Farmer, M.; Sabba, F.; Wells, G. F.

Nitrous oxide (N2O) is a potent greenhouse gas produced as an unintentional, undesired byproduct in many nitrogen removal bioprocesses. Given the considerable challenges in managing N2O emissions from wastewater treatment, N2O could be reframed as a value-added product if intentionally generated and captured. This study assesses N2O production and mechanisms in a Coupled Aerobic-anoxic Nitrous Decomposition Operation with Phosphorus removal (CANDO+P) reactor. Optimal performance was achieved when the reactor was fed with a mixture of propionate and glucose, resulting in N2O was production up to 50% of influent nitrogen. Through 16S rRNA amplicon and shotgun metagenomic sequencing, we found that Candidatus Accumulibacter were the dominant phosphorus accumulating organism (PAO). Assembly of a high-quality metagenome-assembled genome showed that Ca. Accumulibacter encoded a full complete denitrification pathway from nitrite to nitrogen gas. We also found abundant populations of denitrifying glycogen accumulating organisms (GAO) and ordinary heterotrophic organisms (OHO). We also incorporated truncated denitrification pathways into a process model to predict N2O generation. N2O predictions were the most similar to observed results when the denitrification pathways of PAO, GAO, and OHO model populations reflected denitrification gene abundances from the metagenomic sequencing analysis. Our work demonstrates the feasibility of using non-VFA carbon for intentional N2O generation and provides broader insights into N2O generation and truncated denitrification pathways of denitrifying PAO and GAO.