The anaerobic formation and oxidation of methane involve unique enzymatic mechanisms and cofactors, all of which are believed to be specific for C-compounds. Here we show that an anaerobic thermophilic enrichment culture composed of dense consortia of archaea and bacteria apparently uses partly similar pathways to oxidize the C hydrocarbon butane. The archaea, proposed genus 'Candidatus Syntrophoarchaeum', show the characteristic autofluorescence of methanogens, and contain highly expressed genes encoding enzymes similar to methyl-coenzyme M reductase. We detect butyl-coenzyme M, indicating archaeal butane activation analogous to the first step in anaerobic methane oxidation. In addition, Ca. Syntrophoarchaeum expresses the genes encoding β-oxidation enzymes, carbon monoxide dehydrogenase and reversible C methanogenesis enzymes. This allows for the complete oxidation of butane. Reducing equivalents are seemingly channelled to HotSeep-1, a thermophilic sulfate-reducing partner bacterium known from the anaerobic oxidation of methane. Genes encoding 16S rRNA and methyl-coenzyme M reductase similar to those identifying Ca. Syntrophoarchaeum were repeatedly retrieved from marine subsurface sediments, suggesting that the presented activation mechanism is naturally widespread in the anaerobic oxidation of short-chain hydrocarbons.

The anaerobic oxidation of methane coupled to sulfate reduction is a microbially mediated process requiring a syntrophic partnership between anaerobic methanotrophic (ANME) archaea and sulfate-reducing bacteria (SRB). Based on genome taxonomy, ANME lineages are polyphyletic within the phylum Halobacterota, none of which have been isolated in pure culture. Here, we reconstruct 28 ANME genomes from environmental metagenomes and flow sorted syntrophic consortia. Together with a reanalysis of previously published datasets, these genomes enable a comparative analysis of all marine ANME clades. We review the genomic features that separate ANME from their methanogenic relatives and identify what differentiates ANME clades. Large multiheme cytochromes and bioenergetic complexes predicted to be involved in novel electron bifurcation reactions are well distributed and conserved in the ANME archaea, while significant variations in the anabolic C1 pathways exists between clades. Our analysis raises the possibility that methylotrophic methanogenesis may have evolved from a methanotrophic ancestor.

The methanogenic degradation of oil hydrocarbons can proceed through syntrophic partnerships of hydrocarbon-degrading bacteria and methanogenic archaea [1][2][3] . However, recent culture-independent studies have suggested that the archaeon 'Candidatus Methanoliparum' alone can combine the degradation of long-chain alkanes with methanogenesis 4,5 . Here we cultured Ca. Methanoliparum from a subsurface oil reservoir. Molecular analyses revealed that Ca. Methanoliparum contains and overexpresses genes encoding alkyl-coenzyme M reductases and methyl-coenzyme M reductases, the marker genes for archaeal multicarbon alkane and methane metabolism. Incubation experiments with different substrates and mass spectrometric detection of coenzyme-M-bound intermediates confirm that Ca. Methanoliparum thrives not only on a variety of long-chain alkanes, but also on n-alkylcyclohexanes and n-alkylbenzenes with long n-alkyl (C ≥13 ) moieties. By contrast, short-chain alkanes (such as ethane to octane) or aromatics with short alkyl chains (C ≤12 ) were not consumed. The wide distribution of Ca. Methanoliparum 4-6 in oil-rich environments indicates that this alkylotrophic methanogen may have a crucial role in the transformation of hydrocarbons into methane.In subsurface oil reservoirs and marine oil seep sediments, microorganisms use hydrocarbons as a source of energy and carbon 7,8 . The microorganisms preferentially consume alkanes, cyclic and aromatic compounds, leaving an unresolved complex mixture as residue and thereby altering the quality of the oil 7,8 . In the absence of sulfate, microorganisms couple anaerobic hydrocarbon degradation to methane formation 1,9,10 . This reaction was originally demonstrated by Zengler et al 2 as methanogenic 'microbial alkane cracking', and a large number of studies have shown that it can be performed in syntrophic interactions of bacteria and archaea 11 . In this syntrophy, the bacteria ferment the oil to acetate, carbon dioxide and hydrogen, while hydrogenotrophic and/or acetotrophic methanogenic archaea use the products for methanogenesis 1,2,11 .Diverse anaerobic hydrocarbon activation mechanisms exist, including the well-studied fumarate addition pathway catalysed by glycyl radical enzymes 12 . This mechanism is widespread among bacteria that thrive on alkanes of various chain lengths and other hydrocarbons 12,13 . By contrast, several archaeal lineages activate gaseous alkanes with the help of a specific type of methyl-coenzyme M reductase (MCR), an enzyme that was originally described to catalyse the reduction of methyl-coenzyme M (methyl-CoM) to methane in methanogens 14 . Anaerobic methanotrophic archaea use canonical MCRs to activate methane into methyl-CoM, which is then oxidized to CO 2 . Short-chain alkane-oxidizing archaea contain divergent variants of this enzyme, which are known as alkyl-CoM reductases (ACRs). Analogous to the methane-activating MCR, ACRs activate multicarbon alkanes to form CoM-bound alkyl units [15][16][17] . The cultured alkane-oxidizing archaea oxidize sho...