 7 of 12 serology and/or 11 milk positive animals confirmed as MAP culture positive. Conclusion: Milk samples tested undiluted and/or at 1:2 dilution using IDEXX HerdchekTM were similar to sera using the same test. Colostrum may prove to be superior to milk due to concentration of maternal antibodies. This is an appealing sample as most ewes and does are handled at or near parturition and samples could be collected by the owner. Colostrum may result in increased pellet culture positives in a decreased culture time due to increased numbers of macrophages present. A larger project to begin late January 2008 involving 400 to 500 ewes is planned to test these hypotheses. Key words: Johne's, MAP, sheep, goats, milk 92 Comparative Studies on Fluid Resuscitation in Goat under Surgical Hypovolemic Shock M. Awad , I. Ahmed , H. Gohar , A. Ghanem 1 Faculty of Veterinary Medicine Suez Canal University, Veterinary Surgery and Radiology, Ismailia, Egypt 2 Faculty of Veterinary Medicine, Cairo University, Veterinary Surgery and Radiology, Cairo, Egypt Objectives of the study: establishment of a controlled model that simulates severe hemorrhagic shock and to assess the effectiveness of a small volume hypertonic crystalloid solution for treatment of hypovolemic hemorrhagic shock in goats in comparison to different treatments adopted for hypovolemic shock in ruminants. Materials and Methods: This study was applied on Twenty-five goats, divided randomly into five equal groups as the following 1) non treated

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114 XXV. Jubilee World Buiatrics Congress 2008

group 2) received blood 3) received I/V hypertonic saline 4) received oral isotonic electrolyte solution and 5) received intravenous hypertonic saline followed immediately by oral electrolyte solution. Hemorrhagic shock was induced by constant removing of blood over 15-20 minutes. The amount of the withdrawn blood ranged between 532 ml to 616 ml. depending upon body weight of each goat. Immediately before hemorrhagic shock induction, the baseline line values were recorded including heart rate, ECG, respiratory rate, rectal temperature, body weight and a maximum clinical score (MCS), Hematocrit, blood gas analysis) pH, Pco2, Po2 ), serum biochemical analysis (Na, K, Cl, Ca, Phosphate, total protein, albumin and creatinine concentration), blood lactate and plasma volume changes. These parameters were also determined at the time of entry into the treatment phase, 30 minutes, 2 hours, 8 hours, and 24 hours after treatment. Results and Conclusions: results indicated that intravenous administration of hypertonic saline solution together with oral administration of electrolyte solution in hypovolemic goats caused immediate and sustained effect. It was superior to administration of either the two solutions alone and induced the same effect as whole blood. Meanwhile resuscitation with Hypertonic saline solution alone resulted in only transient resuscitation. Key words: goat surgery, hypovolemic, shock 93 Estimation of Gestational Age in Egyptian Goats by Ultrasonographic Fetometry A. Karen , E. Fatouh , S. Shokr 1 Faculty of Veterinary Medicine, Theriogenology, Kafr El-Sheikh, Egypt 2 Faculty of Veterinary Medicine, Anatomy and Embryology,Ismailia, Egypt The present study aimed to determine the earliest day at which the signs of pregnancy can be detected by transrectal (TR) and transabdominal (TA) scanning and to estimate the gestational age in Egyptian goats by ultrasound measurement of embryonic or fetal parts throughout pregnancy.Fifteen pregnant Egyptian goats were used in the present study. The date of their mating was considered as Day 0 for calculating the gestational age. The does were transrectally scanned by using a 7.0 MHz vector probe (Acuson 128 XP/10 computed scanner) at Day 10 post mating on alternate days until Day 25 and then once at 3-5 day intervals until day 50. Linear and convex (3.5 to 5 MHz) probes were used for TA scanning which were carried out from Day 25 to Day 130 at 3 to 5-day intervals. The days after mating at which the embryonic vesicle and the embryo proper with a beating heart firstly imaged by TR and TA scanning were recorded. After imaging the embryo or the fetus, the following parameters were measured: length of the embryo or fetus (CRL), heart rate (FHR), biparital diameter (BPD), trunk diameter (TD), placentome size (PS), umbilical cord diameter (UCD) and femur length (FL).The average of days at which the embryonic vesicle was firstly determined by TR and TA scanning was 16.98±1.97 and 27.87±3.48, respectively. The embryo proper with a beating heart was firstly determined by TR and TA scanning at an average of 22.36±2.66 and 30.36±4.75 days, respectively. All the fetal measures were significantly (P<0.0001) correlated to the gestational age. The fetal TD (R2=96), the BPD (R2=95.6), the CRL (R2=94.1, the UCD (R2=93), PS 2 2 (R =91) and the FL (R =90) were highly correlated with the gestational 2 age. On the other hand, the FHR was poorly correlated (R =55.1) with the gestational age. In conclusion, early pregnancy in Egyptian goats can be accurately diagnosed from Day 22 of gestation by using TR scanning (7MHz) and at Day 30 by TA scanning. The age of embryo or fetus in Egyptian does can be accurately estimated by ultrasound measuring the crown rump length, biparital diameter, trunk diameter, placentome size, umbilical cord diameter and femur length. Key words: ultrasonography, fetometry, goats 94 Diagnosis of Scrapie in Goats by the Detection of Abnormal Prion Protein in the Central Nervous System and Lymphoid Tissues H. Cassard , F. Corbiere , C. Chauvineau-Perrin , C. Lacroux , 3 1 1 F. Barillet , F. Schelcher , O. Andreoletti 1 INRA ENVT UMR 1225, Interactions Hôtes-Agents Pathogenes, Toulouse, France 2 AFSSA, Laboratoire d'Etudes Caprines, NIORT, France 3 INRA, Station Genetique d'Amelioration des Animaux, CASTANET TOLOSAN, France

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Scrapie is a naturally occuring transmissible spongiform encephalopathy of sheep and goats characterized by the accumulation Sc of an abnormal isoform (PrP ) of a cellular protein in infected tissues. Sc After oral exposure, PrP is detected in lymphoid tissues in early stages of the disease, before invasion of the central nervous system (CNS). Post-mortem diagnosis of scrapie in goats currently relies on detection of PrPSc in the CNS by ELISA, immunohistochemistry (IHC) or Western blotting but data concerning reliability of these assays when applied to other tissues are lacking. Moreover, tonsil biopsy can be used as a diagnostic tool in living sheep but this method has not been tested yet in goats. The aims of our field study were (i) to compare the sensitivity of ELISA and IHC when performed on different tissues samples and (ii) to assess the relevance of tonsil biopsy for the detection of preclinical scrapie cases in living goats. Eleven scrapie infected flocks (n=2756 goats), in which a scrapie positive index case had been detected at the rendering plant or at the abattoir, were investigated. Tonsil biopsies were performed in 7 flocks for almost all animals (n=1570). Additionally, obex, palatine tonsils, the ileal mesenteric lymph node (MLN) and the distal ileum were collected in more than 96 % of animals (n=2658) at slaughter. PrPSc was detected using IHC on tonsil biopsies and ELISA and IHC on necropsic samples. For each kind of sample, the concordance between the results of IHC and ELISA and the sensitivity of both methods were assessed. A total of 183 infected animals was detected. Seven out of 11 flocks could be considered sporadically infected, with an observed prevalence below 1.5 %, whereas scrapie was enzootic in the 4 other flocks, with a prevalence ranging from 13.1 % to 31.8 %. Whatever the tissue tested, IHC and ELISA gave only few discordant results (19/9655). The performances of both diagnostic methods applied to obex were poor (sensitivity near 60 %). The highest sensitivities (near 95 %) were obtained with palatine tonsils and MLN. IHC applied to tonsil biopsies had a significantly higher sensitivity (90.6 %) than ELISA and IHC performed on obex. This method was also as efficient as ELISA or IHC performed on MLN and palatine tonsils collected at necropsy. This study shows that NLM and palatine tonsils can be used for post-mortem diagnosis of scrapie in goats and that tonsil biopsy is an efficient preclinical diagnostic tool in living goats. Key words: scrapie, goats 95 Limited Cross-Protection in Goats Following Immunization to Differing Field Isolates of Contagious Ecthyma J. Musser , C. Taylor , D. Waldron 1 Texas A&M University, Veterinary Pathobiology, College Station, United States 2 Texas Agricultural Experiment Station, Sonora, United States 3 Texas Agricultural Experiment Station, San Angelo, United States Contagious ecthyma (CE), also referred to as soremouth in animals and orf in humans, is a common viral disease of ruminants worldwide. A parallel groups cross-protection study in goats, using a live virus vaccine (A&M vaccine) and a wild-type contagious ecthyma virus strain (wtCE strain), was conducted to determine if recent vaccine failures were due to antigenically dissimilar viral strains. The wtCE strain was obtained from lesions during a severe outbreak of contagious ecthyma in a previously vaccinated goat herd. Boer x Spanish cross goat kids (n=117), <14 days of age, from a single herd were allotted to 4 treatment groups: 1) A&M vaccine immunized, A&M vaccine challenged; 2) A&M vaccine immunized, wtCE strain challenged; 3) wtCE strain immunized, A&M vaccine challenged; and, 4) wtCE strain immunized, wtCE strain challenged. Primary immunization site lesions (scabs) were assessed 12 days after inoculation. Ten weeks after primary immunization, goats were challenged; scab formation was assessed 10 days later. Locations for immunization and challenge were the medial aspect of the right and left thigh, respectively. Scab formation was scaled 0 to 4: 0 = no response, 1 = mild response, 2 = moderate response, 3 = heavy response, and 4 = extreme response. No scab formation following challenge was suggestive of protective immunity induced by primary immunization. Positive reactions were seen at the site of primary immunization with the A&M vaccine (79 of 81) and the wtCE strain (33 of 36). Significant differences (p<0.001) were noted between scab formation scores following primary immunization and after challenge with the same viral strain. In groups immunized with A&M vaccine, there was significantly (p<0.001) more scab formation scores with wtCE strain challenge than with A&M vaccine challenge. In the groups immunized with wtCE strain, there was significant difference (p<0.001) in scab formation scores between wtCE strain challenge and A&M vaccine challenge. These results indicate that

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Sheep, Goat and Buffaloes 115

immunization with either the currently produced vaccine (A&M vaccine) or with a wild-type contagious ecthyma viral strain does not result in cross-protection. Antigenically dissimilar CE viral strains may have contributed to vaccine failures, thus contagious ecthyma vaccines should be multivalent or include locally occurring strains. Key words: contagious ecthyma, goats, vaccine, immunology 96 Prevalence and Determinants of Ixodids in Domestic Goats Lower Punjab (Pakistan) S. Sajid , Z. Iqbal , N. Khan , G. Muhammad 1 University of Agriculture, Faisalabad, Parasitology, Faisalabad, Pakistan 2 University of Agriculture, Faisalabad, Clinical Medicine and Surgery, Faisalabad, Pakistan Tick infestation is a serious nuisance in small ruminants of developing countries like Pakistan. The objective of the current study is to determine the prevalence and various determinants of tick infestation in caprines of lower Punjab (Pakistan). For this purpose, a total of 700 goats of various breeds, age and sex were randomly selected from each of district Muzaffargarh and Layyah of lower Punjab. The selected herds were visited thrice a month in order to collect all the relevant information including the species, breed, age and sex of host, infested number of animals, type of housing and average number of ticks per animal on a pre-designed questionnaire. Ticks were collected and processed through standard procedure for their taxonomic identification. Hyalomma anatolicum and Rhipicephalus sanguineus species were identified from the subjected hosts. The overall prevalence of ticks was 51.6% (723/1400) in both the districts. The highest month-wise prevalence in Districts Layyah (53.42%; 374/700) and Muzaffargarh (60.8%; 426/700) were found in July and lowest in November (3.14%; 22/700) and October (3.14%; 22/700), respectively. No significant (P = 0.760) differences in the prevalence of tick infestation as calculated through Z-test (Z = 0.31) were found between district Layyah and Muzaffargarh. However, significant differences were found in prevalence percentages of various tehsils of Layyah and Muzaffargarh. Regarding determinants, prevalence of tick infestation was found higher in males, older animals and Beetal breed goats. The preferred sites of ticks were inside and external ear. The results provided a better understanding of the status of tick infestation in the domestic goats of the study area and will be helpful in the planning of control programme against ticks. Key words: prevalence, ticks, goats, ixodids, Pakistan 97 Effect of Boron on Haemogram and Biochemical Profile of Urine in Buffalo (Bubalus Bubalis) Calves Fed High Fluoride Ration V. Bharti , M. Gupta 1 CCS Haryana Agricultural University, Hisar, Department of Veterinary Physiology, Hisar, India 2 Indian Veterinary Research Institute, Division of Physiology & Climatology, Bareilly, India The present study was carried out with the objective of ameliorative effect of boron (B) on high fluoride (F) intake. As per Randomized Block Design, twelve male Murrah buffalo calves of 6-8 months age were divided into three groups of four calves in each group. They were fed conventional concentrate mixture, roughage and supplemented with sodium fluoride (NaF, 60ppm elemental F) alone or in combination with borax (Na2B4O7.10H2O, 140ppm elemental B) for th th th 90 days. Blood sample were taken on 0, 30 , 60 and 90 day for analysis of Hb, TEC and PCV. Urine were collected at the end of th experiment (90 day) and analyzed for alkaline phosphatase activity (ALP) and minerals (viz, Ca, P, Fe, Zn and Cu).This level of fluoride in ration caused a significant (p<0.05) depression on blood Hb and PCV on day 90 which was marginally improved with B supplementation. Urinary F and ALP was also increased significantly. Boron supplementation with F caused significant (p<0.05) reduction of urinary F and ALP level. Urinary excretions of minerals (P, Fe, Zn and Cu) were remain unaffected on F as well as F+B treatment. However, calcium excretion was significantly (p<0.05) increased on F treatment and significantly (p<0.05) reduced on B+F supplementation. These findings suggest that boron has ameliorative effect on haemogram and biochemical profile of urine in buffalo calves fed high fluoride ration. Key words: boron, fluorine, haemogram, mineral, urine

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98 Estimation of Breeding Value of Milk Production in Iranian Buffaloes M. Sanjabi , A. Lavvaf , H. Naderfard 1 IROST, Animal Science, Tehran, Iran 2 Islamic Azad University, Science & Research Branch, Animal Science, Karaj, Iran This study was conducted on the data of 65534 individual milk records along with fat percentage from 1992-2002 in 452 herds in six provinces of Khuzestan, Mazndaran, W. Azarbaijan, E. Azarbaijan, Ardabil and Gilan. The data has been analyzed by SAS software in GLM procedure. Independent variables were: Milking days, evening and morning milk, total milk yield, fat percentage of evening and morning milk, ME milk in 100 and 200 days. The effects of year and season of calving were significant on milk production (p<0.05), least square mean of milk yield and fat percentage were calculated by using the complete pedigree records. The heritability and breeding value of 1195 animal were calculated by DFRML procedure. The estimated (h2) of milk production was 0.16 which is in the range of reported h2 by other researchers . The LSM of average milk production in the provinces of Gilan, Mazandaran E. Azarbaijan, W. Azarbaijan, Khuzestan and Ardabil were: 1452, 1586, 1382, 1183, 2135 and 1189 kg respectively. These results indicate that the province of Khuzestan has the most potential in the field of milk production. The average milk yield per lactation, days of lactation, fat percentage and LSM of fat percentage were 1513 kg, 202 days, 5.04 and 6.77 respectively. Characteristics of the best 3 female buffalo having the most breeding value the herds number of 60114, 60125, 60165 had higher BV of 91, 85, and 76 respectively and the best 5 male had BV49, 45, 37, 29 and 27 which all best male and female water buffaloes were belong to Khuzestan Providence. The top five highest breeding value bulls have been introduced to buffalo population by artificial insemination. It can be conclude that Iranian buffaloes still have high genetic potential that in case of proper environmental management, have a genetic potential of milk production Key words: breeding value, buffalo, milk production 99 Naloxon's Effects on Post-Thawed Buffalo Sperm Motility, Viability and Capacitation R. Minoia, F. Sassone, A. Guaricci, G. Neglia, L. Pavone, M. Nicassio Facolty di Medicina Veterinaria, Animal production, Valenzano (BA), Italy It is generally accepted that a substantial number (about 50%) of sperm cells are damaged during cryopreservation and several studies are concentrated on research of better standards of semen cryopreservation to optimize viability and, so, the usability for artificial insemination. Poor studies are published on possibilities to improve sperm viability after thawing in buffaloes. In literature, it is evidenced that the administration of opioid antagonists to raw or frozen semen significantly improves the capacity of sperm cells to acquire the conditions essential to fertilize oocytes. Objective of the study: The aim of the present study was to evaluate the effects of naloxone (Nx), an opioid antagonist, added post-thawing, on buffalo sperm cell quality analizing as the motility by CASA system, as the viability and capacitation by Chlortetracycline (CTC)/Hoechst staining. Materials and Methods: Straws of buffalo semen were thawed for 30 sec in a 37 °C water-bath; the semen recovered was washed with a modified Tyrode medium. For the functional test, sperm cells were incubated at the concentration of 25 millions/mL at 38°C in absence (control condition) and in presence of different Nx concentrations (10 3 -6 -8 , 10 and 10 M) or heparin (10 IU/mL) as known capacitation inducer. Viability and capacitation were detected by CTC/Hoechst 33258 staining at 0, 1 and 3 hours incubation time points. Fluorescence microscope analysis clearly Allowed differentiation of dead/live and capacitated/non capacitated cells. Results: Statistical analysis of motility data by CASA showed that only the average path velocity (VAP) at 1h was modified negatively by -3 the concentration of 10 Nx (p<0.05) in respect to control condition, whereas all other parameters were not modified. Statistical analysis of viability and capacitation data revealed that Nx at the concentrations of -3 -6 10 and 10 M significantly (p<0.05; 0.001 respectively) improved -6 sperm cell viability after 1h; whereas the Nx 10 M on capacitation

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116 XXV. Jubilee World Buiatrics Congress 2008

resulted efficacious after 3h of incubation in respect to control -6 condition. Moreover 10 M Nx at 1 and 3h statistically increased (p<0.001) the rate of viable cells compared to heparin determining, at the same time, a comparable rate of capacitated. Conclusions: These results indicate that Nx few or never influences -3 -6 buffalo sperm cell motility (only VAP at 10 M at 1h) but at 10 M could be considered a better capacitating agent in IVF protocols than heparin. Key words: buffaloes, semen, naloxon, CASA analysis 100 Effects of Dexamethasone Administration Following Thiopentone Induced Circulatory Insufficiency in Buffaloes P. Peshin, J. Singh, R. Tyagi, A. Kumar, S. Singh Chaudhary Charan Singh Haryana Agricultural University, College of Veterinary Sciences, Veterinary Surgery and Radiology, Hisar, India Common causes of cardiopulmonary embarrassment during surgery are anaesthetic overdose, to anaesthetics or preanaesthetics or iatrogenic problems. The most serious outcome in such cases follows over dosage with barbiturates. Therapeutic efficacy of massive doses of corticosteroids has been stressed in men, dogs and laboratory animals. Hence, the pertinent study was done to create cardiopulmonary dysfunction in buffalo calves with thiopentone over dosage and its management with dexamethasone. Studies were done on five clinically healthy male buffalo calves (Bubalus bubalis) of 8 months to 1 year age and weighing 55-120 kg. Circulatory insufficiency was induced with thiopentone sodium (5%), and dexamethasone (4.0 mg/kg, I.V.) was administered for treatment. Various parameters investigated were: rectal temperature (RT), haemoglobin (Hb), plasma glucose, plasma creatinine, total plasma proteins, heart rate (HR), systolic pressure (SP), diastolic pressure (DP), mean arterial pressure (MAP), central venous pressure (CVP), electrocardiogram (ECG), electroencephalogram (EEG), arterial carbon dioxide tension (PaCO2), arterial oxygen tension (PaO2) and venous oxygen tension (PvO2).The dilated pupils were constricted within 15 minutes and by 30 to 45 minutes resumption of corneal and palpebral reflex activity started. Electrical activity in the EEG began appear between 40 seconds to 5 minutes and there was an appreciable increase in MAP. The MAP began rising 1.75±0.76 minutes after dexamethasone administration and values were more stable by 3.17±0.77 minutes and near base values by 45 minutes. The MAP was elevated to 14.33±0.707 kPa five minutes after dexamethasone administration as compared to 4.69+0.433 kPa during cardiac insufficiency. No significant changes in the HR were seen. The CVP was lowered 2.33±0.215 kPa as compared to 4.49±0.191 kPa during cardiac insufficiency, but remained higher than base values (1.42±0.131 kPa) until 45 minutes. Three calves showed primary Twave changes and ST segment depression through at the period of study. Arterial oxygen tension remained elevated throughout the period of study, while venous O2 and arterial CO2 tension were within normal limits. There was slight fall in rectal temperature. Other blood biochemical parameters were not affected except for increase in blood glucose at 45 minutes. It was concluded that dexamethasone should be a valuable adjunct to therapy of circulatory insufficiency in buffaloes. Key words: dexamethasone, thiopentone, circulatory insufficiency, buffaloes

study. Forty two (70%) of these abomasums were contaminated by gastro-intestinal Nematodes. By minimum and maximum of counted Ostertagia (67-1300) and mean of 127, Thirty two (53.33%) of abomasums were contaminated by Ostertagia. Also, Eighteen (30%) of them were contaminated by two species of Ostertagia circumcincta and Ostertagia trifurcata. Minimum and maximum of collected Ostertagia was 85-1300 with the mean of 130. Eight abomasums (13.3%) were only contaminated by Ostertagia trifurcata with minimum and maximum of 67-230 and mean of 82. Six abomasums (10%) were also contaminated by Ostertagia circumcincta with minimum and maximum of 80-640 and mean of 110.There was a significant statistical relation between age and contaminating with Ostertagia and maximal contamination had been seen between the ages six to twelve months (P<0. 05). By age increasing, the contamination was decreased. Histopathological study of the abomasums contaminated by Ostertagia had been shown that parietal cells were decreased because of parasitic larval infiltration and inflammatory cells such as neutrophils and eosinophils could be found in lamina properia. Key words: Ostertagia, pathologic lesions, Shahrekord, sheep, species, abomasum 738 A Case of Zoonosis Ecthyma A. Sagkan Ozturk , H. Tol , I. Sen 1 University of Selcuk, Faculty of Vet. Med., Department of Clinical Sciences, Faculty of Veterinary Medicine, Konya, Turkey 2 University of Selcuk, Faculty of Vet. Med., Dermatology, Konya, Turkey Contagious ecthyma, also know as contagious putular dermatitis or orf, is a specific skin disease of sheep and goats, caused by a parapox virus of the family poxviridae. There are few published reports on contagious ectima in Turkey and other countries indicating the occurrence of the disease in both lambs and humans. Objectives of study: Therefore, the aim of this case report is to show occurred the lesion on a sheepherder's fingers that were formed after a lamb with a contagious ectima bites him. Materials and Methods: A lamb with a contagious ectima and sheepherder with orf. Results: In routine clinical examination of lamp, small lesions mouth and around were observed along with high temperature. Lamb's the lower respiratory s