Autoinducer-2 and acyl homoserine lactones have contrasting effects on ammonia and nitrite-oxidizing sludge

Abstract Enhancing nitrification with quorum sensing manipulation has emerged as a promising strategy to overcome rate-limiting steps. This study examined how disrupting microbial cell-to-cell communication by supplementing or quenching signal molecules regulates ammonia– and nitrite-oxidizing activity within activated sludge. Prolonged enrichment of activated sludge over 180 days yielded stable and robust nitrifying consortia, increasing the ammonia oxidation rate (AOR) from 5.4 to 9.3 mg N g⁻¹ VSS h⁻¹ and the nitrite oxidation rate (NOR) from 0.6 to 4.8 mg N g⁻¹ VSS h⁻¹, while reducing the hydraulic residence time by 50 % (from 72 h to 36 h). Exogenous addition of oxoacyl and long-chain acyl homoserine lactone (AHL) signals further boosted AOR up to 4.5-fold higher than the control activated sludge, predominantly through transcriptional activation of the amoA gene in Nitrosomonas eutropha . Acylase-mediated AHL quenching lowered AOR to 4.8 mg N g⁻¹ VSS h⁻¹ but improved functional resilience of nitrite oxidizing bacteria by enhancing mass transfer and oxygen diffusion via smaller flocs (123 µm vs 357 µm in AHL-treated sludge). Conversely, elevated autoinducer-2 (AI-2) levels suppressed ammonia-oxidizing activity (AOR = 2 mg N g⁻¹ VSS h⁻¹) yet stimulated nitrite oxidation (NOR = 36 mg N g⁻¹ VSS h⁻¹), particularly Nitrospira , underscoring the contrasting regulatory requirements of the two nitrifying guilds. Overall, the study demonstrates that AHLs and AI-2 serve as complementary yet opposing regulators of nitrification, primarily activating ammonia oxidizers and nitrite oxidizers, respectively. Maintaining these signaling molecules within an optimal physiological window thereby offers a biologically tunable approach for synchronized ammonia and nitrite oxidation, ultimately maximizing nitrogen removal in biological treatment systems.