Deep-sea environments are largely unexplored habitats where a surprising number of species may be found in large communities, thriving regardless of the darkness, extreme cold, and high pressure. Their unique geochemical features result in reducing environments rich in methane and sulfides, sustaining complex chemosynthetic ecosystems that represent one of the most surprising findings in oceans in the last 40 years. The deep-sea Lucky Strike hydrothermal vent field, located in the Mid Atlantic Ridge, is home to large vent mussel communities where Bathymodiolus azoricus represents the dominant faunal biomass, owing its survival to symbiotic associations with methylotrophic or methanotrophic and thiotrophic bacteria. The recent transcriptome sequencing and analysis of gill tissues from B. azoricus revealed a number of genes of bacterial origin, hereby analyzed to provide a functional insight into the gill microbial community. The transcripts supported a metabolically active microbiome and a variety of mechanisms and pathways, evidencing also the sulfur and methane metabolisms. Taxonomic affiliation of transcripts and 16S rRNA community profiling revealed a microbial community dominated by thiotrophic and methanotrophic endosymbionts of B. azoricus and the presence of a Sulfurovum-like epsilonbacterium.
References
[1]
Boutet, I.; Ripp, R.; Lecompte, O.; Dossat, C.; Corre, E.; Tanguy, A.; Lallier, F.H. Conjugating effects of symbionts and environmental factors on gene expression in deep-sea hydrothermal vent mussels. BMC Genomics 2011, 12.
[2]
De Chaine, E.G.; Bates, A.E.; Shank, T.M.; Cavanaugh, C.M. Off-axis symbiosis found: Characterization and biogeography of bacterial symbionts of Bathymodiolus mussels from Lost City hydrothermal vents. Environ. Microbiol. 2006, 8, 1902–1912, doi:10.1111/j.1462-2920.2005.01113.x.
[3]
Huber, J.A.; Mark Welch, D.B.; Morrison, H.G.; Huse, S.M.; Neal, P.R.; Butterfield, D.A.; Sogin, M.L. Microbial population structures in the deep marine biosphere. Science 2007, 318, 97–100.
[4]
Barry, J.P.; Buck, K.R.; Kochevar, R.K.; Nelson, D.C.; Fujiwara, Y.; Goffredi, S.K.; Hashimoto, J. Methane-based symbiosis in a mussel, Bathymodiolus platifrons, from cold seeps in Sagami Bay, Japan. Invertebr. Biol. 2002, 121, 47–54.
[5]
Dover, C.V. The Ecology of Deep-Sea Hydrothermal Vents; Princeton University Press: Princeton, NJ, USA, 2000.
[6]
Fiala-Médioni, A.; McKiness, Z.; Dando, P.; Boulegue, J.; Mariotti, A.; Alayse-Danet, A.; Robinson, J.; Cavanaugh, C. 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. 2002, 141, 1035–1043, doi:10.1007/s00227-002-0903-9.
[7]
Won, Y.-J.; Hallam, S.J.; O’Mullan, G.D.; Pan, I.L.; Buck, K.R.; Vrijenhoek, R.C. Environmental acquisition of thiotrophic endosymbionts by deep-sea mussels of the genus Bathymodiolus. Appl. Environ. Microbiol. 2003, 69, 6785–6792.
[8]
Duperron, S.; Lorion, J.; Samadi, S.; Gros, O.; Gaill, F. Symbioses between deep-sea mussels (Mytilidae: Bathymodiolinae) and chemosynthetic bacteria: Diversity, function and evolution. C. R. Biol. 2009, 332, 298–310, doi:10.1016/j.crvi.2008.08.003.
[9]
Page, H.M.; Fiala-Medioni, A.; Fisher, C.R.; Childress, J.J. Experimental evidence for filter-feeding by the hydrothermal vent mussel, Bathymodiolus thermophilus. Deep Sea Res. A Oceanogr. Res. Pap. 1991, 38, 1455–1461.
[10]
Distel, D.L.; Lee, H.K.; Cavanaugh, C.M. Intracellular coexistence of methano- and thioautotrophic bacteria in a hydrothermal vent mussel. Proc. Natl. Acad. Sci. USA 1995, 92, 9598–9602, doi:10.1073/pnas.92.21.9598.
[11]
De Long, E.F.; Preston, C.M.; Mincer, T.; Rich, V.; Hallam, S.J.; Frigaard, N.-U.; Martinez, A.; Sullivan, M.B.; Edwards, R.; Brito, B.R.; Chisholm, S.W.; Karl, D.M. Community genomics among stratified microbial assemblages in the ocean’s interior. Science 2006, 311, 496–503.
[12]
Konstantinidis, K.T.; Braff, J.; Karl, D.M.; DeLong, E.F. Comparative metagenomic analysis of a microbial community residing at a depth of 4,000 meters at station ALOHA in the North Pacific subtropical gyre. Appl. Environ. Microbiol. 2009, 75, 5345–5355.
[13]
Frias-Lopez, J.; Shi, Y.; Tyson, G.W.; Coleman, M.L.; Schuster, S.C.; Chisholm, S.W.; Delong, E.F. Microbial community gene expression in ocean surface waters. Proc. Natl. Acad. Sci. USA 2008, 105, 3805–3810.
[14]
Gifford, S.M.; Sharma, S.; Rinta-Kanto, J.M.; Moran, M.A. Quantitative analysis of a deeply sequenced marine microbial metatranscriptome. ISME J. 2011, 5, 461–472, doi:10.1038/ismej.2010.141.
[15]
Harada, M.; Yoshida, T.; Kuwahara, H.; Shimamura, S.; Takaki, Y.; Kato, C.; Miwa, T.; Miyake, H.; Maruyama, T. Expression of genes for sulfur oxidation in the intracellular chemoautotrophic symbiont of the deep-sea bivalve Calyptogena okutanii. Extremophiles 2009, 13, 895–903, doi:10.1007/s00792-009-0277-8.
[16]
Stewart, F.J.; Dmytrenko, O.; DeLong, E.F.; Cavanaugh, C.M. Metatranscriptomic analysis of sulfur oxidation genes in the endosymbiont of solemya velum. Front Microbiol. 2011, 2.
[17]
Bettencourt, R.; Pinheiro, M.; Egas, C.; Gomes, P.; Afonso, M.; Shank, T.; Santos, R.S. High-throughput sequencing and analysis of the gill tissue transcriptome from the deep-sea hydrothermal vent mussel Bathymodiolus azoricus. BMC Genomics 2010, 11.
[18]
Meyer, F.; Paarmann, D.; D’Souza, M.; Olson, R.; Glass, E.; Kubal, M.; Paczian, T.; Rodriguez, A.; Stevens, R.; Wilke, A.; et al. The metagenomics RAST server—A public resource for the automatic phylogenetic and functional analysis of metagenomes. BMC Bioinforma 2008, 9.
[19]
Poretsky, R.S.; Hewson, I.; Sun, S.; Allen, A.E.; Zehr, J.P.; Moran, M.A. Comparative day/night metatranscriptomic analysis of microbial communities in the North Pacific subtropical gyre. Environ. Microbiol. 2009, 11, 1358–1375, doi:10.1111/j.1462-2920.2008.01863.x.
[20]
Hewson, I.; Poretsky, R.S.; Dyhrman, S.T.; Zielinski, B.; White, A.E.; Tripp, H.J.; Montoya, J.P.; Zehr, J.P. Microbial community gene expression within colonies of the diazotroph, Trichodesmium, from the Southwest Pacific Ocean. ISME J. 2009, 3, 1286–1300, doi:10.1038/ismej.2009.75.
[21]
Dam, B.; Ghosh, W.; Das Gupta, S.K. Conjugative Type 4 secretion system of a novel large plasmid from the chemoautotroph Tetrathiobacter kashmirensis and construction of shuttle vectors for Alcaligenaceae. Appl. Environ. Microbiol. 2009, 75, 4362–4373.
[22]
Hurtado, L.A.; Mateos, M.; Lutz, R.A.; Vrijenhoek, R.C. Coupling of bacterial endosymbiont and host mitochondrial genomes in the hydrothermal vent clam Calyptogena magnifica. Appl. Environ. Microbiol. 2003, 69, 2058–2064, doi:10.1128/AEM.69.4.2058-2064.2003.
[23]
Degnan, P.H.; Leonardo, T.E.; Cass, B.N.; Hurwitz, B.; Stern, D.; Gibbs, R.A.; Richards, S.; Moran, N.A. Dynamics of genome evolution in facultative symbionts of aphids. Environ. Microbiol. 2010, 12, 2060–2069.
[24]
Newton, I.; Bordenstein, S. Correlations between bacterial ecology and mobile DNA. Curr. Microbiol. 2011, 62, 198–208.
[25]
Zander, U.; Faust, A.; Klink, B.U.; de Sanctis, D.; Panjikar, S.; Quentmeier, A.; Bardischewsky, F.; Friedrich, C.G.; Scheidig, A.J. Structural basis for the oxidation of protein-bound sulfur by the sulfur cycle molybdohemo-enzyme sulfane dehydrogenase SoxCD. J. Biol. Chem. 2011, 286, 8349–8360.
[26]
Frigaard, N.-U.; Dahl, C. Sulfur Metabolism in Phototrophic Sulfur Bacteria. In Advances in Microbial Physiology; Poole, R.K., Ed.; Academic Press: New York, NY, USA, 2008; Volume 54, pp. 103–200.
Beller, H.R.; Chain, P.S.G.; Letain, T.E.; Chakicherla, A.; Larimer, F.W.; Richardson, P.M.; Coleman, M.A.; Wood, A.P.; Kelly, D.P. The genome sequence of the obligately chemolithoautotrophic, facultatively anaerobic bacterium Thiobacillus denitrificans. J. Bacteriol. 2006, 188, 1473–1488.
[29]
Kappler, U.; Dahl, C. Enzymology and molecular biology of prokaryotic sulfite oxidation. FEMS Microbiol. Lett. 2001, 203, 1–9.
[30]
Loy, A.; Duller, S.; Baranyi, C.; Mu？mann, M.; Ott, J.; Sharon, I.; Béjà, O.; Le Paslier, D.; Dahl, C.; Wagner, M. Reverse dissimilatory sulfite reductase as phylogenetic marker for a subgroup of sulfur-oxidizing prokaryotes. Environ. Microbiol. 2009, 11, 289–299, doi:10.1111/j.1462-2920.2008.01760.x.
[31]
Duperron, S.; Sibuet, M.; MacGregor, B.J.; Kuypers, M.M.M.; Fisher, C.R.; Dubilier, N. Diversity, relative abundance and metabolic potential of bacterial endosymbionts in three Bathymodiolus mussel species from cold seeps in the Gulf of Mexico. Environ. Microbiol. 2007, 9, 1423–1438, doi:10.1111/j.1462-2920.2007.01259.x.
[32]
Esparza, M.; Cárdenas, J.; Bowien, B.; Jedlicki, E.; Holmes, D.S. Genes and pathways for CO2 fixation in the obligate, chemolithoautotrophic acidophile, Acidithiobacillus ferrooxidans, Carbon fixation in A. ferrooxidans. BMC Microbiol. 2010, 10.
[33]
Byrne, N.; Strous, M.; Crépeau, V.; Kartal, B.; Birrien, J.-L.; Schmid, M.; Lesongeur, F.; Schouten, S.; Jaeschke, A.; Jetten, M.; et al. Presence and activity of anaerobic ammonium-oxidizing bacteria at deep-sea hydrothermal vents. ISME J. 2009, 3, 117–123, doi:10.1038/ismej.2008.72.
Petersen, J.M.; Zielinski, F.U.; Pape, T.; Seifert, R.; Moraru, C.; Amann, R.; Hourdez, S.; Girguis, P.R.; Wankel, S.D.; Barbe, V.; et al. Hydrogen is an energy source for hydrothermal vent symbioses. Nature 2011, 476, 176–180, doi:10.1038/nature10325.
[36]
Bettencourt, R. Department of Oceanography and Fisheries, University of Azores, Horta, Portugal. Unpublished work, 2012.
[37]
Biddle, J.F.; Fitz-Gibbon, S.; Schuster, S.C.; Brenchley, J.E.; House, C.H. Metagenomic signatures of the peru margin subseafloor biosphere show a genetically distinct environment. Proc. Natl. Acad. Sci. USA 2008, 105, 10583–10588.
[38]
McCarren, J.; Becker, J.W.; Repeta, D.J.; Shi, Y.; Young, C.R.; Malmstrom, R.R.; Chisholm, S.W.; DeLong, E.F. Microbial community transcriptomes reveal microbes and metabolic pathways associated with dissolved organic matter turnover in the sea. Proc. Natl. Acad. Sci. USA 2010, 107, 16420–16427.
Nakagawa, S.; Takaki, Y.; Shimamura, S.; Reysenbach, A.-L.; Takai, K.; Horikoshi, K. Deep-sea vent epsilon-proteobacterial genomes provide insights into emergence of pathogens. Proc. Natl. Acad. Sci. USA. 2007, 104, 12146–12150.
[41]
Kato, S.; Kobayashi, C.; Kakegawa, T.; Yamagishi, A. Microbial communities in iron-silica-rich microbial mats at deep-sea hydrothermal fields of the Southern Mariana Trough. Environ. Microbiol. 2009, 11, 2094–2111, doi:10.1111/j.1462-2920.2009.01930.x.
[42]
Sylvan, J.B.; Toner, B.M.; Edwards, K.J. Life and death of deep-sea vents: Bacterial diversity and ecosystem succession on inactive hydrothermal sulfides. mBio 2012, 3.
[43]
Stingl, U.; Desiderio, R.A.; Cho, J.-C.; Vergin, K.L.; Giovannoni, S.J. The SAR92 clade: An abundant coastal clade of culturable marine bacteria possessing proteorhodopsin. Appl. Environ. Microbiol. 2007, 73, 2290–2296.
[44]
Buchan, A.; González, J.M.; Moran, M.A. Overview of the marine Roseobacter lineage. Appl. Environ. Microbiol. 2005, 71, 5665–5677, doi:10.1128/AEM.71.10.5665-5677.2005.
[45]
Letunic, I.; Bork, P. Interactive tree of life v2: Online annotation and display of phylogenetic trees made easy. Nucleic Acids Res. 2011, 39, W475–W478, doi:10.1093/nar/gkr201.
[46]
Cole, J.R.; Wang, Q.; Cardenas, E.; Fish, J.; Chai, B.; Farris, R.J.; Kulam-Syed-Mohideen, A.S.; McGarrell, D.M.; Marsh, T.; Garrity, G.M.; et al. The ribosomal database project: Improved alignments and new tools for rRNA analysis. Nucleic Acids Res. 2009, 37, D141–D145.
[47]
Xu, Y.; Nogi, Y.; Kato, C.; Liang, Z.; Rüger, H.-J.; de Kegel, D.; Glansdorff, N. Psychromonas profunda sp. nov., a psychropiezophilic bacterium from deep Atlantic sediments. Int. J. Syst. Evol. Microbiol. 2003, 53, 527–532, doi:10.1099/ijs.0.02227-0.
[48]
Tokuda, G.; Yamada, A.; Nakano, K.; Arita, N.O.; Yamasaki, H. Colonization of Sulfurovum sp. on the gill surfaces of Alvinocaris longirostris, a deep-sea hydrothermal vent shrimp. Mar. Ecol. 2007, 29, 106–114.
[49]
Husmann, G.; Gerdts, G.; Wichels, A. Spirochetes in crystalline styles of marine bivalves: Group-specific PCR detection and 16S rRNA sequence analysis. J. Shellfish Res. 2010, 29, 1069–1075, doi:10.2983/035.029.0409.
[50]
Takai, K.; Campbell, B.J.; Cary, S.C.; Suzuki, M.; Oida, H.; Nunoura, T.; Hirayama, H.; Nakagawa, S.; Suzuki, Y.; Inagaki, F.; et al. Enzymatic and genetic characterization of carbon and energy metabolisms by deep-sea hydrothermal chemolithoautotrophic isolates of epsilonproteobacteria. Appl. Environ. Microbiol. 2005, 71, 7310–7320, doi:10.1128/AEM.71.11.7310-7320.2005.
[51]
Corre, E.; Reysenbach, A.L.; Prieur, D. Epsilon-proteobacterial diversity from a deep-sea hydrothermal vent on the Mid-Atlantic Ridge. FEMS Microbiol. Lett. 2001, 205, 329–335.
[52]
Suzuki, Y.; Kojima, S.; Sasaki, T.; Suzuki, M.; Utsumi, T.; Watanabe, H.; Urakawa, H.; Tsuchida, S.; Nunoura, T.; Hirayama, H.; et al. Host-symbiont relationships in hydrothermal vent gastropods of the genus Alviniconcha from the Southwest Pacific. Appl. Environ. Microbiol. 2006, 72, 1388–1393, doi:10.1128/AEM.72.2.1388-1393.2006.
[53]
Hügler, M.; G？rtner, A.; Imhoff, J.F. Functional genes as markers for sulfur cycling and CO2 fixation in microbial communities of hydrothermal vents of the Logatchev field. FEMS Microbiol. Ecol. 2010, 73, 526–537.
[54]
Petersen, J.M.; Ramette, A.; Lott, C.; Cambon-Bonavita, M.-A.; Zbinden, M.; Dubilier, N. Dual symbiosis of the vent shrimp Rimicaris exoculata with filamentous gamma- and epsilonproteobacteria at four Mid-Atlantic Ridge hydrothermal vent fields. Environ. Microbiol. 2010, 12, 2204–2218.
[55]
Chevreux, B.; Pfisterer, T.; Drescher, B.; Driesel, A.J.; Müller, W.E.G.; Wetter, T.; Suhai, S. Using the miraEST assembler for reliable and automated mRNA transcript assembly and SNP detection in sequenced ESTs. Genome Res. 2004, 14, 1147–1159, doi:10.1101/gr.1917404.
[56]
DeepSeaVent Database. Available online: http://transcriptomics.biocant.pt:8080/deepSeaVent (accessed on 19 June 2010).
[57]
Stajich, J.E.; Block, D.; Boulez, K.; Brenner, S.E.; Chervitz, S.A.; Dagdigian, C.; Fuellen, G.; Gilbert, J.G.R.; Korf, I.; Lapp, H.; et al. The Bioperl toolkit: Perl modules for the life sciences. Genome Res. 2002, 12, 1611–1618, doi:10.1101/gr.361602.
[58]
MG-RAST Metagenomics Analysis Server. Available online: http://metagenomics.anl.gov (accessed on 5 February 2012).
[59]
Wang, Y.; Qian, P.-Y. Conservative fragments in bacterial 16S rRNA genes and primer design for 16S ribosomal DNA amplicons in metagenomic studies. PLoS One 2009, 4.
[60]
Edgar, R.C. Search and clustering orders of magnitude faster than BLAST. Bioinformatics 2010, 26, 2460–2461, doi:10.1093/bioinformatics/btq461.
[61]
Huang, X.; Madan, A. CAP3: A DNA sequence assembly program. Genome Res. 1999, 9, 868–877, doi:10.1101/gr.9.9.868.
[62]
Felsenstein, J. PHYLIP: Phylogenetic Inference Package, Version 3.5c; University of Washington: Seattle, WA, USA, 1993.
[63]
Tamura, K.; Peterson, D.; Peterson, N.; Stecher, G.; Nei, M.; Kumar, S. MEGA5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol. Biol. Evol. 2011, 28, 2731–2739, doi:10.1093/molbev/msr121.
[64]
Jukes, T.; Cantor, C.R. Evolution of Protein Molecules. In Mammalian Protein Metabolism; Munro, H.N., Ed.; Academic Press: New York, NY, USA, 1969; Volume 3, pp. 121–132.
[65]
Felsenstein, J. Confidence limits on Phylogenies: An approach using bootstrap. Evolution 1985, 39, 783–791, doi:10.2307/2408678.
[66]
Mid-Atlantic Ridge 2008 (MAR08) Homepage. Available online: http://www.deepseavoyage.research.pdx.edu (accessed on 5 February 2010).
