Molecular and isotopic analysis of anaerobic methane-oxidizing communities in marine sediments

Convergent lines of molecular, carbon-isotopic, and phylogenetic evidence have previously indicated (Hinrichs, K.-U., Hayes, J.M., Sylva, S.P., Brewer, P.G., DeLong, E. F., 1999. Methane-consuming archaebacteria in marine sediments. Nature 398, 802–805.) that archaea are involved in the anaerobic oxidation of methane in sediments from the Eel River Basin, offshore northern California. Now, further studies of those same sediments and of sediments from a methane seep in the Santa Barbara Basin have confirmed and extended those results. Mass spectrometric and chromatographic analyses of an authentic standard of sn-2-hydroxyarchaeol (hydroxylated at C-3 in the sn-2 phytanyl moiety) have confirmed our previous, tentative identification of this compound but shown that the previously examined product was the mono-TMS, rather than di-TMS, derivative. Further analyses of ^(13)C-depleted lipids, appreciably more abundant in samples from the Santa Barbara Basin, have shown that the archaeal lipids are accompanied by two sets of products that are only slightly less depleted in ^(13)C. These are additional glycerol ethers and fatty acids. The alkyl substituents in the ethers (mostly monoethers, with some diethers) are non-isoprenoidal. The carbon-number distributions and isotopic compositions of the alkyl substituents and of the fatty acids are similar, suggesting strongly that they are produced by the same organisms. Their structures, n-alkyl and methyl-branched n-alkyl, require a bacterial rather than archaeal source. The non-isoprenoidal glycerol ethers are novel constituents in marine sediments but have been previously reported in thermophilic, sulfate- and nitrate-reducing organisms which lie near the base of the rRNA-based phylogenetic tree. Based on previous observations that the anaerobic oxidation of methane involves a net transfer of electrons from methane to sulfate, it appears likely that the non-archaeal, ^(13)C-depleted lipids are products of one or more previously unknown sulfate-reducing bacteria which grow syntrophically with the methane-utilizing archaea. Their products account for 50% of the fatty acids in the sample from the Santa Barbara Basin. At all methane-seep sites examined, the preservation of aquatic products is apparently enhanced because the methane-oxidizing consortium utilizes much of the sulfate that would otherwise be available for remineralization of materials from the water column