Synergistic activity of colistin and stilbenoids against Pseudomonas aeruginosa clinical isolates

ABSTRACT Pseudomonas aeruginosa is a major opportunistic pathogen responsible for chronic lung infections in cystic fibrosis (CF) patients, contributing significantly to morbidity and mortality. In this study, we investigated the synergistic activity of colistin combined with two natural stilbenoids, pterostilbene and (±)- trans -ε-viniferin, against clinical P. aeruginosa isolates from CF patients. Synergy was assessed through minimum inhibitory concentration (MIC) determination and time–kill assays. The combination of colistin with pterostilbene reduced colistin MIC values in all tested strains, with the strongest effect observed in a colistin-resistant isolate (eightfold reduction, fractional inhibitory concentration index = 0.188). Time–kill assays confirmed enhanced bactericidal activity, with faster and more effective bacterial killing compared to colistin alone. According to the known mechanism of action of colistin, medium supplementation with purified lipopolysaccharide (LPS) reduced colistin activity on P. aeruginosa . Medium supplementation with stilbenoids partially restored colistin activity, reducing the negative effect of LPS. Flow cytometry analyses showed increased membrane permeability in the presence of the combination of colistin and pterostilbene, whereas elevated concentrations of divalent cations reduced this effect, highlighting the role of electrostatic interactions in colistin/pterostilbene membrane damage. Overall, these findings suggest that pterostilbene acts as a colistin adjuvant in vitro , enhancing its antimicrobial activity against P. aeruginosa through a mechanism involving cell membrane damage. IMPORTANCE Pseudomonas aeruginosa is a major cause of chronic lung infections in cystic fibrosis patients and is increasingly difficult to treat because of antibiotic resistance. Colistin remains one of the few effective last-line therapies against this bacterial pathogen, but its clinical use is limited by severe dose-dependent toxicity. In this study, we show that pterostilbene, a natural stilbenoid compound, enhances in vitro the activity of colistin against clinical P. aeruginosa isolates. The combination improved colistin efficacy, increased membrane damage, and partially overcame lipopolysaccharide-mediated reduction of colistin activity. Importantly, these effects were achieved using low concentrations of pterostilbene under the experimental conditions tested.