Nitrate removal by anammox-ZnS biohybrid system utilizing hydraulic kinetic energy

Nitrate (NO₃^-) removal from low-carbon wastewater is challenging for conventional denitrification due to the need for external electron donors. Anammox bacteria could reduce nitrate via the dissimilatory nitrate reduction to ammonium (DNRA) pathway using intracellular glycogen, but this endogenous process suffered from slow kinetics and finite glycogen reserves. Here, we coupled anammox bacteria with piezoelectric ZnS nanoparticles under mechanical agitation (simulating hydraulic kinetic energy) to construct a piezoelectric-driven nitrate reduction system. The system achieved complete NO₃^- and total nitrogen removal from synthetic wastewater (50 mg-N/L NO₃^-) within 30 h, with the nitrate reduction efficiency enhanced approximately 2.7-fold compared to the sole endogenous DNRA-driven group, and produced no detectable nitrous oxide (N₂O). Inhibitor assays confirmed that NO₃^- conversion followed the DNRA-anammox coupled pathway and that piezoelectrons (generated by mechanical agitation of ZnS nanoparticles) served as a sustainable exogenous electron source. Transcriptomic analysis revealed that genes related to extracellular electron transfer, DNRA, and the respiratory chain were coordinately upregulated, while genes related to glycolysis were downregulated, reflecting a shift in metabolism from endogenous glycogen utilization to exogenous electron utilization. Overall, this study overcame the kinetic and electron-source limitations of endogenous DNRA in anammox, providing a hydraulic kinetic energy-driven, low-carbon, and N₂O-free strategy for removing NO₃^- from low-carbon wastewater.