Abiotic Conversion of Extracellular NH₂OH Contributes to N₂O Emission during Ammonia Oxidation

Abiotic processes involving the reactive ammonia-oxidation intermediates nitric oxide (NO) or hydroxylamine (NH₂OH) for N₂O production have been indicated recently. The latter process would require the availability of substantial amounts of free NH₂OH for chemical reactions during ammonia (NH₃) oxidation, but little is known about extracellular NH₂OH formation by the different clades of ammonia-oxidizing microbes. Here we determined extracellular NH₂OH concentrations in culture media of several ammonia-oxidizing bacteria (AOB) and archaea (AOA), as well as one complete ammonia oxidizer (comammox) enrichment (Ca. Nitrospira inopinata) during incubation under standard cultivation conditions. NH₂OH was measurable in the incubation media of Nitrosomonas europaea, Nitrosospira multiformis, Nitrososphaera gargensis, and Ca. Nitrosotenuis uzonensis, but not in media of the other tested AOB and AOA. NH₂OH was also formed by the comammox enrichment during NH₃ oxidation. This enrichment exhibited the largest NH₂OH:final product ratio (1.92%), followed by N. multiformis (0.56%) and N. gargensis (0.46%). The maximum proportions of NH₄⁺ converted to N₂O via extracellular NH₂OH during incubation, estimated on the basis of NH₂OH abiotic conversion rates, were 0.12%, 0.08%, and 0.14% for AOB, AOA, and Ca. Nitrospira inopinata, respectively, and were consistent with published NH₄⁺:N₂O conversion ratios for AOB and AOA.