Electron transfer mechanisms underlying magnetite-mediated nitrate removal in electro-assisted mixotrophic denitrification for recirculating mariculture wastewater treatment

Efficient nitrate removal from recirculating mariculture wastewater is constrained by low carbon-to-nitrogen ratio and high nitrate concentrations. Although mixotrophic denitrification can integrate organic- and sulfur-driven pathways, its performance is still limited by inefficient electron transfer and poorly coordinated sulfur and nitrogen metabolism. In this study, magnetite was immobilized on a carbon-felt anode, and an electro-assisted mixotrophic denitrification system was constructed to regulate electron flow. At a shortened hydraulic retention time of 12 h, more than 99% NO x - -N removal was maintained in the magnetite-modified system, while sulfate accumulation was reduced. Elemental sulfur accumulation of 168.7 mg/L was achieved, corresponding to 2.3 and 1.9 times the levels measured in the non-electrode and bare-anode system, respectively, indicating that sulfur conversion was redirected toward elemental sulfur. Following anode modification, increased redox responses associated with flavin- and cytochrome c -like components in extracellular polymeric substances were observed. Intracellular electron transport was concurrently intensified, as evidenced by a 61.5% increase in electron transport system activity and enhanced nitrate and nitrite reductase activities. Distinct microbial community restructuring was observed, while coordinated changes in functional profiles related to sulfur oxidation ( sox and fccAB ), electron transport, and denitrification ( napAB , nirK , and nosZ ) were identified. Linkages among sulfur oxidation, respiratory electron transport, and nitrate/nitrite reduction were further revealed by correlation analysis. These findings improve understanding of electron-transfer regulation and its role in nitrate removal.