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Abstract
Antibiotic resistant infections are a significant and increasing cause of morbidity and mortality worldwide. In particular, infections caused by multi-drug resistant Gram negative bacteria recalcitrant to commonly used classes of antibiotics are of particular concern, as the antibiotic colistin is often the only remaining treatment option. Given the importance of this last-resort antibiotic, it is imperative that we fully understand the scope of colistin resistance mechanisms so that novel treatment strategies can be identified. In this work, we utilized an Enterobacter cloacae model of colistin heteroresistance to gain insight into this elusive resistance mechanism. The heteroresistant strain harbored both colistin resistant and susceptible subpopulations that were genetically indistinguishable yet transcriptionally distinct. The resistant subpopulation increased during antibiotic treatment, receded to baseline after subculture without drug, and was distinct from persisters. Presence of the resistant subpopulation and modification of colistin drug target lipid A was dependent on the histidine kinase gene phoQ. Colistin therapy failed to rescue mice infected with the heteroresistant strain, however treatment of mice infected with the phoQ mutant was successful, indicating that resistant bacterial subpopulations can cause antibiotic treatment failures. As a further approach to reduce the impacts of colistin resistance, we utilized a model of colistin resistant Acinetobacter baumannii to develop a resistance inhibition strategy. We identified a small molecule inhibitor of the naxD-controlled colistin resistance lipid A modification pathway via tandem in vitro and in silico screens. The inhibitor restored colistin susceptibility in a naxD-dependent manner and blocked the addition of galactosamine onto lipid A. Finally, the inhibitor reduced colistin resistance in a panel of clinical A. baumannii isolates, highlighting the potential for modulation of resistance expression to restore colistin efficacy for diverse strains. Altogether, these results provide fundamental insights into the biology of colistin resistance, and set the stage for continued development of novel therapeutics to combat antibiotic resistance in the clinic.