We report here a molecular survey based on 16S rRNA genes of the bacterial diversity found in two deep-sea vent niches at the Mid-Atlantic Ridge: hydrothermal sediment (Rainbow site), and microcolonizers made of three different substrates (organic-rich, iron-rich and pumice) that were exposed for 15 days to a vent emission. Bacterial diversity in sediment samples was scattered through many bacterial divisions. The most abundant and diverse environmental sequences (phylotypes) in our libraries corresponded to the Gammaproteobacteria, followed by the Acidobacteria. We detected members of all the subdivisions within the Proteobacteria. Myxobacterial lineages were the most represented within the delta subdivision. Phylotypes ascribing to the Cytophaga-Flavobacterium-Bacteroides, Planctomycetales, high and low G + C Gram-positives, Nitrospirae, and the candidate division TM7 were also identified. Compared to this broad taxonomic coverage, microcolonizers were almost exclusively colonized by epsilonproteobacteria, although these exhibited considerable morphological and phylogenetic in-group diversity. No specificity for any of the substrates tested was seen. This observation further supports the idea of the ecological dominance of epsilonproteobacteria in the fluid–seawater interface environment. Because oxidation of reduced S species and/or sulphur-reduction is thought to be essential for their energetic metabolism in these areas, we mapped different oxidation states of S in individual bacterial filaments from the iron-rich microcolonizer. For this, we used high-resolution, non-destructive synchrotron micro-X-ray Absorption Near-Edge Spectroscopy (micro-XANES), which revealed the co-existence of different S oxidation states, from sulphide to sulphate, at the level of individual cells. This suggests that these cells were metabolizing sulphur in situ.