Lithosyntrophy: Obligate syntrophy in a phosphite-oxidizing, methanogenic culture

The anaerobic conversion of organic matter to methane and carbon dioxide typically relies on obligate syntrophic interactions between bacteria and methanogenic archaea, where interspecies hydrogen (H2) transfer enables thermodynamically constrained reactions to proceed near equilibrium. Syntrophs couple the oxidation of fermentation products such as fatty acids and alcohols to the reduction of protons to form H2. These reactions can only proceed if low H2 concentrations are maintained by H2-consuming syntrophic partners. Here, we describe "lithosyntrophy," a novel mode of syntrophic interaction in which electrons that drive hydrogenotrophic methanogenesis originate from an inorganic compound rather than from the canonical organic substrates. Candidatus Phosphitivorax anaerolimi strain Phox-21 oxidizes phosphite (HPO32-, oxidation state +3) to phosphate coupled to hydrogenogenesis in an obligate energetic dependency on a hydrogenotrophic methanogen, Methanoculleus sp. Physiology experiments, thermodynamic calculations, genomic annotation, and metaproteomics analysis collectively revealed a mechanism for syntrophic phosphite oxidation in Phox-21, which requires phosphite, acetate, and CO2 as co-substrates. In this pathway, electrons derived from phosphite drive H2 production via an electron-confurcating hydrogenase. Unlike previously characterized acetogenic phosphite oxidizers that grow without exogenous acetate, Phox-21 requires acetate to regenerate AMP, a cofactor required by the phosphite dehydrogenase, PtdF. Lithosyntrophic phosphite oxidizers may play important roles both in transferring reducing equivalents as well as biologically available phosphorus to other members of their surrounding microbial communities. We infer that lithosyntrophic DPO emerged before acetoclastic methanogenesis and was a major sink for acetate in the Archaean when phosphite was more abundant. Significance statementDissimilatory phosphite-oxidizing microorganisms (DPOM) use phosphite as an energy source, producing phosphate. While the two previously isolated DPOM couple phosphite oxidation to carbon fixation via the Wood-Ljungdahl pathway, Candidatus Phosphitivorax anaerolimi strain Phox-21 instead performs lithosyntrophic metabolism, coupling DPO to proton reduction in obligate partnership with a hydrogenotrophic methanogen. This establishes a novel link between phosphorus and carbon redox cycles: like organosyntrophy, DPO can fuel methanogenesis. Lithosyntrophy expands our understanding of syntrophic interactions and suggests similar processes may occur with other inorganic substrates in anoxic ecosystems. We propose lithosyntrophic DPO predates acetoclastic methanogenesis, serving as an early acetate sink during the Archaean. Following Earths oxygenation, phosphite depletion likely reduced competition for acetate, enabling the later evolution of acetoclastic methanogenesis.