Mechanistic shifts driving nitrous oxide mitigation in mainstream partial nitritation/anammox: Transitioning from nitrifier denitrification to hydroxylamine oxidation

Achieving stable partial nitritation/anammox (PN/A) for mainstream municipal wastewater treatment while minimizing nitrous oxide (N₂O) emissions remains a key challenge due to slow anammox bacteria growth and nitrite oxidizing bacteria (NOB) proliferation. In this study, a stable mainstream PN/A system was established in a biological aerated filter (BAF) using an alternating influent strength and concentration strategy, leveraging a high-strength ammonium model as a mechanistic baseline. Results demonstrated that the alternating strategy sustained a robust nitrogen removal efficiency of >82 % under mainstream conditions, while reducing the N₂O emission factor by 80 % compared to high-load operation. Spatially, the upper filter bed functioned as an effective N₂O sink, actively consuming residual dissolved N₂O via heterotrophic denitrification supported by the selective enrichment of nosZ genes. Crucially, isotopic analysis revealed a fundamental metabolic shift: high-strength conditions favored stress-induced nitrifier denitrification (F_ND, ∼59 %) driven by nitrite accumulation, whereas mainstream conditions shifted dominance to hydroxylamine oxidation (F_NN, ∼78 %), representing a lower-intensity metabolic leakage. Furthermore, strategic hydraulic backwashing combined with periodic high free ammonia (FA) pulses effectively suppressed NOB activity through metabolic hysteresis and selective washout of outer biofilm layers. These findings elucidate the key metabolic drivers governing N₂O transformation and validate the alternating influent BAF process as a robust and scalable operational strategy for achieving simultaneous nitrogen removal and greenhouse gas mitigation in mainstream municipal wastewater treatment.