Combination of Sequencing Batch Operation and A/O Process to Achieve Partial Mainstream Anammox: Pilot-Scale Demonstration and Microbial Ecological Mechanism

In this study, sequencing batch operation was successfully combined with a pilot-scale anaerobic biofilm-modified anaerobic/aerobic membrane bioreactor to achieve anaerobic ammonium oxidation (anammox) without inoculation of anammox aggregates for municipal wastewater treatment. Both total nitrogen and phosphorus removal efficiencies of the reactor reached up to 80% in the 250-day operation, with effluent concentrations of 4.95 mg-N/L and 0.48 mg-P/L. In situ enrichment of anammox bacteria with a maximum relative abundance of 7.86% was observed in the anaerobic biofilm, contributing to 18.81% of nitrogen removal, with denitrification being the primary removal pathway (38.41%). Denitrifying phosphorus removal (DPR) (40.54%) and aerobic phosphorus uptake (48.40%) played comparable roles in phosphorus removal. Metagenomic sequencing results showed that the biofilm contained significantly lower abundances of NO-reducing functional genes than the bulk sludge (p < 0.01), favoring anammox catabolism in the former. Interactions between the anammox bacteria and flanking community were dominated by cooperation behaviors (e.g., nitrite supply, amino acids/vitamins exchange) in the anaerobic biofilm community network. Moreover, the hydrolytic/fermentative bacteria and endogenous heterotrophic bacteria (Dechloromonas, Candidatus competibacter) were substantially enriched under sequencing batch operation, which could alleviate the inhibition of anammox bacteria by complex organics. Overall, this study provides a feasible and promising strategy for substantially enriching anammox bacteria and achieving partial mainstream anammox as well as DPR.