Identification of Methanogens and Geochemical Controls on the Production of Methane in Cape Lookout Bight, NC

Methane, the most abundant hydrocarbon in Earth’s atmosphere, is produced in large quantities in marine sediments. Very little is known about the actual microbes that are responsible for the actual production of methane in these systems. We endeavored to identify the organisms that are responsible for the methane produced in a coastal marine site, and to determine whether that production is under thermodynamic control based on hydrogen concentrations. We demonstrated with a bottle incubation of methane seep sediment taken from Cape Lookout Bight, NC, that hydrogen is the controlling substrate in methanogenic sediments. The bottles acted as temporal analog for depth and while sulfate was present, the hydrogen concentration was maintained at below 2 nM. Only after the depletion of sulfate allowed hydrogen concentrations to rise above 5 nM was methane produced. Quantitative PCR data suggest that ANME-2, other Methanosarcinales, and Methanomicrobiales increase when sulfate is depleted. 16s rRNA gene analysis supports the increase of ANME’s and other methanogens relative to other organisms after sulfate concentrations have declined while sulfate reducing bacteria maintain similar population levels throughout the duration of the experiment. 16s rRNA gene analysis also illuminated a relatively uncharacterized euryarchaeota order, Kazan 3A-21, that trended up in relative abundance alongside expected methanogens and in similar abundance suggesting that it is also a methanogen. Total cell counts demonstrate a decline in cells with the decrease of sulfate until a recovery corresponding with production of methane. Our results suggest that hydrogen concentrations strongly influence what metabolic processes can occur in marine sediments, as well as identify the potential diversity of methanogens at this methane seep.