Nanosized Combined Antimicrobial Drugs Decreased Emergence of Resistance in Escherichia coli: A Future Promise.

Abstract

Antibiotic combinations remain the frontline therapy for severe infections to reduce mortality. However, conventional antibiotic combinations have some limitations such as the low bioavailability and the rise of resistant strains. Nanoparticles are increasingly used as antibiotic delivery systems to promote bioavailability and hence improve efficacy of antibiotics. In this work, we hypothesize that the simultaneous delivery of two antibiotic-loaded nanoparticles will improve the intracellular bioavailability and thus inhibit emergence of resistance. Accordingly, Chitosan-pluronic nanoparticles were used to construct nanosized ciprofloxacin and meropenem and the antibacterial activity of nanosized combined antibiotics were compared versus unloaded single, unloaded combined, and nanosized single antibiotics. Thirty-six stepwise mutants were selected by exposing two E. coli strains to increasing concentrations of free-unloaded and nanosized antibiotics, and mutants were tested for antimicrobial susceptibilities using broth microdilution and disc diffusion methods. The change in expression levels of acrB efflux pump and porins (ompC and ompF) was assessed by real-time reverse transcription-PCR. The in vitro evaluation of combined ciprofloxacin and meropenem-loaded nanoparticles demonstrated that this nanosystem exhibited enhanced antibacterial effect. Step mutants selected with nanosized combined antibiotics showed higher sensitivity to both drugs, exhibited lower mutation frequencies, and less cross-resistance to other antimicrobial classes. Moreover, for all steps of selection, nanosized combined antibiotic mutants expressed significantly lower levels of acrB as well as higher levels of ompC and ompF (p-value <0.01). In view of these results, the use of nanosized combined antibiotics may be considered among the new promising strategies to combat infections through their potential efficacy in reducing microorganisms' ability to form resistant mutants.