Unravelling mechanisms of meropenem induced persistence facilitates identification of GRAS compounds with anti-persister activity against Acinetobacter baumannii

Abstract

Acinetobacter baumannii is recognized as one of the "critical" pathogens by the World Health Organisation (WHO) due to its unprecedented ability to acquire resistance genes and undergo genetic modifications. Carbapenem classes of antibiotics are considered as the "drugs of choice" against A. baumannii infections, although increasing incidence of carbapenem resistant isolates have greatly limited their efficacy in clinical settings. Nonetheless, the phenomenon of multi-drug tolerance or persistence exhibited by A. baumannii has further led to therapeutic failure of carbapenems against chronic and recurring infections. Exploring the underlying mechanisms of persistence hosted by the nosocomial pathogen, A. baumannii can facilitate the development of effective anti-persister strategies against them. Accordingly, this study investigates the characteristics and mechanisms responsible for meropenem induced persistence in A. baumannii. Furthermore, it describes the adaptation of a screening strategy for identification of potent anti-persister compounds that cumulatively act by targeting the A. baumannii membrane, inhibiting antibiotic efflux and inducing oxidative stress mediated killing. The screen identified the phytochemical compound, thymol to display excellent activity against persisters of mechanically distinct antibiotics. While meropenem exposed A. baumannii persisters exhibited multi-drug tolerance and indicated the ability to enter a Viable But Non Culturable (VBNC) state, thymol efficiently eradicated all persister cells, irrespective of their culturability. Thymol exhibited no propensity for resistance generation and also inhibited persisters of other Gram-negative pathogens, Pseudomonas aeruginosa and Klebsiella pneumoniae. Collectively, our results establish thymol to have immense potential to act either alone or as an adjunct in combination therapies against persistent infections. IMPORTANCEApart from the global catastrophe of antibiotic resistance, the phenomenon of "antibiotic tolerance" exhibited by a subpopulation of bacterial cells known as "persisters" ensue a major clinical threat. Eradication of the persister populations holds extreme importance for an improved long-term recovery from chronic and recurring bacterial infections. This study addresses the problem of antibiotic persistence prevailing in clinics and investigates its associated mechanisms in the nosocomial pathogen, Acinetobacter baumannii in reference to the antibiotic meropenem. It further describes the use of a mechanism-based screening approach for the identification of potent multi-targeting anti-persister compounds, thereby leading to the identification of GRAS (Generally Regarded As Safe) molecules exhibiting promising activity against A. baumannii persisters. This strategy can further be utilized for repurposing of FDA approved drugs or other available compound libraries, in order to identify novel anti-persister compounds.