Thermogenic hydrocarbons sustain diverse subseafloor microbial communities in deep sea cold seep sediments

At marine cold seeps, gaseous and liquid hydrocarbons migrate from deep subsurface origins to the sediment-water interface. Cold seep sediments are known to host taxonomically diverse microorganisms, but little is known about their metabolic potential and depth distribution. In this work, we combined geochemical, metagenomic and metabolomic measurements to profile activities of benthic microbial communities spanning different redox regimes within the uppermost 350 cm of Scotian Basin cold seep sediment in the deep sea (2.3 km water depth). Depth profiling revealed compositional and functional differentiation of microbial communities between near-surface sediments (dominated by Proteobacteria) and deeper subsurface zones (dominated by Atribacteria, Chloroflexi, Euryarchaeota and Lokiarchaeota). Metabolic capabilities of community members were inferred from 376 metagenome-assembled genomes spanning 46 phyla. Evaluation of replication rates suggested that several lineages have high in situ microbial activities at different sediment depths. Various subsurface community members are capable of oxidizing short-chain alkanes (alkyl-CoM reductase pathway), longer-chain alkanes (fumarate addition pathway), and aromatic hydrocarbons (fumarate addition and subsequent benzoyl-CoA pathways), including novel lineages within Methanosarcinales and Chloroflexi. Geochemical profiling demonstrated that hydrocarbon substrates are abundant in this location, thermogenic in origin, and subject to biodegradation. The detection of alkyl-/arylalkylsuccinate metabolites, together with carbon isotopic signatures of ethane, propane and carbon dioxide, support that microorganisms are actively degrading hydrocarbons in these sediments. Capacities for reductive dehalogenation, sulfide oxidation, hydrogen oxidation, carbon fixation, and fermentation are also widespread. Most community members may indirectly benefit from thermogenic hydrocarbons through interspecies transfer of electrons and metabolites, and degradation of necromass. Thus, we conclude that upward migrated thermogenic hydrocarbons are important carbon and energy sources that sustain diverse subseafloor microbial communities at permanently cold hydrocarbon seeps.