Characterization of phytocompounds inhibitors from Commelina communis against AcrAB-TolC efflux pump of a pathogenic multidrug-resistant Aeromonas hydrophila: By virtual screening and bioassays

Multidrug-resistant (MDR) pathogens are a significant threat to aquaculture due to their resistance to most conventional drugs. AcrAB-TolC is a known MDR efflux pump that increases antibiotic resistance. Inhibiting such pumps is a promising strategy in drug discovery. In this study, phytocompounds derived from Commelina communis (Asiatic dayflower) have been identified as candidate inhibitors for AcrB of Aeromonas hydrophila through virtual screening, encompassing molecular docking and molecular dynamics (MD) simulation. Consequently, five phytocompounds from the Asiatic dayflower met the criteria as suggested by the TCMSP database. Among them, flavocommelin, luteolin, and delphin_qt, exhibiting values higher than or equal to the control PAβN, emerged as candidate inhibitors of AcrB protein according to molecular docking analysis. Luteolin, which adhered to Lipinski's rule of five, was stable in binding to AcrB in a 50 ns molecular dynamics simulation. Luteolin also attenuated the reduction in Nile Red fluorescence in a dose-dependent manner, indicating the actual inhibition of the AcrAB-TolC efflux pump of the MDR A. hydrophila Y-1. The checkerboard synergy analysis revealed that luteolin exerted a positive impact on the six tested antibiotics, with fractional inhibitory concentration (FIC) indexes ranging from 0.125 to 0.375. Luteolin and florfenicol were recognized for having the best synergistic activity. When Ctenopharyngodon idella (grass carp) and Plecoglossus altivelis (ayu) were infected by A. hydrophila Y-1, the combination of luteolin and florfenicol markedly enhanced the survival rate and declined the bacterial load, as compared to either luteolin or florfenicol alone. Thus, luteolin is an effective efflux pump inhibitor for AcrAB-TolC of A. hydrophila , enhancing antibiotic sensitivity and exhibiting synergistic effects with antibiotics.