Anaerobic microbial hydrocarbon degradation is a major biogeochemical process at marine seeps. Here we studied the response of the microbial community to petroleum seepage simulated for 190 days in a sediment core from the Caspian Sea using a sediment-oil-flow-through (SOFT) system. Untreated (without simulated petroleum seepage) and SOFT sediment microbial communities shared 43% bacterial genus-level 16S rRNA-based operational taxonomic units (OTU0.945) but shared only 23% archaeal OTU0.945. The community differed significantly between sediment layers. The detection of fourfold higher deltaproteobacterial cell numbers in SOFT than in untreated sediment at depths characterized by highest sulfate reduction rates and strongest decrease of gaseous and mid-chain alkane concentrations indicated a specific response of hydrocarbon-degrading Deltaproteobacteria. Based on an increase in specific CARD-FISH cell numbers, we suggest the following groups of sulfate-reducing bacteria to be likely responsible for the observed decrease in aliphatic and aromatic hydrocarbon concentration in SOFT sediments: clade SCA1 for propane and butane degradation, clade LCA2 for mid- to long-chain alkane degradation, clade Cyhx for cycloalkanes, pentane and hexane degradation, and relatives of Desulfobacula for toluene degradation. Highest numbers of archaea of the genus Methanosarcina were found in the methanogenic zone of the SOFT core where we detected preferential degradation of long-chain hydrocarbons. Sequencing of masD, a marker gene for alkane degradation encoding (1-methylalkyl)succinate synthase, revealed a low diversity in SOFT sediment with two abundant species-level MasD OTU0.96.

The microbial community response to petroleum seepage was investigated in a whole round sediment core (16 cm length) collected nearby natural hydrocarbon seepage structures in the Caspian Sea, using a newly developed Sediment-Oil-Flow-Through (SOFT) system. Distinct redox zones established and migrated vertically in the core during the 190 days-long simulated petroleum seepage. Methanogenic petroleum degradation was indicated by an increase in methane concentration from 8 μM in an untreated core compared to 2300 μM in the lower sulfate-free zone of the SOFT core at the end of the experiment, accompanied by a respective decrease in the δ13C signal of methane from -33.7 to -49.5‰. The involvement of methanogens in petroleum degradation was further confirmed by methane production in enrichment cultures from SOFT sediment after the addition of hexadecane, methylnapthalene, toluene, and ethylbenzene. Petroleum degradation coupled to sulfate reduction was indicated by the increase of integrated sulfate reduction rates from 2.8 SO42-m-2 day-1 in untreated cores to 5.7 mmol SO42-m-2 day-1 in the SOFT core at the end of the experiment, accompanied by a respective accumulation of sulfide from 30 to 447 μM. Volatile hydrocarbons (C2–C6 n-alkanes) passed through the methanogenic zone mostly unchanged and were depleted within the sulfate-reducing zone. The amount of heavier n-alkanes (C10–C38) decreased step-wise toward the top of the sediment core and a preferential degradation of shorter (C30) was seen during the seepage. This study illustrates, to the best of our knowledge, for the first time the development of methanogenic petroleum degradation and the succession of benthic microbial processes during petroleum passage in a whole round sediment core.