Hydrazine-mediated control of denitrifiers overgrowth: Dual mechanisms for suppressing denitrifiers while stimulating anammox bacteria in anammox-denitrification systems

Abstract

In simultaneous anammox-denitrification (SAD) systems, the overgrowth of denitrification bacteria severely impairs nitrogen removal efficiency by competing with anammox bacteria for nitrite under organic-rich conditions. In this study, we propose a hydrazine (N₂H₄)-based strategy to selectively inhibit denitrification bacteria while stimulating the anammox activity. Batch experiments revealed a concentration-dependent dual role of N₂H₄: at 10 mg N/L, the anammox activity increased to 123 % of the baseline performance, whereas denitrification activity declined to 59 %. Higher concentrations (≥50 mg N/L) progressively suppressed both anammox and denitrification processes. A dual inhibitory mechanism for N₂H₄ was proposed: (1) disrupting electron carriers (e.g., cytochrome c) and (2) inactivating key metabolic enzymes like hydrazine dehydrogenase (HDH) or nitrate reductase (Nar). In SAD systems, short-term (5 h) exposure to 10 mg N/L N₂H₄ effectively curtailed denitrifier overgrowth, elevating the ammonium removal rate from 96.2 ± 0.3 to 116.4 ± 2.5 mg N/g VSS·d, and reducing COD degradation rates from 345.5 ± 13.0 to 268.0 ± 9.7 mg N/g VSS·d. However, the total nitrogen (TN) removal efficiency and the proportional contribution of anammox to TN removal remained suboptimal compared to the control, suggesting that future studies should explore either higher concentrations or extended exposure durations of N₂H₄ shock to achieve optimal performance. These findings highlight the dual role of N₂H₄ in modulating microbial competition and propose its application as a control agent to optimize the SAD performance under the organic stress.