A Novel Screening Strategy Reveals ROS-Generating Antimicrobials That Act Synergistically against the Intracellular Veterinary Pathogen Rhodococcus equi

Abstract

: Rhodococcus equi is a facultative intracellular pathogen that causes infections in foals and many other animals such as pigs, cattle, sheep, and goats. Antibiotic resistance is rapidly rising in horse farms, which makes ineffective current antibiotic treatments based on a combination of macrolides and rifampicin. Therefore, new therapeutic strategies are urgently needed to treat R. equi infections caused by antimicrobial resistant strains. Here, we employed a R. equi mycoredoxin-null mutant strain highly susceptible to oxidative stress to screen for novel ROS-generating antibiotics. Then, we used the well-characterized Mrx1-roGFP2 biosensor to confirm the redox stress generated by the most promising antimicrobial agents identified in our screening. Our results suggest that different combinations of antibacterial compounds that elicit oxidative stress are promising anti-infective strategies against R. equi. In particular, the combination of macrolides with ROS-generating antimicrobial compounds such as norfloxacin act synergistically to produce a potent antibacterial effect against R. equi. Therefore, our screening approach could be applied to identify novel ROS-inspired therapeutic strategies against intracellular pathogens.

Oxidative Stress-Generating Antimicrobials, a Novel Strategy to Overcome Antibacterial Resistance

“…These antibiotics may alter the pathogen's central metabolism and/or its iron homeostasis, which results in the production of intracellular hydrogen peroxide [107]. Recent studies have identified RONS-producing-antibiotics by expressing redox biosensors in several bacterial species (Table 1) [107][108][109][110]. In addition, the combination of different RONS-generating antimicrobials may act synergistically against some bacterial pathogens ( Figure 4A) [109].…”

“…Recent studies have identified RONS-producing-antibiotics by expressing redox biosensors in several bacterial species (Table 1) [107][108][109][110]. In addition, the combination of different RONS-generating antimicrobials may act synergistically against some bacterial pathogens ( Figure 4A) [109]. Similarly, the combination of some antibiotics with silver nanoparticles may enhance RONS biosynthesis and, therefore, increase the efficacy of the combinatorial treatment.…”

“…These novel therapeutic strategies can improve the antimicrobial activity of some drugs with repurposing potential. It is even possible that antimicrobials clinically approved to treat common infections might be used as part of combinatorial therapies against new multi-drug resistant (MDR) bacteria [90,109]. However, most of this evidence is still only based on in vitro experiments, and therefore further research is required to demonstrate the efficacy of these novel treatments against pathogenic bacteria in vivo.…”

Novel Treatments against Mycobacterium tuberculosis Based on Drug Repurposing

“…Other strategies can be based on the analysis of oxidative stress-generating compounds and their combination to increase the biosynthesis of radical oxygen species (ROS) during host cell infection [ 44 , 45 ]. During phagocytosis, M. tuberculosis is exposed to oxidative stress, but this is not sufficient to kill the pathogen [ 46 ].…”

“…Several antituberculosis drugs have been tested as promising ROS-generating antimicrobials against M. tuberculosis , and the majority of them have produced an oxidative shift during infection, especially clofazimine [ 46 ], but also rifampicin and isoniazid [ 47 ]. The combination of several ROS-generating compounds could be a new solution against intracellular M. tuberculosis as it was demonstrated for other intracellular pathogens [ 44 , 45 , 48 ].…”