This expedition represents the first time that subsurface microbial samples have been collected from a hydrothermal system using modern microbially-motivated ocean drilling techniques with the aim of obtaining direct evidence for a functionally active, metabolically diverse subvent biosphere in relation to the hydrothermally active mounds located in the Okinawa Trough. This proposal seeks to expand on our post-cruise results with the focus directed towards a metagenomic community analysis using two different avenues of next-gen sequencing. We are motivated by the recent discovery of unique and possibly endemic populations of subsurface Zetaproteobacteria phylotypes. We are also motivated by our initial cultivation results of multiple enrichments that were obtained using both microaerophilic and anaerobic FeOB culturing conditions and that we were able to detect up to 13% Zetas, by qPCR analysis, in two habitats with very different fluid flow regimes. We propose to use pyrosequencing of 16S genes to assay community diversity and massively-parallel sequencing to reconstruct genomes of the most abundant community members, focusing on these two habitat extremes. We hypothesize that this approach will allow insights into the physiology of these FeOB communities thereby demonstrating key features that they use to survive, compete and grow in the deep “tepid” biosphere.