Decoupling biodegradation from substrate concentration via quorum sensing: A cell density-responsive strategy for robust chlorobenzene removal.

Chlorinated aromatic compounds are important industrial pollutants that pose persistent environmental risks due to their toxicity and recalcitrance. Biological treatment is a sustainable approach for chlorinated VOC removal. However, conventional bioremediation is constrained by an inherent dilemma in native regulatory systems: trace substrate concentrations trigger insufficient gene expression, while shock loads induce severe metabolic inhibition and cytotoxicity. To overcome these limitations, we leveraged quorum sensing (QS) to reprogram gene regulation in Pseudomonas putida, redirecting degradation control from substrate concentration to cell density. The native, pollutant-inducible promoter of the tod operon was replaced with the QS-responsive promoter PlasA, generating the engineered strain F1-QS. At low chlorobenzene concentrations, F1-QS increased tod transcription by 9.7‑fold, reduced the lag phase from 6 h to 2 h, and enhanced the degradation rate by 38.91%. Critically, this advantage extended to high pollution loads. At 325 mg/L chlorobenzene, F1-QS achieved 88.0% removal within 32 h, outperforming the wild-type strain (35.9%) by more than two-fold. In a synthetic microbial community, the F1-QS strain accelerated consortium degradation by 70.93% and maintained stable performance during long-term operation, highlighting its superior efficacy and resilience under substrate-limited conditions. Metagenomic analysis revealed that enhanced degradation performance was associated with higher abundance of key chlorobenzene degradation genes, suggesting an enhanced functional potential of the microbial consortium. This study shows that QS‑based rewiring resolves the trade‑off between weak induction at low concentrations and toxicity at high concentrations, offering a robust strategy for chlorinated VOC bioremediation.