Genetic requirements for Staphylococcus aureus nitric oxide resistance and virulence.

Melinda R. Grosser,Marcus M. Dillon,Anthony R. Richardson,Lance R. Thurlow,Elyse Paluscio,Vaughn S. Cooper,Thomas H. Kawula,Andreas Peschel

doi:10.1371/journal.ppat.1006907

Melinda R. Grosser, Marcus M. Dillon + Show 6 more

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https://doi.org/10.1371/journal.ppat.1006907

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Abstract

Staphylococcus aureus exhibits many defenses against host innate immunity, including the ability to replicate in the presence of nitric oxide (NO·). S. aureus NO· resistance is a complex trait and hinges on the ability of this pathogen to metabolically adapt to the presence of NO·. Here, we employed deep sequencing of transposon junctions (Tn-Seq) in a library generated in USA300 LAC to define the complete set of genes required for S. aureus NO· resistance. We compared the list of NO·-resistance genes to the set of genes required for LAC to persist within murine skin infections (SSTIs). In total, we identified 168 genes that were essential for full NO· resistance, of which 49 were also required for S. aureus to persist within SSTIs. Many of these NO·-resistance genes were previously demonstrated to be required for growth in the presence of this immune radical. However, newly defined genes, including those encoding SodA, MntABC, RpoZ, proteins involved with Fe-S-cluster repair/homeostasis, UvrABC, thioredoxin-like proteins and the F1F0 ATPase, have not been previously reported to contribute to S. aureus NO· resistance. The most striking finding was that loss of any genes encoding components of the F1F0 ATPase resulted in mutants unable to grow in the presence of NO· or any other condition that inhibits cellular respiration. In addition, these mutants were highly attenuated in murine SSTIs. We show that in S. aureus, the F1F0 ATPase operates in the ATP-hydrolysis mode to extrude protons and contribute to proton-motive force. Loss of efficient proton extrusion in the ΔatpG mutant results in an acidified cytosol. While this acidity is tolerated by respiring cells, enzymes required for fermentation cannot operate efficiently at pH ≤ 7.0 and the ΔatpG mutant cannot thrive. Thus, S. aureus NO· resistance requires a mildly alkaline cytosol, a condition that cannot be achieved without an active F1F0 ATPase enzyme complex.

Highlights

Staphylococcus aureus is a highly invasive human pathogen that is responsible for significant morbidity each year[1]
The human pathogen Staphylococcus aureus is remarkably resistant to many facets of the host immune response, including the antibacterial radical nitric oxide (NOÁ)
Because our primary interest was to identify genes required for both NOÁ resistance and persistence in soft tissue infections (SSTIs), we wanted to select a Methicillin resistant S. aureus (MRSA) strain commonly associated with SSTIs in the current communityacquired MRSA (CA-MRSA) pandemic

Summary

Introduction

Staphylococcus aureus is a highly invasive human pathogen that is responsible for significant morbidity each year[1]. Historically a nosocomial pathogen, in recent decades otherwise healthy individuals have begun to contract MRSA outside of hospital settings[3]. These communityacquired MRSA (CA-MRSA) strains are typically characterized as hypervirulent and most frequently cause skin and soft tissue infections (SSTIs), infections often progress to more invasive and systemic disease[4]. Understanding the factors contributing to the persistence and invasiveness of CA-MRSA is of paramount importance to limiting its current spread through both community and hospital settings. A major factor contributing to S. aureus persistence within mammalian hosts is resistance to the antimicrobial radical nitric oxide (NOÁ), a membrane permeable gas that is produced by activated phagocytes in response to bacterial infection[5]. NOÁ resistance is important for S. aureus persistence in murine infection models because deletion of genes required for NOÁ resistance attenuates S. aureus virulence[6,8,9]

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