Molecular indicators of methane metabolisms at cold seeps along the United States Atlantic Margin

Abstract

Anaerobic oxidation of methane (AOM) and the environmental conditions supporting AOM on continental margins is an essential component to global methane budgets. Diagnostic lipid biomarkers and their compound specific isotope analysis preserved in authigenic carbonates at cold seeps can serve as “fingerprints” to archaeal−bacterial consortia involved in AOM. However, despite the discovery of several hundreds of seeps along the United States Atlantic Margin (USAM), there are relatively few biomarker investigations of cold seep carbonates along this passive margin. A lipid biomarker, carbon isotope, and DNA marker gene study was therefore undertaken to determine the microbial origins of authigenic carbonates from two USAM seeps, Norfolk and the Baltimore Canyon seep fields. Results from this study capture a distinct archaeal lipid signature from putative methanotrophic archaea, including archaeol (I), sn-2-hydroxyarchaeol, 2,6,10,15,19-pentamethylicosane (PMI), and crocetane. The 13C-depleted AOM-related archaeal lipid samples (i.e., archaeol: −91.6‰, sn-2-hydroxyarchaeol: −129.2‰, PMI −92.8‰, and crocetane: −70.9‰) confirm the dominance of methane assimilation and isotope fractionation during AOM. These results are consistent with the detection of archaeal anaerobic methanotrophs (ANMEs) based on 16S rRNA gene sequencing. The Norfolk authigenic carbonate contained ANME-1a, -1b, 2a-2b, and 2c whereas only the ANME-2 clade was detected at Baltimore and present as the subclusters 2a-2b and -2b. The ANME-2d clade may also be present, particularly at the Baltimore seep site, given the high abundance of Candidatus Methanoperedens nitroreducens detected in the mcrA gene sequencing. The presence of terminally branched fatty acids, antesio- and iso-C15:0 components, as well as C16:1ω7 with δ13C values as low as −107.6‰, are indicative of sulfate-reducing bacteria (SRB) at the Norfolk seep site and supports syntrophy of SRB with methane-oxidizing archaea. In contrast, nitrate-driven AOM in syntrophy M. nitroreducens at the Baltimore seep site is consistent with elevated fatty acid δ13C values and lack of Deltaproteobacteria at the Baltimore seep site. Taken together, the range in lipid composition, distribution, and carbon isotopic composition observed at the Norfolk and Baltimore seep sites suggests AOM is performed by multiple archaea instead of a single species and may be paired with either or both nitrate- and sulfate-reduction. Given the heterogeneous nature of cold seep ecosystems, this study fills a critical spatial gap in our knowledge of AOM activity at two seep sites along a passive margin.