Achieving high-level nitrogen removal in mainstream by coupling anammox with denitrifying anaerobic methane oxidation in a membrane biofilm reactor

To achieve energy neutral wastewater treatment, mainstream anaerobic ammonium oxidation (anammox) has attracted extensive attention in the past decade. However, the relatively high effluent nitrogen concentration (>10 mg N L^-1) remains a significant barrier hindering its practical implementation. A novel technology integrating the anammox and denitrifying anaerobic methane oxidation (DAMO) reactions in a membrane biofilm reactor (MBfR) was developed in this study to enhance the mainstream anammox process. With the hydraulic retention time (HRT) progressively decreased from 12 to 4 h, the total nitrogen (TN) removal rate increased stepwise from 0.09 to 0.28 kg N m⁻³ d⁻¹, with an effluent TN concentration below 3.0 mg N L^-1 achieved. Mass balance analysis showed that 30–60% of the nitrate produced by the anammox reaction was reduced back to nitrite by DAMO archaea, and the anammox and DAMO bacteria were jointly responsible for nitrite removal with contributions of >90% and <10%, respectively. Additionally, the established MBfR was robust and achieved consistently high effluent quality with >90% TN removal when the influent nitrite to ammonium molar ratio varied in the range of 1.17−1.55. Fluorescence in situ hybridization (FISH) and 16S rRNA gene sequencing indicated that anammox bacteria, DAMO bacteria and DAMO archaea jointly dominated the biofilm, and were likely the key contributors to nitrogen removal. This is the first study that a high nitrogen removal rate (>0.2 kg N m⁻³ d⁻¹) and satisfactory effluent quality (∼3 mg TN L^-1) were achieved simultaneously by integrating anammox and DAMO reactions in mainstream wastewater treatment.