Polyphosphate metabolism genes are associated with hydroxyapatite biomineralization capacity in a novel hydrogenotrophic methanogen Methanobacterium AMB_03

Methanogenic archaea play crucial roles in sedimentary phosphorus cycling, but their capacity for phosphate biomineralization remains poorly understood. Here we demonstrate that Methanobacterium AMB_03, isolated from a serpentinizing environment, mediates extracellular precipitation of carbonate-substituted hydroxyapatite (HAp) under phosphate-enriched conditions. Comparative genomic analysis of AMB_03 versus non-mineralizing strains (Methanobacterium oryzae, Methanoculleus bourgensis MAB1) shows that AMB_03 encodes polyphosphate kinase 1 (ppk1), enabling polyphosphate synthesis from ATP. In contrast, M. oryzae and MAB1 completely lack ppk1, preventing polyphosphate accumulation. Phylogenomic analysis of 320 Methanobacteriota genomes further shows that complete polyP cycling capacity is present in only ~25% of representatives, placing AMB_03 among a minority of methanogens with this capability. Experimental cultivation across phosphate gradients (0-6 mM) showed HAp early-onset precipitation was preferential at [≥]3 mM. Precipitation was eventually observed at all phosphate concentrations (except P0) by 49 weeks and was associated with extracellular polymeric substance production and localized pH increases of up to 8.08. Raman spectroscopy confirmed carbonate-substituted HAp, with characteristic peaks at 960.5 cm-1 (PO43-) and 1070-1072 cm-1 (CO32-). These findings identify a strong genomic association between polyphosphate metabolism genes and biomineralization capacity, suggesting that methanogens with complete polyP cycling systems may create localized phosphate-rich environments that promote authigenic mineral formation in sedimentary systems.