Methanotrophs play an important role in controlling methane balance in the biosphere. For over 100 years, methanotrophs of the Proteobacteria phylum have been studied under oxic conditions, with molecular oxygen serving as the major electron acceptor. However, recent metagenomic data suggest that proteobacterial methanotrophs may be more metabolically versatile. In this work, we demonstrate methane-dependent ferrihydrite reduction by a methanotroph of the Methylomonas genus (Gammaproteobacteria) as well as by a methanotroph of the Methylosinus genus (Alphaproteobacteria). This suggests that solid minerals may serve as alternative electron acceptors for aerobic methanotrophs, under hypoxia. This novel type of energy metabolism likely serves as a survival strategy in the absence of molecular oxygen. Our findings expand the range of known microorganisms capable of methane-dependent mineral reduction and provide a deeper understanding of microbial methane metabolism across redox niches.

Methane-dependent reduction of ferrihydrite by Methylomonas cells. (A) Dynamics of accumulation of Fe(II) in experimental mixtures, along negative controls. (B) Change in color of ferrihydrite particles after 20-day incubation under hypoxic conditions indicating transformation of Fe(III) into Fe(II) in the presence of methane (right) but not in control (left). (C) High-resolution transmission electron microscopy, (D) selected area electron diffraction, and (E) lattice images of ferrihydrite particles after reduction by Methylomonas cells at day 30. (F) Dynamics of Fe(III), Fe(II), and CO₂ concentrations over the course of the experiment. Two biological replicates and three technical replicates were used. (G) Dynamics of accumulation of Fe(II) in experiments involving AQDS and corresponding negative controls. Error bars represent standard deviations of values measured in three biological replicates.