Furthermore, simple organic compounds are formed by thermal decomposition of biomass (McCollom & Seewald, 2007), homoacetogenesis (Drake et al., 2008 Lever et al., 2010) or by the vent-associated macrofauna (Pimenov et al., 2002). Organic acids, lipids, and hydrocarbons are formed in the deep subsurface by serpentinization and subsequent Fischer–Tropsch-type processes under elevated temperature and pressure (Shock & Schulte, 1998 Holm & Charlou, 2001). However, diffuse and end-member hydrothermal fluids can also contain various organic compounds other than methane (Holm & Charlou, 2001 Rogers & Amend, 2006 Konn et al., 2009 Charlou et al., 2010 Lang et al., 2010 Reeves et al., 2014). In submarine hydrothermal systems, inorganic carbon is the primary carbon source (Shively et al., 1998 Nakagawa & Takai, 2008). Our data provide first insights into the heterotrophic microbial community, catalyzing an under-investigated part of microbial carbon cycling at hydrothermal vents.Įpsilonproteobacteria, Gammaproteobacteria, heterotrophy, 16S rRNA gene, nanoSIMS, stable isotopes Introduction The rapid growth of microorganisms upon acetate addition suggests that acetate consumers in diffuse fluids are copiotrophic opportunists, which quickly exploit their energy sources, whenever available under the spatially and temporally highly fluctuating conditions. Here, 16S rRNA gene sequences were mostly related to mesophilic Marinobacter, reflecting the high content of seawater in these fluids. In contrast, Gammaproteobacteria dominated the 13C-acetate assimilation in incubations of 37 ☌-fluids from the back-arc hydrothermal system in the Manus Basin/Papua New Guinea. In 55 ☌-fluids from the Menez Gwen hydrothermal system/Mid-Atlantic Ridge, a novel epsilonproteobacterial group accounted for nearly all assimilation of acetate, representing the first aerobic acetate-consuming member of the Nautiliales. We followed cell growth and assimilation of 13C into single cells by nanoSIMS combined with fluorescence in situ hybridization (FISH). To identify potential heterotrophs without prior cultivation, we combined microbial community analysis with short-term incubations using 13C-labeled acetate at two distinct hydrothermal systems. Microorganisms consuming these compounds at hydrothermal sites are so far only known from cultivation-dependent studies. Diffuse hydrothermal fluids often contain organic compounds such as hydrocarbons, lipids, and organic acids.