Spatial Coupling of Feammox and NDFO Processes in an Iron-Integrated Membrane-Aerated Biofilm Reactor

Abstract Iron-reduction coupled with anaerobic ammonium oxidation (Feammox) often coexists with multiple Fe–N transformation pathways, such as nitrate-dependent Fe (II) oxidation (NDFO). However, an imbalance between iron supply and consumption across these pathways impedes the establishment of a stable Feammox-based process. Here, we established an Fe–O dual-driven strategy using an Fe2O3-integrated membrane-aerated biofilm reactor (MABR) to sustain Feammox activity. Results demonstrated that the pump-free diffusive aeration mode effectively attenuated the interfacial oxygen concentration, while the integrated Fe2O3 enhanced the oxygen transfer rate (OTR) by 26.87% via physicochemical retention and biological regulation. This concerted mechanism restricted oxygen penetration depth, thereby broadening the colonization niche for anaerobes. Cryosectioning-16S rRNA sequencing combined with microelectrode analysis confirmed this spatial reorganization: Feammox bacteria migrated to the middle-outer regions, and NDFO bacteria were enriched across all layers, whereas nitrifiers were confined to the inner biofilm. This stratified architecture promoted O2-driven Fe (II) oxidation and nitrification-derived nitrate generation, effectively regenerating the Fe (III) pool for a stable Fe–N–O cycle. Consequently, the system achieved a nitrogen removal rate of 0.67 g N m–2 d–1 with 68.15% N2 selectivity. These findings deepen our understanding of microbial interactions within biofilms and advance Feammox-based nitrogen removal in wastewater treatment.