Hydroxylamine stabilizes low-ammonium comammox-anammox cooperation through selective metabolic regulation of complete...

In low-ammonium municipal wastewater treatment, the coupling of comammox and anammox represents a promising strategy for energy-efficient nitrogen removal. However, the process exhibits considerable instability under fluctuating influent ammonium concentrations and low-ammonium conditions. Substrate limitation often drives comammox to shift from partial to complete nitrification, thereby disrupting the supply of nitrite to anammox. Moreover, the mechanisms underlying their synergistic interactions at the metabolic regulation level have yet to be elucidated. Herein, a sequencing batch biofilm reactor (SBBR) was operated with stepwise decreased ammonia concentration (33 to 10 mg-N·L^-1) and subsequently amended with 5 mg L^-1 hydroxylamine (NH₂OH) to probe system performance and microbial synergy. Results demonstrated that NH₂OH addition recovered the total nitrogen removal efficiency to 66.7% by selectively enhancing comammox-driven partial nitritation while suppressing complete nitrification, as evidenced by upregulated amoA expression and inhibited nxrAB activity. Additionally, hydroxylamine enhanced the microbial network's resilience in biofilms under substrate-limited conditions and stimulated the expression of dissimilatory nitrate reduction to ammonium (DNRA) genes, providing an alternative nitrogen retention route. The stimulated electron recycling and interspecies communication further reinforced the metabolic coupling between comammox and anammox bacteria. This study elucidates a novel hydroxylamine-mediated regulatory strategy for the comammox-anammox coupling system, providing critical insights into stabilizing synergistic nitrogen removal in carbon-limited mainstream environments.