C4-HSL-mediated quorum sensing regulates nitrogen removal in activated sludge process at Low temperatures

The activated sludge process faces challenges in achieving adequate nitrification ability under low-temperature conditions. Therefore, we investigated the effects of different concentrations of exogenous N-butyryl-homoserine lactone (C4-HSL) on nitrogen removal in lab-scale sequencing batch reactors (SBRs) at 10 °C. The results revealed that both 10 and 100 μg/L of C4-HSL could improve NH₄⁺-N removal efficiency by 26% and reduce the effluent TN concentration to below 15 mg/L. Analysis of extracellular polymeric substances (EPS) revealed that adding C4-HSL (especially 100 μg/L) reduced the protein-like substance content while increasing the humic and fulvic acid-like substance content in EPS. Protein-like substances could serve as carbon sources for denitrifiers, thus promoting denitrification. Moreover, exogenous C4-HSL increased the abundance of bacteria and genes associated with nitrification and denitrification. Further analysis of quorum sensing (QS) of microorganisms indicated that exogenous C4-HSL (especially 100 μg/L) promoted regulation, transportation, and decomposition functions in the QS process. Furthermore, CS, sdh, fum, and mdh gene expressions involved in the tricarboxylic acid (TCA) cycle were enhanced by 100 μg/L C4-HSL. Exogenous C4-HSL promoted microbial communication, microbial energy metabolism, and nitrogen metabolism, thereby improving the nitrogen removal efficiency of activated sludge systems at low temperatures. This study provides a feasible strategy for enhancing denitrogenation performance at low temperatures through exogenous C4-HSL.