Abstract
The emergence of antimicrobial resistance severely threatens our ability to treat bacterial infections. While acquired resistance has received considerable attention, relatively little is known of intrinsic resistance that allows bacteria to naturally withstand antimicrobials. Gene products that confer intrinsic resistance to antimicrobial agents may be explored for alternative antimicrobial therapies, by potentiating the efficacy of existing antimicrobials. In this study, we identified the intrinsic resistome to a broad spectrum of antimicrobials in the human pathogen, Staphylococcus aureus. We screened the Nebraska Transposon Mutant Library of 1920 single-gene inactivations in S. aureus strain JE2, for increased susceptibility to the anti-staphylococcal antimicrobials (ciprofloxacin, oxacillin, linezolid, fosfomycin, daptomycin, mupirocin, vancomycin, and gentamicin). Sixty-eight mutants were confirmed by E-test to display at least twofold increased susceptibility to one or more antimicrobial agents. The majority of the identified genes have not previously been associated with antimicrobial susceptibility in S. aureus. For example, inactivation of genes encoding for subunits of the ATP synthase, atpA, atpB, atpG and atpH, reduced the minimum inhibitory concentration (MIC) of gentamicin 16-fold. To elucidate the potential of the screen, we examined treatment efficacy in the Galleria mellonella infection model. Gentamicin efficacy was significantly improved, when treating larvae infected with the atpA mutant compared to wild type cells with gentamicin at a clinically relevant concentration. Our results demonstrate that many gene products contribute to the intrinsic antimicrobial resistance of S. aureus. Knowledge of these intrinsic resistance determinants provides alternative targets for compounds that may potentiate the efficacy of existing antimicrobial agents against this important pathogen.
Highlights
Antibiotic resistant bacteria are a growing global issue that reduces our ability to cure bacterial infections (Gould, 2009; Davies and Davies, 2010)
We identified genetic determinants contributing to the intrinsic resistance of S. aureus to eight different antimicrobials
The complete Nebraska Transposon Mutant Library (NTML) of 1920 single-gene inactivations (Fey et al, 2013) was screened for mutants displaying increased susceptibility to eight antimicrobial agents with different mechanism of action, by inspecting for lack of growth on agar plates supplemented with 0.5x minimum inhibitory concentration (MIC) of the respective antimicrobials
Summary
Antibiotic resistant bacteria are a growing global issue that reduces our ability to cure bacterial infections (Gould, 2009; Davies and Davies, 2010). Determinants of Antimicrobial Intrinsic Resistance in S. aureus bacteria to an antimicrobial agent by potentiating the efficacy of an antimicrobial with a helper-drug. Intrinsic resistance to antimicrobials has traditionally been attributed to reduced permeability of the cell envelope, presence of inactivating enzymes or efflux pumps that can extrude the antimicrobial agents (Cox and Wright, 2013). An analogous approach has been pursued by limiting the active efflux of antimicrobial agents by efflux pump inhibitors (Lomovskaya and Bostian, 2006), which have been shown to potentiate the efficacy of, e.g., levofloxacin in Pseudomonas aeruginosa (Renau et al, 1999) and norfloxacin in Staphylococcus aureus (Stermitz et al, 2000). Efflux pumps inhibitors have not yet been approved for treatment of human infections due to tolerability issues (Fernebro, 2011)
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