Role of Microoxic Control in Alternating Anoxic/Oxic Systems for Selective Enrichment of N ₂ O-Reducing Bacteria and N ₂ O Mitigation in Wastewater Treatment

The biological reduction of N2O to N2 relies on N2O-reducing bacteria (N2ORB), which involves enhancement of the N2O-reducing enzyme (N2OR) and selection of N2ORB. This study investigates the short-term effects of microoxic pretreatment on N2OR and long-term operational strategies for wastewater treatment aimed at achieving N2O mitigation. In denitrification batch experiments, microoxic pretreatment (with dissolved oxygen (DO) at 0.2 mg/L for 2 h) enhanced N2OR levels to 1.27 times that of the control group, resulting in a 35% to 65% reduction in N2O emission flux. Through long-term reactor operation (294 days), controlling the microoxic environment during the oxic stage revealed that the second anoxic stages of the anoxic/oxic/anoxic (AOA) or anoxic/oxic/anoxic/oxic (AOAO) processes emitted no N2O. Isotope analysis confirmed a high N2O reduction ratio of 90.46%. The coexpression of nosZ with cytochrome c oxidase and CRP/FNR transcriptional regulators in clade I N2ORB reflects the metabolic versatility of denitrifiers, driven by the integrated environmental selection of long-term microoxic conditions in oxic stages coupled with step-feeding strategies in anoxic stages. These findings highlight the AOA and AOAO processes under such controls as effective strategies for achieving both efficient nitrogen removal and N2O mitigation.