Effects of cooperative regulation by long-and short-chain quorum sensing signaling molecules on metabolic efficacy of sulfamethoxazole in electroactive biofilms

Wastewater containing high concentrations of sulfamethoxazole （SMX） significantly inhibits anaerobic microorganisms， leading to reduced organic matter degradation efficiency and decreased methane yield in anaerobic digestion processes. This study investigated the effects of adding short-chain （C6-HSL） and long-chain （C12-HSL） signaling molecules on the construction， performance， and antibiotic resistance genes （ARGs） of anaerobic electroactive biofilm systems within a microbial electrochemically coupled anaerobic digestion （MEC-AD） system. Results indicated that， compared to the control group （without added signaling molecules）， the relative SMX removal rates in the C6-HSL group （T1）， the C12-HSL group （T2）， and the dual-treated group （T3） increased by 9.26%， 7.44%， and 10.67%， respectively， while methane production rates increased by 20.4%， 16.9%， and 23.1%， respectively. The signaling molecule N-acyl-γ-lactones （AHLs） significantly enhanced the attachment of electroactive microorganisms to the anode electrode by promoting extracellular polymer production. Microbial community analysis revealed that AHLs enhanced microbial diversity and regulated the abundance of key functional bacteria. At the genus level， Georgenia exhibited significantly differential responses to different signal molecules： relative abundance increased by 16.77% and 36.47% in T1 and T3 groups， respectively， while decreasing by 15.99% in T2 group. ARGs analysis results showed that single signaling molecules increased the abundance of intl1， sul1， and sul2. However， the T3 group exhibited a milder response， with sul2 abundance decreasing by 4.92% compared to the control group. This finding indicates that the synergistic effect of composite AHLs helps mitigate the amplification of potential ARG-carrying bacteria. This study reveals， for the first time， the role of short-chain and long-chain composite AHLs in promoting electroactive biofilm formation， maintaining microbial community stability， and regulating ARG transmission risks. It can provide a feasible pathway for developing antibiotic wastewater treatment and risk management strategies based on quorum sensing regulation.