Petrochemical and geological evidence suggest that petroleum in most reservoirs is anaerobically biodegraded to some extent. However, the conditions for this metabolism and the cultivation of the requisite microorganisms are rarely established. Here, we report on microbial hydrocarbon metabolism in two distinct oilfields on the North Slope of Alaska (designated Fields A and B). Signature anaerobic hydrocarbon metabolites were detected in produced water from the two oilfields offering evidence of in situ biodegradation activity. Rate measurements revealed that sulfate reduction was an important electron accepting process in Field A (6–807 µmol S l−1 day−1), but of lesser consequence in Field B (0.1–10 µmol S l−1 day−1). Correspondingly, enrichments established at 55°C with a variety of hydrocarbon mixtures showed relatively high sulfate consumption but low methane production in Field A incubations, whereas the opposite was true of the Field B enrichments. Repeated transfer of a Field B enrichment showed ongoing methane production in the presence of crude oil that correlated with ≥ 50% depletion of several component hydrocarbons. Molecular-based microbial community analysis of the methanogenic oil-utilizing consortium revealed five bacterial taxa affiliating with the orders Thermotogales, Synergistales, Deferribacterales (two taxa) and Thermoanaerobacterales that have known fermentative or syntrophic capability and one methanogen that is most closely affiliated with uncultured clones in the H2-using family Methanobacteriaceae. The findings demonstrate that oilfield-associated microbial assemblages can metabolize crude oil under the thermophilic and anaerobic conditions prevalent in many petroleum reservoirs.