A high-throughput screening approach for bacterial quorum sensing inhibitors (QSIs) against Aeromonas hydrophila infection

Bacterial quorum sensing (QS) is a well-known behavior for bacterial communication and plays an important role in various bacterial biological functions. Several approaches have been developed to interrupt the QS mechanism in bacterial pathogens. This study developed a novel, high-throughput screening method combined with phage peptide display libraries and a QS biosensor system (PDQBS) to identify quorum-sensing inhibitors (QSIs). We screened 11 hit peptide candidates for QSI activity, which inhibited the C4-AHL mediated quorum-sensing system. Although the synthetic peptides showed moderate inhibitory effects and six candidate peptides ligated with a cell-penetrating peptide (CPP) dramatically inhibited QS signals. The protective efficacies of candidate DNA expression vector ligated with CPP-QSIs were tested against Aeromonas hydrophila infection in the zebrafish model. The obtained result revealed that at least two peptides (CPP21 and CPP23) exhibited an increased protective ability against A. hydrophila infection with RPSs of >50%. Furthermore, the three candidates CPP-QSIs (CPP21, CPP23, and CPP24), also showed high protective abilities with RPSs of >50% against A. hydrophila infection in the mice model. The qPCR analysis further confirmed that these selected CPP-QSIs might stimulate host immunity and inhibit the virulence of A. hydrophila. Finally, the results of dot blot and ELISA assays indicated that these CPP-QSI peptides could bind to the QS receptor protein, AhyR/CviR. Overall, this study provides a promising approach to screening novel QSI candidates for controlling QS-mediated bacterial infections.