Read A Lesson in Deep Sea Vent Symbioses: Riftia pachyptila and its chemolithoautotrophic symbiont -proteobacterium Joel Migliaccio text version

A Lesson in Deep Sea Vent Symbioses: Riftia pachyptila and its chemolithoautotrophic symbiont -proteobacterium
Introduction Physiology Biochemistry Free living symbiont More to come...
Joel Migliaccio

· Riftia pachyptila was first discovered by M.L. Jones in 1981 at the East Pacific Rise (EPR), Galapagos Spreading Center, and Guaymas Basin · 3 meters in length living at depths of ~2600 meters! The ever changing sea floor rifts are in various stages of succession
·
· Microbial floc, to Tevnia jerichonana, to R. pachyptila

Physiology of R. pachyptila and its symbiont:
The trophosome is composed of groups of cells forming lobules (Bright and Sorgo 2003)

3-D ultrastructural reconstruction of the R. pachyptila lobule using ultrathin section electron micrographs
· trophosome is more likely derived from mesodermal rather that endodermal tissue, unique in comparative phys. · symbionts make up a conservative 24.1%, and bacteriocytes make up 70.5% of the trophosomes volume!

Micrograph cross-section of lobule

Biochemical relationships between host and symbiont: Energy !
The highly reduced chemical sulfide (H2S) is used as an electron donor, which drives the reaction turning inorganic carbon into organic molecules by the basic reactions:
H2S + 1/2 O2 ---> So + H2O + energy So + 1 1/2 O2 + H2O ---> SO42- + 2 H+ + energy energy ---> HCO3- ---> [CHO's] (succinate, glutamate)

S-Sulfohemoglobin
Hemoglobins allow Riftia to maintain high concentrations of sulfide in its body fluids (up to 12 mmol/l in the blood; Pruski and Fiala-Medioni, 2003) Two were found in the vascular blood (V1 and V2), and one found in the coelomic fluid (C1) (A) HBL Hb V1 (B) Similar HBL V1 (C) Small ring HB V2 (D) Small ring HB C1

Conversion of Sulfide to hypo and thiotaurine: transport, storage, stabilization
· Lipid
soluble
· Safe accumulation · Osmotic balance · Low energy cost

Carbon fixation, transfer and utilization in the Riftia pachyptila symbiosis (Bright et al, 2000):
Trophosome
Lobule
Opithosome Glands

Carbon uptake and transfer from symbiont to host

Arginine Metabolism (Minic and Herve, 2003):
· Symbiont: nitrate ­ nitrite ­ ammonia · Host: carbonic anhydrase to maintain bicarbonate · Symbiont: ODase and ADase for polyamines · Analogous to protozoan parasites: Giardia lamblia,
Trichomonas vaginalis, and Tritrichomonas fetus
Who's using who?

Biosynthesis of Pyrimidine Nucleotides:
(Minic et al, 2001 and 2002)
· Symbiont: Controls the de novo pathway · Host: uses compounds produced by symbiont, uses "salvage" pathway · Also seen in protozoan parasites · Source of nitrogen and carbon for host

Millikan et al, 1999: Flagellin Gene!
Previous studies: FISH ­ symbiont not in gametes (horizontal transmission) Transient mouth and ciliated gut in larvae: Still no symbiont!
coli
Cloned a fliC flagellin gene Expressed gene in mutant E.
Negative control Positive control using S. typhimurium fliC gene

Methods for isolating R. pachyptila's free living symbiont: (Gros et al, 2003)
· Extracted sediment DNA · PCR detection of C. obicularis symbiont in gills of T. testudinum, sediment, and mangrove sediments · FISH probe (eubacteria) · FISH probe (Simco 2) · DAPI stain for viability Fish DAPI

Current Research: Hot off the Press!! "Stalking The Wild Symbiont" (Harmer and Cavanaugh, coming soon to a peer reviewed journal near you!)
· Collect basalts, sulfides, and other surfaces, as well as water samples from vent sites and peripheral locations · Analyze using molecular techniques · Preliminary data: Riftia symbiont or a similar phylotype is present on some of the tested surfaces · pressurized incubation chambers · FISH probes, etc... to characterize method of transmission

Literature Cited:
Bright, M., H. Keckeis, and C.R. Fisher. 2000. An autoradiographic examination of carbon fixation, transfer and utilization in the Riftia pachyptila symbiosis. Marine Biology. 136: pp. 621-632. Bright, M., and A. Sorgo. 2003. Ultrastructural reinvestigation of the trophosome in adults of Riftia pachyptila (Annelida, Siboglinidae). Invertebrate Biology. 122(4): pp. 347-368. Cavanaugh, C. M., S. L. Gardiner, M. L. Jones, H. W. Jannasch, and J. B. Waterbury. 1981. Prokaryotic cells in the hydrothermal vent tube worm Riftia pachyptila: possible chemoautotrophic symbionts. Science 213: pp. 340-342. Gros, O., M. Liberge, A. Heddi, C. Khatchadourian, and H. Felbeck. 2003. Detection of the Free-Living Forms of Sulfide-Oxidizing Gill Endosymbionts in the Lucinid Habitat (Thalassia testudinum Environment). Applied and Environmental Microbiology. 69(10): pp. 6264-6267. Hassler, D.R., L. Goehring, and C. Fisher. 2002. The Complex World along Mid-Ocean Ridges. Geotimes. December issue: pp. 14-18. Jones, M. L. 1981. Riftia pachyptila, new genus, new species, the vestimentiferan tubeworm from the Galapagos Rift geothermal vents. Proc. Biol. Soc. Wash. 93: pp. 1295-1313. Millikan, D.S., H. Felbeck, and J.L. Stein. 1999. Identification and Characterization of a Flagellin Gene from the Endosymbiont of the Hydrothermal Vent Tubeworm Riftia pachyptila. Applied and Environmental Microbiology. 65(7): pp. 3129-3133. Minic, Z., V. Simon, B. Penverne, F. Gaill, and G. Herve. 2001. Contribution of the Bacterial Endosymbiont to the Biosynthesis of Pyrimidine Nucleotides in the Deep-sea Tube Worm Riftia pachyptila. The Journal of Biological Chemistry. 276(26): pp. 23777-23784. Minic, Z., S. Pastra-Landis, F. Gaill, and G. Herve. 2002. Catabolism of Pyrimidine Nucleotides in the Deep-sea Tube Worm Riftia pachyptila. The Journal of Biological Chemistry. 277(1): pp. 127-134. Minic, Z., and G. Herve. 2003. Arginine Metabolism in the Deep Sea Tube Worm Riftia pachyptila and Its Bacterial Endosymbiont. The Journal of Biological Chemistry. 278(42): pp. 40527-40533. Pruski, A.M., and A. Fiala-Medioni. 2003. Stimulatory effect of sulphide on thiotaurine synthesis in three hydrothermal-vent species from the East Pacific Rise. The Journal of Experimental Biology. 206: pp. 2923-2930. Zal, F., F.H. Lallier, J.S. Wall, S.N. Vinogradov, and A. Toulmond. 1996. The Multi-hemoglobin System of the Hydrothermal Vent Tube Worm Riftia pachyptila I. REEXAMINATION OF THE NUMBER AND MASSES OF ITS CONSTITUENTS. Journal of Biochemistry. 271(15): pp. 8869-8874. Zal, F., E. Leize, F.H. Lallier, A. Toulmond, A.V. Dorsselaer, and J.J. Childress. 1998. S-Sulfohemoglobin and disulfide exchange: The mechanisms of sulfide binding by Riftia pachyptila hemoglobins. Proceedings of the National Academy of Sciences. 95(15): pp. 8997-9002.