Evidence for Nitrous Oxide Emissions by Nitrite-Dependent Anaerobic Methane Oxidizing Bacteria

Nitrite-dependent anaerobic methane oxidizing (n-DAMO) bacteria generally convert nitrite to dinitrogen and bypass the nitrous oxide (N₂O) formation step. However, N₂O is often detected in n-DAMO bacteria dominated cultures and it remains an open question as to the microbial origin of N₂O in these enrichments. Using a stable nitrite consuming microbial community enriched for n-DAMO bacteria, we demonstrated that N₂O production was coupled to methane oxidation and the higher initial nitrite concentrations led to increased quantities of N₂O being formed. Moreover, continuous exposure of the enrichment culture to about 5 mg of N L^-1 nitrite resulted in constant N₂O being produced (12.5% of nitrite was reduced to N₂O). Metatranscriptomic analyses revealed that nitrite reductase (nirS) and nitric oxide reductase (norZ) transcripts from n-DAMO bacteria increased in response to nitrite exposure. No other bacteria significantly expressed nor genes under these conditions, suggesting n-DAMO bacteria are responsible for N₂O being produced. In a 35-day bioreactor experiment, N₂O produced by the n-DAMO bacteria accumulated when nitrite was in excess; this was found to be up to 3.2% of the nitrogen that resulted from nitrite removal. Together, these results suggested that excess nitrite is an important driver of N₂O production by n-DAMO bacteria. To this end, proper monitoring and control of nitrite levels in wastewater treatment plants would be effective strategies for mitigating N₂O emissions to the atmosphere.