Extensive carbon isotopic heterogeneity among methane seep microbiota.

To assess and study the heterogeneity of delta(13)C values for seep microorganisms of the Eel River Basin, we studied two principally different sample sets: sediments from push cores and artificial surfaces colonized over a 14 month in situ incubation. In a single sediment core, the delta(13)C compositions of methane seep-associated microorganisms were measured and the relative activity of several metabolisms was determined using radiotracers. We observed a large range of archaeal delta(13)C values (> 50 per thousand) in this microbial community. The delta(13)C of ANME-1 rods ranged from -24 per thousand to -87 per thousand. The delta(13)C of ANME-2 sarcina ranged from -18 per thousand to -75 per thousand. Initial measurements of shell aggregates were as heavy as -19.5 per thousand with none observed to be lighter than -57 per thousand. Subsequent measurements on shell aggregates trended lighter reaching values as (13)C-depleted as -73 per thousand. The observed isotopic trends found for mixed aggregates were similar to those found for shell aggregates in that the initial measurements were often enriched and the subsequent analyses were more (13)C-depleted (with values as light as -56 per thousand). The isotopic heterogeneity and trends observed within taxonomic groups suggest that ANME-1 and ANME-2 sarcina are capable of both methanogenesis and methanotrophy. In situ microbial growth was investigated by incubating a series of slides and silicon (Si) wafers for 14 months in seep sediment. The experiment showed ubiquitous growth of bacterial filaments (mean delta(13)C = -38 +/- 3 per thousand), suggesting that this bacterial morphotype was capable of rapid colonization and growth.

Methyl sulfides as intermediates in the anaerobic oxidation of methane.

While it is clear that microbial consortia containing Archaea and sulfate-reducing bacteria (SRB) can mediate the anaerobic oxidation of methane (AOM), the interplay between these microorganisms remains unknown. The leading explanation of the AOM metabolism is 'reverse methanogenesis' by which a methanogenesis substrate is produced and transferred between species. Conceptually, the reversal of methanogenesis requires low H(2) concentrations for energetic favourability. We used (13)C-labelled CH(4) as a tracer to test the effects of elevated H(2) pressures on incubations of active AOM sediments from both the Eel River basin and Hydrate Ridge. In the presence of H(2), we observed a minimal reduction in the rate of CH(4) oxidation, and conclude H(2) does not play an interspecies role in AOM. Based on these results, as well as previous work, we propose a new model for substrate transfer in AOM. In this model, methyl sulfides produced by the Archaea from both CH(4) oxidation and CO(2) reduction are transferred to the SRB. Metabolically, CH(4) oxidation provides electrons for the energy-yielding reduction of CO(2) to a methyl group ('methylogenesis'). Methylogenesis is a dominantly reductive pathway utilizing most methanogenesis enzymes in their forward direction. Incubations of seep sediments demonstrate, as would be expected from this model, that methanethiol inhibits AOM and that CO can be substituted for CH(4) as the electron donor for methylogenesis.

Methyl sulfide production by a novel carbon monoxide metabolism in Methanosarcina acetivorans.

We observed dimethyl sulfide and methanthiol production in pure incubations of the methanogen Methanosarcina acetivorans when carbon monoxide (CO) served as the only electron donor. Energy conservation likely uses sodium ion gradients for ATP synthesis. This novel metabolism permits utilization of CO by the methanogen, resulting in quantitative sulfide methylation.