Metatranscriptomic Evidence for Magnetite Nanoparticle-Stimulated Acetoclastic Methanogenesis under Continuous Agitation

Previous studies have shown that conductive nanoparticles, such as MNPs, accelerate methanogenesis and suggested that MNPs facilitate DIET between exoelectrogenic bacteria and methanogenic archaea. In these methanogens, electrons thus obtained are considered to be used for hydrogenotrophic methanogenesis. However, the present work provides evidence that shows that MNPs accelerate DIET-independent acetoclastic methanogenesis under continuous agitation. Since most of previous studies have examined effects of MNPs in static or weakly agitated methanogenic cultures, results obtained in the present work suggest that hydraulic conditions definitively determine how MNPs accelerate methanogenesis. In addition, the knowledge obtained in this study is useful for engineers operating stirred-tank anaerobic digesters, since we show that MNPs accelerate methanogenesis under continuous agitation. Conductive nanomaterials have been reported to accelerate methanogenesis by promoting direct interspecies electron transfer (DIET), while their effects seem to vary depending on operational conditions. The present study examined the effects of magnetite nanoparticles (MNPs) on methanogenesis from acetate by soil-derived anaerobic cultures under continuous agitation. We found that MNPs accelerated methanogenesis in agitated cultures, as has been observed previously for static cultures. Metabarcoding of 16S rRNA gene amplicons showed that Methanosarcina substantially increased in the presence of MNPs, while DIET-related Geobacter did not occur. Metagenomic and metatranscriptomic analyses confirmed the predominance of Methanosarcina in MNP-supplemented agitated cultures. In addition, genes coding for acetoclastic methanogenesis, but not those for hydrogenotrophic methanogenesis, were abundantly expressed in the dominant Methanosarcina in the presence of MNPs. These results suggest that MNPs stimulate acetoclastic methanogenesis under continuous agitation. IMPORTANCE Previous studies have shown that conductive nanoparticles, such as MNPs, accelerate methanogenesis and suggested that MNPs facilitate DIET between exoelectrogenic bacteria and methanogenic archaea. In these methanogens, electrons thus obtained are considered to be used for hydrogenotrophic methanogenesis. However, the present work provides evidence that shows that MNPs accelerate DIET-independent acetoclastic methanogenesis under continuous agitation. Since most of previous studies have examined effects of MNPs in stati