Methane-Fueled Syntrophy through Extracellular Electron Transfer: Uncovering the Genomic Traits Conserved within Diverse Bacterial Partners of Anaerobic Methanotrophic Archaea

Abstract

The anaerobic oxidation of methane by anaerobic methanotrophic (ANME) archaea in syntrophic partnership with deltaproteobacterial sulfate-reducing bacteria (SRB) is the primary mechanism for methane removal in ocean sediments. The mechanism of their syntrophy has been the subject of much research as traditional intermediate compounds, such as hydrogen and formate, failed to decouple the partners. Recent findings have indicated the potential for extracellular electron transfer from ANME archaea to SRB, though it is unclear how extracellular electrons are integrated into the metabolism of the SRB partner. We used metagenomics to reconstruct eight genomes from the globally distributed SEEP-SRB1 clade of ANME partner bacteria to determine what genomic features are required for syntrophy. The SEEP-SRB1 genomes contain large multiheme cytochromes that were not found in previously described free-living SRB and also lack periplasmic hydrogenases that may prevent an independent lifestyle without an extracellular source of electrons from ANME archaea. Metaproteomics revealed the expression of these cytochromes at in situ methane seep sediments from three sites along the Pacific coast of the United States. Phylogenetic analysis showed that these cytochromes appear to have been horizontally transferred from metal-respiring members of the Deltaproteobacteria such as Geobacter and may allow these syntrophic SRB to accept extracellular electrons in place of other chemical/organic electron donors.

Gene expression and ultrastructure of meso‐ and thermophilic methanotrophic consortia

“…As previously reported for HotSeep‐1 and Seep‐SRB1 (Krukenberg et al ., ; Skennerton et al ., ), both Seep‐SRB2 draft genomes encode the core gene set for dissimilatory sulfate reduction, including membrane‐bound electron transport complexes (Fig. ), but lack genes related to methane metabolism (see Supporting Information Tables S6 and S7).…”

“…The draft genomes of the two types of Seep‐SRB2 bacteria associated with the mesophilic ANME‐2c and ‐1a had a size of 3.6 Mb (E20) and 2.6 Mb (G37), and an estimated completeness of 91%–94% and 91%–95%, respectively (see Table ). This is in the range of what has been reported for HotSeep‐1 (2.5 Mb, 97% completeness; Krukenberg et al ., ) and Seep‐SRB1a (∼ 3 Mb; Skennerton et al ., ). Congruent with our observations from the ANME draft genomes, the GC content of the partner bacteria draft genomes did not show temperature‐dependent variations (47% in E20 Seep‐SRB2, 49% in G37 Seep‐SRB2 and 37% in G60 HotSeep‐1).…”

“…Recent studies provided insights into the draft genome of the AOM partner bacteria HotSeep‐1 (Wegener et al ., ; Krukenberg et al ., ) and Seep‐SRB1 (Skennerton et al ., ). Here we analyzed the draft genome and transcriptome of two Seep‐SRB2 types, representing environmentally relevant partner bacteria from a clade described by Kleindienst and colleagues ().…”

“…This complex may also work in reverse to catalyze the reverse electron transfer from NADH to ferredoxin driven by a proton gradient (Müller et al ., ). Like the Seep‐SRB1 draft genome (Skennerton et al ., ), both Seep‐SRB2 encode the cytoplasmic Flx/Hdr complex (Ramos et al ., ), which is not encoded in the HotSeep‐1 draft genome. This complex couples the reduction of ferredoxin by NADH to the reduction of a disulfide bond, possibly in the DsrC protein, via electron bifurcation.…”

“…Like the ANME genomes, the genomes of the three partner bacteria contain the genes encoding autotrophic carbon fixation. The Seep‐SRB2 genomes encode the reductive acetyl‐CoA pathway, a common feature among autotrophic sulfate reducers including Seep‐SRB1 (Skennerton et al ., ). Notably, in the E20 and G37 enrichment, the expression of genes encoding the reductive acetyl‐CoA pathway in ANME and their partner bacteria was very similar, documenting the coupled growth patterns of the two consortial members.…”