A permeability-increasing drug synergizes with bacterial efflux pump inhibitors and restores susceptibility to antibiotics in multi-drug resistant Pseudomonas aeruginosa strains

Raquel Ferrer-Espada,Philip S Stewart,Susana Sánchez-Gómez,Hawraa Shahrour,Betsey Pitts,Guillermo Martínez-De-Tejada

doi:10.1038/s41598-019-39659-4

Abstract

Resistance to antibiotics poses a major global threat according to the World Health Organization. Restoring the activity of existing drugs is an attractive alternative to address this challenge. One of the most efficient mechanisms of bacterial resistance involves the expression of efflux pump systems capable of expelling antibiotics from the cell. Although there are efflux pump inhibitors (EPIs) available, these molecules are toxic for humans. We hypothesized that permeability-increasing antimicrobial peptides (AMPs) could lower the amount of EPI necessary to sensitize bacteria to antibiotics that are efflux substrates. To test this hypothesis, we measured the ability of polymyxin B nonapeptide (PMBN), to synergize with antibiotics in the presence of EPIs. Assays were performed using planktonic and biofilm-forming cells of Pseudomonas aeruginosa strains overexpressing the MexAB-OprM efflux system. Synergy between PMBN and EPIs boosted azithromycin activity by a factor of 2,133 and sensitized P. aeruginosa to all tested antibiotics. This reduced several orders of magnitude the amount of inhibitor needed for antibiotic sensitization. The selected antibiotic-EPI-PMBN combination caused a 10 million-fold reduction in the viability of biofilm forming cells. We proved that AMPs can synergize with EPIs and that this phenomenon can be exploited to sensitize bacteria to antibiotics.

Highlights

At the beginning of 2017, the World Health Organization issued for the first time in its history a global priority list of antibiotic-resistant bacteria[1]
We have demonstrated for the first time that a permeabilizing agent at subinhinbitory concentrations is able to sensitize strains that overexpress MexAB-OprM to a combination of a substrate antibiotic and an efflux pump inhibitors (EPIs), being the combination unsuccessful in the absence of the permeabilizing agent
We found that at subinhibitory concentrations, polymyxin B nonapeptide (PMBN)-mediated potentiation of PAβN resulted in sensitization to antibiotics by factors of up to 2,133 in the mutant strain P. aeruginosa LC1-6 (Table 2; Supplementary Table 1a)

Summary

Introduction

At the beginning of 2017, the World Health Organization issued for the first time in its history a global priority list of antibiotic-resistant bacteria[1]. P. aeruginosa possesses both intrinsic and adaptive resistance to a wide variety of antimicrobials and frequently causes bacteremia, healthcare related pneumonia and urinary tract infections[2]. This organism is the most common bacterial species infecting the respiratory tract in cystic fibrosis patients[3]. P. aeruginosa has 12 resistance-nodulation-division (RND)-type efflux systems, being MexAB-OprM the best characterized[4]. This pump is constitutively expressed and exhibits an incredibly high ability to capture and extrude very structurally different antimicrobials including β-lactams, fluoroquinolones, macrolides, tetracyclines, trimethoprim, sulfamides and chloramphenicol[12]. The deletion of some regulatory genes such as mexR, nalD and nalC derepresses the system, thereby increasing bacterial resistance to its substrates[13]

Methods

Results

Conclusion