Advertisement

Symbiosis

, Volume 63, Issue 1, pp 19–29 | Cite as

Comparative analysis of symbiont ratios and gene expression in natural populations of two Bathymodiolus mussel species

  • H. Guezi
  • I. Boutet
  • A. C. Andersen
  • F. H. Lallier
  • A. Tanguy
Article

Abstract

Bathymodiolus mussels associated with deep-sea hydrothermal vents and cold seeps harbor chemosynthetic endosymbiotic bacteria in bacteriocytes located in the gill epithelium. Two distinct morphotypes of γ-proteobacteria, sulfur- and methane-oxidizing, have been identified and form a dual symbiosis in B. azoricus mussels from the Mid-Atlantic Ridge and in B. aff. boomerang from cold seeps in the Gulf of Guinea. Thiotrophic bacteria (SOX) are capable of fixing CO2 in the presence of sulfide or thiosulfate and methanotrophic bacteria (MOX) use methane both as a carbon and an energy source. In this study we used quantitative real-time PCR to test whether symbiont abundance and gene expression varied between the two mussel species. Results showed that B. azoricus from two hydrothermal sites had similar ratios and gene expression pattern for both symbiont types. In B. aff. boomerang, SOX ratio and ATP sulfurylase gene expression show differences between specimens collected on the different sites. Analysis of symbiont ratios in both species indicated a clear dominance of MOX symbionts in B. aff. boomerang and SOX symbionts in B. azoricus. We also evidenced that the species from the deeper sites (B. aff. boomerang) and mussels collected from sulfur and methane rich habitats showed higher symbiont ratio suggesting that environmental parameters may have significant impacts on the symbiont ratios in Bathymodiolus mussels.

Keywords

Hydrothermal vent Cold seeps Bathymodiolus Symbiont Fish 

Abbreviations

DAPI

4’,6-DiAmidino-2-Phenyl-Indole double-stranded DNA staining

Cy3 and Cy5

Cyanine dyes

FISH

Fluorescencent In Situ Hybridization

MOX

Methane OXidizing bacteria

pmoA

particulate methane oxygenase subunit A

ROV

Remotely Operated Vehicle

SOX

Sulfur OXidizing bacteria

Notes

Acknowledgments

We thank the crew and pilots of the NO Pourquoi Pas? and the ROV Victor 6,000 for their assistance and technical support, as well as the chief scientist Dr. Karine Olu Le Roy during the cruise WACS (2011) and Dr. François Lallier, during the cruise BIOBAZ (2011). We thank Dr. Sophie Le Panse, manager of Optical Imaging Platform Merimage of the Station Biologique de Roscoff, for having introduced us (HG and AA) to the confocal microscopy. We thank the anonymous referees for useful comments and suggestions. This work was funded by the Région Bretagne with the help of GIS Europole Mer (HG), and the JST/CNRS Bathymodiolus program (AT, FHL).

References

  1. Amann R, Binder BJ, Olson RJ, Chisholm SW, Devereux R, Stahl DA (1990) Combination of 16S rRNA-targeted oligonucleotide probes withflow cytometry for analyzing mixed microbial populations. Appl Environ Microbiol 56:1919–1925PubMedCentralPubMedGoogle Scholar
  2. Barry JP, Buck KR, Kochevar RK, Nelson DC, Fujiwara Y, Goffredi SK, Hashimoto J (2002) Methane-based symbiosis in a mussel, Bathymodiolus platifrons, from cold seeps in sagami bay, Japan. Invert Biol 121:47–54CrossRefGoogle Scholar
  3. Boetius A, Suess E (2004) Hydrate ridge: a natural laboratory for the study of microbial life fueled by methane from near-surface gas hydrates. Chem Geol 205:291–310CrossRefGoogle Scholar
  4. Boutet I, Ripp R, Lecompte O, Dossat C, Corre E, Tanguy A, Lallier F (2011) Conjugating effects of symbionts and environmental factors on gene expression in deep-sea hydrothermal vent mussels. BMC Genomics 12:530PubMedCentralPubMedCrossRefGoogle Scholar
  5. Caro A, Gros O, Got P, De Wit R, Troussellier M (2007) Characterization of the population of the sulfur-oxidizing symbiont of Codakia orbicularis (Bivalvia, Lucinidae) by single-cell analyses. Appl Environ Microbiol 73:2101–2109PubMedCentralPubMedCrossRefGoogle Scholar
  6. Cavanaugh CM (1983) Symbiotic chemoautotrophic bacteria in marine invertebrates from sulfide-rich habitats. Nature 302:58–61CrossRefGoogle Scholar
  7. Cavanaugh CM, McKiness ZP, Newton ILG, Stewart FJ (2006) Marine chemosynthetic symbioses. In: Dworkin M, Falkow S, Rosenberg E, Schleifer K-H, Stackebrandt E (eds) The prokaryotes. Springer New York, New York, pp 475–507CrossRefGoogle Scholar
  8. Charlou JL, Donval JP, Fouquet Y, Jean-Baptiste P, Holm N (2002) Geochemistry of high H2 and CH4 vent fluids issuing from ultramafic rocks at the rainbow hydrothermal field (36°14’ N, MAR). Chem Geol 191:345–359CrossRefGoogle Scholar
  9. Charlou JL, Donval J, Fouquet Y, Ondreas H, Knoery J, Cochonat P, Levaché D, Poirier Y, Jean-Baptiste P, Fourré E, Chazallon B (2004) Physical and chemical characterization of gas hydrates and associated methane plumes in the Congo–Angola basin. Chem Geol 205:405–425CrossRefGoogle Scholar
  10. Childress JJ, Fisher CR, Brooks JM, Kennicutt MC, Bidigare R, Anderson AE (1986) A methanotrophic marine Molluscan (Bivalvia, Mytilidae) symbiosis: mussels fueled by gas. Science 233:1306–1308PubMedCrossRefGoogle Scholar
  11. Childress JJ, Fisher CR, Favuzzi JA, Kochevar RE, Sanders NK, Alayse AM (1991) Sulfide-driven autotrophic balance in the bacterial symbiont-containing hydrothermal vent tubeworm, Riftia pachyptila Jones. Biol Bull 180:135–153CrossRefGoogle Scholar
  12. Deana A, Belasco J (2005) Lost in translation: the influence of ribosomes on bacterial mRNA decay. Genes Develop 19:2526–2533Google Scholar
  13. De Beer D, Sauter EJ, Niemann H, Kaul N, Foucher J-P, Witte U, Schlüter M, Boetius A (2006) In situ fluxes and zonation of microbial activity in surface sediments of the haakon mosby mud volcano. Lim Oceano 51:1315–1331CrossRefGoogle Scholar
  14. DeChaine EG, Cavanaugh CM (2006) Symbioses of methanotrophs and deep-sea mussels (Mytilidae: Bathymodiolinae). Molecular basis of symbiosis. Springer, pp 227–249Google Scholar
  15. Desbruyères D, Biscoito M, Caprais J-C, Colaço A, Comtet T, Crassous P, Fouquet Y, Khripounoff A, Le Bris N, Olu K, Riso R, Sarradin P-M, Segonzac M, Vangriesheim A (2001) Variations in deep-sea hydrothermal vent communities on the mid-atlantic ridge near the azores plateau. Deep Sea Research Part I Oceano Res Pap 48:1325–1346CrossRefGoogle Scholar
  16. Dickens GR (2003) Rethinking the global carbon cycle with a large, dynamic and microbially mediated gas hydrate capacitor. Earth Plan Sci Lett 213:169–183CrossRefGoogle Scholar
  17. Distel DL, Lee HK, Cavanaugh CM (1995) Intracellular coexistence of methano-and thio-autotrophic bacteria in a hydrothermal vent mussel. Proc Natl Acad Sci 92:9598–9602PubMedCentralPubMedCrossRefGoogle Scholar
  18. Douville E, Charlou J, Oelkers E, Bienvenu P, Jove Colon C, Donval J, Fouquet Y, Prieur D, Appriou P (2002) The rainbow vent fluids (36°14’N, MAR): the influence of ultramafic rocks and phase separation on trace metal content in mid-atlantic Ridge hydrothermal fluids. Chem Geol 184:37–48CrossRefGoogle Scholar
  19. Dubilier N, Bergin C, Lott C (2008) Symbiotic diversity in marine animals: the art of harnessing chemosynthesis. Nat Rev Microbiol 6:725–740PubMedCrossRefGoogle Scholar
  20. Duperron S, Nadalig T, Caprais J-C, Sibuet M, Fiala-Medioni A, Amann R, Dubilier N (2005) Dual symbiosis in a Bathymodiolus sp. mussel from a methane seep on the gabon continental margin (southeast atlantic): 16S rRNA phylogeny and distribution of the symbionts in gills. Appl Env Microbiol 71:1694–1700CrossRefGoogle Scholar
  21. Duperron S, Bergin C, Zielinski F, Blazejak A, Pernthaler A, McKiness ZP, DeChaine E, Cavanaugh CM, Dubilier N (2006) A dual symbiosis shared by two mussel species, Bathymodiolus azoricus and Bathymodiolus puteoserpentis (Bivalvia: Mytilidae), from hydrothermal vents along the northern mid-atlantic ridge. Environ Microbiol 8:1441–1447PubMedCrossRefGoogle Scholar
  22. Duperron S, Fiala-Médioni A, Caprais JC, Olu K, Sibuet M (2007) Evidence for chemoautotrophic symbiosis in a mediterranean cold seep clam (Bivalvia: Lucinidae): comparative sequence analysis of bacterial 16S rRNA, APS reductase and RubisCO genes: symbiosis in a cold-seep lucinid. FEMS Microbiol Ecol 59:64–70PubMedCrossRefGoogle Scholar
  23. Duperron S, Halary S, Lorion J, Sibuet M, Gaill F (2008) Unexpected co-occurrence of six bacterial symbionts in the gills of the cold seep mussel Idas sp. (Bivalvia: Mytilidae). Environ Microbiol 10:433–445PubMedCrossRefGoogle Scholar
  24. Duperron S, Guezi H, Gaudron SM, Pop Ristova P, Wenzhöfer F, Boetius A (2011) Relative abundances of methane- and sulphur-oxidising symbionts in the gills of a cold seep mussel and link to their potential energy sources. Geobiology 9:481–491PubMedCrossRefGoogle Scholar
  25. Felbeck H, Childress JJ, Somero GN (1981) Calvin-Benson cycle and sulfide oxidation enzymes in animals from sulfide-rich habitats. Nature 293:291–293CrossRefGoogle Scholar
  26. Fiala-Médioni A, McKiness Z, Dando P, Boulegue J, Mariotti A, Alayse-Danet A, Robinson J, Cavanaugh C (2002) Ultrastructural, biochemical, and immunological characterization of two populations of the mytilid mussel Bathymodiolus azoricus from the mid-atlantic Ridge: evidence for a dual symbiosis. Mar Biol 141:1035–1043CrossRefGoogle Scholar
  27. Fisher CR, Childress JJ, Oremland RS, Bidigare RR (1987) The importance of methane and thiosulfate in the metabolism of the bacterial symbionts of two deep-sea mussels. Mar Biol 96:59–71CrossRefGoogle Scholar
  28. Fisher CR, Brooks JM, Vodenichar JS, Zande JM, Childress JJ (1993) The Co-occurrence of methanotrophic and chemoautotrophic sulfur-oxidizing bacterial symbionts in a deep-sea mussel. Mar Ecol 14:277–289CrossRefGoogle Scholar
  29. Fujiwara Y, Takai K, Uematsu K, Tsuchida S, Hunt JC, Hashimoto J (2000) Phylogenetic characterization of endosymbionts in three hydrothermal vent mussels: influence on host distributions. Mar Ecol Prog Ser 208:147–155CrossRefGoogle Scholar
  30. Génio L, Johnson SB, Vrijenhoek RC, Cunha MR, Tyler PA, Kiel S, Little CT (2008) New record of “Bathymodiolus” mauritanicus cosel 2002 from the gulf of cadiz (NE atlantic) mud volcanoes. J Shellfish Res 27:53–61CrossRefGoogle Scholar
  31. Geret F, Riso R, Sarradin PM, Caprais JC, Cosson RP (2002) Metal bioaccumulation and storage forms in the shrimp, Rimicaris exoculata, from the rainbow hydrothermal field (mid-atlantic ridge); preliminary approach to the fluid–organism relationship. Cah Biol Mar 43:43–52Google Scholar
  32. German CR, Lin J (2004) The thermal structure of the oceanic crust, ridge-spreading and hydrothermal circulation: how well do we understand their inter-connections? In: German CR, Lin J, Parson LM (eds) Geophysical monograph series. American Geophysical Union, Washington, pp 1–18Google Scholar
  33. Girguis PR, Childress JJ (2006) Metabolite stoichiometry and chemoautotrophic function of the hydrothermal vent tubeworm Riftia pachyptila: responses to environmental variations in substrate concentrations and temperature. J Exp Biol 209:3516–3528Google Scholar
  34. Halary S, Riou V, Gaill F, Boudier T, Duperron S (2008) 3D FISH for the quantification of methane- and sulphur-oxidizing endosymbionts in bacteriocytes of the hydrothermal vent mussel bathymodiolus azoricus. ISME J 2:284–292PubMedCrossRefGoogle Scholar
  35. Kádár E, Bettencourt R, Costa V, Santos RS, Lobo-da-Cunha A, Dando P (2005) Experimentally induced endosymbiont loss and re-acquirement in the hydrothermal vent bivalve bathymodiolus azoricus. J Exp Mar Biol Ecol 318:99–110CrossRefGoogle Scholar
  36. Kenk VC, Wilson BR (1985) A new mussel (bivalvia, mytilidae) from hydrothermal vents in the galapagos rift zone. Malacologia 26:253–271Google Scholar
  37. Kojima S (2002) Deep-sea chemoautosynthesis-based communities in the northwestern pacific. J Oceano 58:343–363CrossRefGoogle Scholar
  38. Le Bris N, Sarradin PM, Caprais JC (2003) Contrasted sulphide chemistries in the environment of 13 °N EPR vent fauna. Deep Sea Research Part I Oceano Res Pap 50:737–747CrossRefGoogle Scholar
  39. Le Pennec M, Diouris M, Herry A (1988) Endocytosis and lysis of bacteria in gill epithelium of bathymodiolus thermophilus, thyasira flexuosa and lucinella divaricata (bivalve, molluscs). J Shellfish Res 7:483–489Google Scholar
  40. Levin LA (2005) Ecology of cold seep sediments: interactions of fauna with flow, chemistry, and microbes. Oceanogr Mar Biol Ann Rev 43:1–46Google Scholar
  41. Lonsdale P, Becker K (1985) Hydrothermal plumes, hot springs, and conductive heat flow in the Southern Trough of guaymas basin. Earth Plan Sci Lett 73:211–225CrossRefGoogle Scholar
  42. MacDonald IR, Boland GS, Baker JS, Brooks JM, Kennicutt MC II, Bidigare RR (1989) Gulf of Mexico hydrocarbon seep communities. Mar Biol 101:235–247CrossRefGoogle Scholar
  43. Marcon Y, Sahling H, Allais AG, Bohrmann G, Olu K (2013) Distribution and temporal variation of mega-fauna at the regab pockmark (northern Congo fan), based on a comparison of videomosaics and geographic information systems analyses. Mar Ecol SSN 0173–9565:1–19. doi: 10.1111/maec.12056 Google Scholar
  44. Martins I, Colaço A, Dando PR, Martins I, Desbruyères D, Sarradin P-M, Marques JC, Serrão-Santos R (2008) Size-dependent variations on the nutritional pathway of bathymodiolus azoricus demonstrated by a C-flux model. Ecol Model 217:59–71CrossRefGoogle Scholar
  45. Nelson DC, Fisher CR (1995) Chemoautotrophic and methanotrophic endosymbiotic bacteria at deep-sea vents and seeps. In Microbiology of deep-sea hydrothermal vents Karl DM (ed) Boca Raton FL: CRC Press inc pp 125–167Google Scholar
  46. Nelson DC, Hagen KD, Edwards DB (1995) The gill symbiont of the hydrothermal vent mussel bathymodiolus thermophilus is a psychrophilic, chemoautotrophic, sulfur bacterium. Mar Biol 121:487–495CrossRefGoogle Scholar
  47. Niemann H, Lösekann T, de Beer D, Elvert M, Nadalig T, Knittel K, Amann R, Sauter EJ, Schlüter M, Klages M, Foucher JP, Boetius A (2006) Novel microbial communities of the Haakon Mosby mud volcano and their role as a methane sink. Nature 443:854–858PubMedCrossRefGoogle Scholar
  48. Olu-Le Roy K, Caprais J-C, Fifis A, Fabri M-C, Galéron J, Budzinsky H, Le Ménach K, Khripounoff A, Ondreas H, Sibuet M (2007) Cold-seep assemblages on a giant pockmark off West Africa: spatial patterns and environmental control. Mar Ecol 28:115–130CrossRefGoogle Scholar
  49. Ondréas H, Olu K, Fouquet Y, Charlou JL, Gay A, Dennielou B, Donval JP, Fifis A, Nadalig T, Cochonat P, Cauquil E, Bourillet JF, Moigne ML, Sibuet M (2005) ROV study of a giant pockmark on the Gabon continental margin. Geo Mar Lett 25:281–292CrossRefGoogle Scholar
  50. Pimenov NV, Kalyuzhnaya MG, Khmelenina VN, Mityushina LL, Trotsenko YA (2002) Utilization of methane and carbon dioxide by symbiotrophic bacteria in gills of Mytilidae (bathymodiolus) from the Rainbow and Logachev hydrothermal fields on the mid-atlantic ridge. Microbiology 71:587–594CrossRefGoogle Scholar
  51. Riou V, Duperron S, Halary S, Dehairs F, Bouillon S, Martins I, Colaço A, Serrão Santos R (2010) Variation in physiological indicators in Bathymodiolus azoricus (bivalvia: mytilidae) at the Menez Gwen mid-atlantic ridge deep-sea hydrothermal vent site within a year. Mar Environ Res 70:264–271PubMedCrossRefGoogle Scholar
  52. Robinson JJ, Polz MF, Fiala-Medioni A, Cavanaugh CM (1998) Physiological and immunological evidence for two distinct C1-utilizing pathways in bathymodiolus puteoserpentis (bivalvia: mytilidae), a dual endosymbiotic mussel from the mid-atlantic ridge. Mar Biol 132:625–633CrossRefGoogle Scholar
  53. Salerno JL, Macko SA, Hallam SJ, Bright M, Won Y-J, McKiness Z, Van Dover CL (2005) Characterization of symbiont populations in life-history stages of mussels from chemosynthetic environments. Biol Bull 208:145–155PubMedCrossRefGoogle Scholar
  54. Sibuet M, Olu K (1998) Biogeography, biodiversity and fluid dependence of deep-sea cold-seep communities at active and passive margins. Deep-Sea Res II 45:517–567CrossRefGoogle Scholar
  55. Thorsen BK, Enger O, Norland S, Hoff KJ (1992) Long-term starvation survival of Yersinia ruckeri at different salinities studied by microscopical and flow cytometric methods. Appl Environ Microbiol 58:1624–1628PubMedCentralPubMedGoogle Scholar
  56. Tivey MK (1995). Modeling chimney growth and associated fluid flow at seafloor hydrothermal vent sites. In Humphris SE, Zierenberg RA, Mullineaux LS, and Thomson RE. (Eds.), Seafloor Hydrothermal Systems: Physical, Chemical, Biol Geol Interac, Am. Geophys. Union, Geophys. Monogr., 91:158–177Google Scholar
  57. Van Dover CL (2000) The ecology of deep-sea hydrothermal vents. Princeton University Press, PrincetonGoogle Scholar
  58. Van Dover CL (2002) Evolution and biogeography of deep-sea vent and seep invertebrates. Science 295:1253–1257PubMedCrossRefGoogle Scholar
  59. Van Dover CL, Trask JL (2000) Diversity at deep-sea hydrothermal vent and intertidal mussel beds. Mar Ecol Prog Ser 195:169–178CrossRefGoogle Scholar
  60. Van Dover CL, Aharon P, Bernhard JM, Caylor E, Doerries M, Flickinger W, Gilhooly W, Goffredi SK, Knick KE, Macko SA, Rapoport S, Raulfs EC, Ruppel C, Salerno JL, Seitz RD, Sen Gupta BK, Shank T, Turnipseed M, Vrijenhoek R (2003) Blake ridge methane seeps: characterization of a soft-sediment, chemosynthetically based ecosystem. Deep Sea Research Part I Oceano Res Pap 50:281–300CrossRefGoogle Scholar
  61. Vanreusel A, Fonseca G, Danovaro R, Da Silva MC, Esteves AM, Ferrero T, Gad G, Galtsova V, Gambi C, Da Fonsêca GV, Ingels J, Ingole B, Lampadariou N, Merckx B, Miljutin D, Miljutina M, Muthumbi A, Netto S, Portnova D, Radziejewska T, Raes M, Tchesunov A, Vanaverbeke J, Van Gaever S, Venekey V, Bezerra TN, Flint H, Copley J, Pape E, Zeppilli D, Martinez PA, Galeron J (2010) The contribution of deep-sea macrohabitat heterogeneity to global nematode diversity: Nematode diversity and habitat heterogeneity. Mar Ecol 31:6–20CrossRefGoogle Scholar
  62. Von Damm KL (1995) Controls on the chemistry and temporal variability of seafloor hydrothermal fluids. In: Humphris SE, Zierenberg RA, Mullineaux LS, Thomson RE (eds) Geophysical monograph series. American Geophysical Union, Washington, pp 222–247Google Scholar
  63. Wendeberg A, Zielinski FU, Borowski C, Dubilier N (2012) Expression patterns of mRNAs for methanotrophy and thiotrophy in symbionts of the hydrothermalvent mussel Bathymodiolus puteoserpentis. ISME J 6:104–112Google Scholar
  64. Won YJ, Hallam SJ, O’Mullan GD, Pan IL, Buck KR, Vrijenhoek RC (2003) Environmental acquisition of thiotrophic endosymbionts by deep-sea mussels of the genus bathymodiolus. Appl Environ Microbiol 69:6785–6792PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  • H. Guezi
    • 1
    • 2
  • I. Boutet
    • 1
    • 2
  • A. C. Andersen
    • 1
    • 2
  • F. H. Lallier
    • 1
    • 2
  • A. Tanguy
    • 1
    • 2
  1. 1.CNRS, UMR 7144, Adaptation et Diversité en Milieu MarinRoscoffFrance
  2. 2.Sorbonne UniversitésRoscoffFrance

Personalised recommendations