Insight into the mechanism of chemoautotrophic denitrification using pyrite (FeS₂) as electron donor

In this study, denitrification was performed using pyrite as sole electron donor. The nitrate reducing rate ranged from 0.61 to 0.95 mM/d. The production of nitrous oxide (N₂O) was observed, accounting for 20% of the total nitrate reduction. The isotope fractionation indicated that N₂O production was mainly caused by the bacterial denitrification, instead of chemical denitrification by Fe(Ⅱ). Thiobacillus was the predominant genus, of which relative abundance decreased after the incubation with pyrite. Conversely, other genera belonging to Actinobacteria, like Rhodococcus, increased by more than 10 times. These Actinobacteria-like bacteria lack nitrous oxide reductase, which might be the reason for high N₂O production. Furthermore, the predicted microbial functions analysis by PICRUSt2 showed that the genes (menC/E/G) involved in the biosynthesis of electron shuttles (menaquinone-related redox-active molecule), which were remarkably enriched during the process, suggesting that the first step of pyrite oxidation might be driven by the microbial derived electron shuttles.