Ultra-high anammox-based nitrogen removal contribution and robust bacterial abundance in a pure-biofilm anoxic/oxic system for real municipal wastewater treatment.

[Display omitted] • Anammox coupled with partial denitrification enhanced to nitrogen removal above 55%. • Ultra-high abundance of Ca. Brocadia (5.5, 6.5%, 5.8%) in connected anoxic biofilms. • Ca. Brocadia sapporoensis is acetate-resistant, explaining its high enrichment in biofilms. • Anammox activity increased with decreasing denitrifying activity along anoxic zones. For municipal wastewater treatment via partial-denitrification/anammox (PD/A) in the anoxic zone, the primary challenge is effective enrichment of anaerobic ammonium oxidation (anammox) bacteria with a stable autotrophic nitrogen removal performance. This long-term study investigated a pure-biofilm anoxic/oxic reactor using real municipal wastewater with an ammonia concentration of 49.1 ± 3.6 mg N/L, achieving a total inorganic nitrogen concentration of 11.2 ± 3.5 mg N/L in the effluent, resulting in a removal efficiency of 77.4 ± 7.2%. Driven by partial denitrification, the anammox contribution to total nitrogen removal represented an ultra-high level over 55%, and was considerable in the first anoxic zone. Candidatus Brocadia, the only anammox genus detected by 16S rRNA gene sequencing, reached robust abundances of 5.46, 6.49 and 5.79% in the three sequentially connected anoxic zones. Metagenomic sequencing further identified that the main species was Ca. Brocadia sapporoensis, which is inhibited by nitrite but not by acetate, supporting its high-level enrichment under acetate conditions in the anoxic zone. Batch tests indicate that heterotrophic denitrification activity decreased while anammox activity increased along the anoxic zones, and that excessive heterotrophic denitrifying activity was not conducive to increasing the anammox contribution. This study achieved robust anammox bacterial abundance using a pure-biofilm system, shedding fresh light on the application of mainstream anammox in municipal wastewater treatment. [ABSTRACT FROM AUTHOR]