Reduction of nitrate to nitrite by microbes under oxic conditions

Human activities have greatly increased the input of nitrate to natural and managed ecosystems, but the fate of excess soil nitrate is still unclear. Many studies assume that dissimilatory reduction of nitrate to nitrite is an anaerobic process, but this first step of denitrification can occur in some bacteria at oxygen concentrations that are high enough to repress downstream reduction of nitrite to gaseous products. Here, we examine whether dissimilatory reduction of nitrate under aerobic conditions is an additional, underappreciated fate of nitrate in soil. Aerobic nitrate reduction occurred in soils when provided with both nitrate and a carbon source, with the greatest nitrite accumulation in the wetland sites. The addition of a nitrification inhibitor did not significantly reduce aerobic nitrate reduction activity, nor did an assimilation repressor. Average nitrite production in soils with added carbon, nitrate, and nitrification inhibitor ranged from 7.5 to 50% of added N-nitrate in a five-hour incubation. Bacteria capable of aerobic nitrate reduction were readily isolated from these soils, comprising approximately 35% of the isolates retrieved. Sequencing16S rDNA of these isolates revealed both gram-negative and gram-positive bacteria, with the majority being gram-negative proteobacteria. In six of the isolates, onset of nitrate reduction occurred at 45–86% of atmospheric oxygen concentrations. Reduction of nitrate under aerobic and semi-aerobic conditions did not result in significant enhancements in carbon dioxide production or total electron flow rate to electron acceptors. The genomes of these six isolates were sequenced and targeted RT-qPCR revealed a wide diversity of regulatory controls on the nitrate reductase(s). The results suggest that aerobic nitrate reduction can occur in diverse bacteria, have multiple types of physiological controls, and can occur independently of the gas-forming reactions of denitrification. Thus, it is an unappreciated fate of nitrate in soil.