Methanogen and bacterial diversity and distribution in deep gas hydrate sediments from the Cascadia Margin as revealed by 16S rRNA molecular analysis

The microbial community of a deep (to 234 m below the sea floor) sediment gas hydrate deposit (Cascadia Margin Ocean Drilling Program Site 889/890, Leg 146) was analysed for the first time by molecular genetic techniques. Both bacterial and methanogen diversity were determined by phylogenetic analysis of ribosomal DNA sequences. High molecular mass DNA, indicative of active bacteria, was present in all of the samples. Ribosomal RNA genes were amplified from extracted DNA extracted from sediment using bacteria, and methanogen specific PCR primers, the latter designed in this study. Phylogenetic analysis of approximately 400 bacterial clones demonstrated that 96% were members of the Proteobacteria. These clones were affiliated with the alpha, beta and gamma subdivisions, with Caulobacter (Zymomonas group), Ralstonia and Pseudomonas phylotypes predominating. The methanogen clones were of low diversity and clustered in three sub-groups. Two of these sub-groups (contained 96% of the 400 clones) were closely related to Methanosarcina mazeii, while the third sub-group clustered in the Methanobacteriales. This analysis of a deep sediment gas hydrate environment shows a bacteria and methanogen community of limited diversity and confirms that the gas hydrate zone is biogeochemically active. These results are consistent with the presence of bacterial populations capable of methanogenesis throughout the core, and suggest that the methane hydrate at this site is at least partially biogenic in origin.