Marine cold seep ANME-2/SRB consortia produce their lipid biomass from inorganic carbon
In cold seeps, anaerobic methanotrophic archaea (ANME) and sulfate-reducing bacteria (SRB) oxidize methane to inorganic carbon (IC) coupled to sulfate reduction. While catabolic pathways are well resolved, carbon flow into biomass remains unclear. We conducted lipid stable isotope probing (lipid-SIP) experiments with Astoria Canyon sediments dominated by ANME-2/SRB consortia and incubated samples with either 13C-labeled methane (13CH4) or dissolved IC (DI13C). Lipid-specific {delta}13C analysis showed higher 13C incorporation from DI13C than from 13CH4. After 30 days, {delta}13C values were up to +417{per thousand} in SRB-specific fatty acids (e.g., C16:1{omega}5c, cyC17:0{omega}5,6) and +126{per thousand} in ANME-2-specific isoprenoid lipids (e.g., archaeol, crocetane). Based on these values, we calculated carbon assimilation rates and found that both partners primarily assimilate IC. Remarkedly, IC assimilation in SRB lipids was eight times higher than in ANME lipids, suggesting that ANME either rely on fixation of internally generated CO2 from 13C-label-free methane before it can be exchanged with the environment or they utilize an additional, yet unknown carbon source for lipid biosynthesis. By examining the step-wise 13C-enrichment of ANME- and SRB-derived lipids, we further delineate biosynthetic pathways for archaeal and bacterial diether lipid formation and highlight crocetane as a bilayer-modulating isoprenoid hydrocarbon potentially affecting membrane fluidity and proton permeability. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=88 SRC="FIGDIR/small/667857v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@a70354org.highwire.dtl.DTLVardef@cb95acorg.highwire.dtl.DTLVardef@1cf6d5corg.highwire.dtl.DTLVardef@114b912_HPS_FORMAT_FIGEXP M_FIG C_FIG Lipid-SIP experiments in Astoria Canyon sediments revealed that both ANME-2 and SRB primarily assimilate inorganic carbon (IC), not methane, into biomass. SRB-specific lipids showed eightfold higher I13C-assimilation than ANME lipids, suggesting SRB directly assimilate environmental IC, while ANME rely on internally produced CO2 or alternative carbon sources, potential provided by SRBs.