Irreversible nitrite-oxidising bacteria inhibition by operational shutdowns for high‑ammonia wastewater treatment: from mechanism insights to pilot‑scale granular sludge reactor verification

• Operational shutdowns selectively suppressed NOB. • Combined starvation led to irreversible NOB gene loss and gene transcription. • Pilot-scale tests showed shutdown increased and stabilised NAR at about 98% • Short shutdowns enrich N.europaea and the prolonged enrich N. eutropha. • Operational shutdowns can be a practical strategy for PN enhancement. Short-cut nitrogen removal is a promising strategy for treating high-ammonia wastewaters but is hindered by the persistence of nitrite-oxidising bacteria (NOB). This study evaluated whether operational shutdowns could selectively suppress NOB while preserving ammonia-oxidising bacteria (AOB) in granular sludge reactors (GSRs). Batch assays exposed sludge to 3–20 days of shutdown followed by reactivation. Shutdowns longer than 3 days caused irreversible NOB activity loss and four-log decreases in nxrB abundance, whereas AOB maintained stable amoA copies and showed increased expression after reactivation. Pilot-scale validation in a 275-L GSR treating high-ammonia centrate confirmed these findings, demonstrating stabilised ammonia removal, enhanced granulation, and improved biomass retention and settling. Microbial community analysis revealed enrichment of Nitrosomonas europaea after short shutdowns and Nitrosomonas eutropha under prolonged starvation. Overall, shutdowns provide a simple and effective strategy for selective NOB suppression, enabling resilient partial nitritation and practical sludge preservation for full-scale emergency management.