Autotrophy as a predominant mode of carbon fixation in anaerobic methane-oxidizing microbial communities

Abstract

The methane-rich, hydrothermally heated sediments of the Guaymas Basin are inhabited by thermophilic microorganisms, including anaerobic methane-oxidizing archaea (mainly ANME-1) and sulfatereducing bacteria (e.g., HotSeep-1 cluster). We studied the microbial carbon flow in ANME-1/ HotSeep-1 enrichments in stable-isotopeprobing experiments with and without methane. The relative incorporation of 13 C from either dissolved inorganic carbon or methane into lipids revealed that methane-oxidizing archaea assimilated primarily inorganic carbon. This assimilation is strongly accelerated in the presence of methane. Experiments with simultaneous amendments of both 13 C-labeled dissolved inorganic carbon and deuterated water provided further insights into production rates of individual lipids derived from members of the methane-oxidizing community as well as their carbon sources used for lipid biosynthesis. In the presence of methane, all prominent lipids carried a dual isotopic signal indicative of their origin from primarily autotrophic microbes. In the absence of methane, archaeal lipid production ceased and bacterial lipid production dropped by 90%; the lipids produced by the residual fraction of the metabolically active bacterial community predominantly carried a heterotrophic signal. Collectively our results strongly suggest that the studied ANME-1 archaea oxidize methane but assimilate inorganic carbon and should thus be classified as methane-oxidizing chemoorganoautotrophs.methanotrophy | biomarker | acetyl-CoA pathway | syntrophy M ethane is an important greenhouse gas and the most abundant hydrocarbon in marine sediments. Its upward flux to the sediment-water interface is strongly reduced by sulfate-dependent anaerobic oxidation of methane (AOM) (1, 2). AOM is performed by syntrophic associations of anaerobic methane-oxidizing archaea (ANMEs) (3, 4) and their sulfate-reducing bacterial partners (SRBs) (mainly relatives of Desulfosarcina or Desulfobulbus) (5-8). The free energy yield of the AOM net reaction is one of the lowest known for catabolic reactions under environmental conditions (ΔG ranges from -20 to -40 kJ·mol -1 ; e.g., refs. 9, 10). Consequently, activity and biomass doubling times determined under optimized laboratory conditions range from 2 to 5 mo (11) and growth yields are extremely low, around 1% relative to oxidized methane (11,12). The biomass of ANMEs and SRBs involved in AOM is usually strongly depleted in 13 C. For instance, at methane seep locations, the δ 13 C values of specific bacterial fatty acids and archaeal ether lipids range from -60 to -100‰ and -70 to -130‰, respectively (e.g., refs. 13-17). Such low values have been interpreted as evidence for the incorporation of 13 C-depleted methane into biomass (e.g., refs. 3, 4, 18).One way to identify the carbon sources of microbial biomass is to perform stable-isotope-probing (SIP) experiments (19), followed by analysis of biomolecules such as membrane lipids (lipid-SIP hereafter). Application of lipid-SIP to cold seep sediments and micro...

High rates of anaerobic methane oxidation in freshwater wetlands reduce potential atmospheric methane emissions

“…10-Methylhexadecanoic acid, a characteristic lipid for 'Methylomirabilis oxyfera', a freshwater bacterium capable of oxidizing methane with nitrite 33 , was either not detected at these sites or found in rather low abundance (o1.5% of all fatty acids; data not shown). This proposed biomarker for n-damo 48 was not sufficiently depleted in 13 C to unambiguously support a relationship with AOM (Supplementary Table 3); a conclusive relationship of this biomarker to AOM is complicated by the evidence for chemoorganoautotrophy of M. oxyfera 49 , in analogy to some ANME archaea 47 , and additional sources of this biomarker such as sulphate-reducing 50 and anammox bacteria 51 . The other proposed diagnostic lipid for M. oxyfera, 10-methyl-hexadec-7-enoic acid 48 , was not detected in any sample.…”

“…The isotopic enrichment of archaeal lipids relative to methane is best explained by additional biological sources of these compounds, for example, from heterotrophic archaea 45 or autotropic archaea such as ammonium-oxidizing thaumarcharchaea 46 . Alternatively, the incorporation of DIC rather than methane into methane-oxidizing archaeal lipids 47 could explain the observed 13 C-enriched values. Notably, the archaeal biphytanes are 13 C-depleted relative to CO 2 by À 42 to À 17%, consistent with a predominant origin from autotrophic, methane-oxidizing archaea 47 .…”

“…Alternatively, the incorporation of DIC rather than methane into methane-oxidizing archaeal lipids 47 could explain the observed 13 C-enriched values. Notably, the archaeal biphytanes are 13 C-depleted relative to CO 2 by À 42 to À 17%, consistent with a predominant origin from autotrophic, methane-oxidizing archaea 47 .…”

“…Some have argued the extreme isotopic depletions in archaeal lipids attributed to AOM in vent and seep communities may be entirely due to autotrophic methanogenesis 58 . Recent isotope-based inquiries, however, have demonstrated that microorganisms involved in AOM assimilate carbon dioxide 47,57 in addition to methane 57 . Phylogenetic analyses indicate variable AOM pathways likely occur, possibly within a single group of archaea in response to environmental factors 57 .…”

Unveiling microbial activities along the halocline of Thetis, a deep-sea hypersaline anoxic basin

“…The ANME-1 archaea were assumed to be obligatory methanotrophs, but recent studies provide evidence of co-occurrence of AOM and methanogenesis (Lloyd et al, 2011;Bertram et al, 2013;Wang et al, 2013). ANMEs were shown to assimilate inorganic carbon (Kellermann et al, 2012), but also to be capable of growing on acetate, pyruvate or butyrate using thiosulfate as an electron acceptor (Jagersma et al, 2012). The acetate-metabolizing potential in ANME2a was supported by the identification of the ion-motive electron transport complex Rnf that is independent of H 2 (Wang et al, 2013).…”