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Deep-sea mussels of the genus Bathymodiolus (Bivalvia: Mytilidae) harbor symbiotic bacteria in their gills and are among the dominant invertebrate species at cold seeps and hydrothermal vents. An undescribed Bathymodiolus species was collected at a depth of 3,150 m in a newly discovered cold seep area on the southeast Atlantic margin, close to the Zaire channel. Transmission electron microscopy, comparative 16S rRNA analysis, and fluorescence in situ hybridization indicated that this Bathymodiolus sp. lives in a dual symbiosis with sulfide-and methane-oxidizing bacteria. A distinct distribution pattern of the symbiotic bacteria in the gill epithelium was observed, with the thiotrophic symbiont dominating the apical region and the methanotrophic symbiont more abundant in the basal region of the bacteriocytes. No variations in this distribution pattern or in the relative abundances of the two symbionts were observed in mussels collected from three different mussel beds with methane concentrations ranging from 0.7 to 33.7 M. The 16S rRNA sequence of the methanotrophic symbiont is most closely related to those of known methanotrophic symbionts from other bathymodiolid mussels. Surprisingly, the thiotrophic Bathymodiolus sp. 16S rRNA sequence does not fall into the monophyletic group of sequences from thiotrophic symbionts of all other Bathymodiolus hosts. While these mussel species all come from vents, this study describes the first thiotrophic sequence from a seep mussel and shows that it is most closely related (99% sequence identity) to an environmental clone sequence obtained from a hydrothermal plume near Japan.

Cold seeps, located along the Sonora Margin transform fault in the Guaymas Basin, were extensively explored during the 'BIG' cruise in June 2010. They present a seafloor mosaic pattern consisting of different faunal assemblages and microbial mats. To investigate this mostly unknown cold and hydrocarbon-rich environment, geochemical and microbiological surveys of the sediments underlying two microbial mats and a surrounding macrofaunal habitat were analyzed in detail. The geochemical measurements suggest biogenic methane production and local advective sulfate-rich fluxes in the sediments. The distributions of archaeal communities, particularly those involved in the methane cycle, were investigated at different depths (surface to 18 cm below the sea floor (cmbsf)) using complementary molecular approaches, such as Automated method of Ribosomal Intergenic Spacer Analysis (ARISA), 16S rRNA libraries, fluorescence in situ hybridization and quantitative polymerase chain reaction with new specific primer sets targeting methanogenic and anaerobic methanotrophic lineages. Molecular results indicate that metabolically active archaeal communities were dominated by known clades of anaerobic methane oxidizers (archaeal anaerobic methanotroph (ANME)-1, -2 and -3), including a novel 'ANME-2c Sonora' lineage. ANME-2c were found to be dominant, metabolically active and physically associated with syntrophic Bacteria in sulfate-rich shallow sediment layers. In contrast, ANME-1 were more prevalent in the deepest sediment samples and presented a versatile behavior in terms of syntrophic association, depending on the sulfate concentration. ANME-3 were concentrated in small aggregates without bacterial partners in a restricted sediment horizon below the first centimetres. These niche specificities and syntrophic behaviors, depending on biological surface assemblages and environmental availability of electron donors, acceptors and carbon substrates, suggest that ANME could support alternative metabolic pathways than syntrophic anaerobic oxidation of methane.

A giant pockmark colonised by dense cold-seep assemblages near 3160 m depth along the Congo-Angola margin has been surveyed by the ROV Victor 6000. The quantitative distribution of chemosynthetic communities was mapped along the dive tracks from a video study using GIS and image mosaicking. Several types of faunal assemblages, either dominated by bivalves of the families Mytilidae (Bathymodiolus sp.) or Vesicomyidae (Calyptogena sp., 'Vesicomya' aff. chuni), or by Siboglinidae polychaetes (Escarpia southwardae) were mapped over the 800-m diameter pockmark area and sampled for fauna, water and sediment. The isotopic analyses (d 13 C) of tissues from symbiont-bearing species were within the range typical of nutrition via symbiosis using methane for mussels and sulphide for vesicomyids and siboglinids. The living chemosynthetic communities were distributed on a SW-NE axis, corresponding to the expression at the sediment surface of a main buried channel providing fluids to the pockmark. The site was characterised by a more active central part in a depression with abundant carbonate concretions where high-density clusters of siboglinids and mytilids dominate. Large fields of dead and live vesicomyids with a lower mean density were observed in the external areas. The mean coverage of each of the three symbiotic taxa in these two contrasted areas was estimated from mosaic analysis and was up to 30% in the central area dominated by E. southwardae bushes (23%). Symbiont-bearing species distribution was consistent with methane concentrations in seawater that were generally higher in mytilid beds than in the vicinity of siboglinids and vesicomyids. A Principal Component Analysis performed on environmental factors at the ten sampling sites revealed that 37% of the observed variance in the distribution of symbiont-bearing species may be explained by variation in both methane and oxygen concentrations, while a Canonical Redundancy Analysis selected methane concentration as the only variable which explains symbiont-bearing species densities. This spatial distribution of chemosynthetic species at the pockmark scale may reflect temporal patterns of succession of both substrate and fauna, and may be related to different individual pockmarks visible on the microbathymetry mapped using ROV data.