Collateral effects of bacteriophage resistance on antibiotic sensitivity in Pseudomonas aeruginosa

Abstract

Collateral effect (CEs) of phage resistance can alter antibiotic sensitivity, leading to collateral sensitivity (CS). CS of phage resistance can potentially be used to design more effective phage--antibiotic combination treatments, yet insufficient knowledge on the occurrence of such phage resistance associated CS is currently available. Here, we characterized CEs across diverse Pseudomonas aeruginosa strains, antibiotics, and phages. For each of eight strains (seven clinical isolates and PAO1), we generated mutants resistant to phage PAO1-1 and, separately, mutants resistant to each of six antibiotics. Phage-resistant mutants were also generated using four additional phages (M32, NP1, NP3, and PNM), and double phage- and antibiotic-resistant mutants from one strain. We measured growth curves across antibiotic concentration ranges after which the area-under-the-curve (AUC) was quantified as a measure of bacterial burden. Dose-response curves for AUC against antibiotic concentrations were constructed to estimate a fold change in EC50 of mutants relative to their parental strains. PAO1-1-resistant mutants showed significant CS to colistin and polymyxin B. For other antibiotics, CEs varied by phage and parental strain: PAO1-1-resistant mutants showed CS to tobramycin, whereas mutants resistant to the other four phages tested were associated with CR. For double phage- and antibiotic resistant mutants we found CS to colistin and tobramycin relative to the parental strain. Resistance mutations occurred predominantly in genes involved in metabolism, virulence, and DNA replication. Antibiotic-resistant mutants also showed increased phage sensitivity, with the strongest effect in colistin-resistant mutants. In conclusion this study demonstrates collateral effects of phage resistance are most specific to both strain- and antibiotic-specific.