Antibiotics Used in Empiric Treatment of Ocular Infections Trigger the Bacterial Rcs Stress Response System Independent of Antibiotic Susceptibility.

Nathaniel S. Harshaw,Regis P. Kowalski,Kara M. Lehner,Eric G. Romanowski,Nicholas A. Stella,Robert M. Q. Shanks

doi:10.3390/antibiotics10091033

Abstract

The Rcs phosphorelay is a bacterial stress response system that responds to envelope stresses and in turn controls several virulence-associated pathways, including capsule, flagella, and toxin biosynthesis, of numerous bacterial species. The Rcs system also affects antibiotic tolerance, biofilm formation, and horizontal gene transfer. The Rcs system of the ocular bacterial pathogen Serratia marcescens was recently demonstrated to influence ocular pathogenesis in a rabbit model of keratitis, with Rcs-defective mutants causing greater pathology and Rcs-activated strains demonstrating reduced inflammation. The Rcs system is activated by a variety of insults, including β-lactam antibiotics and polymyxin B. In this study, we developed three luminescence-based transcriptional reporters for Rcs system activity and used them to test whether antibiotics used for empiric treatment of ocular infections influence Rcs system activity in a keratitis isolate of S. marcescens. These included antibiotics to which the bacteria were susceptible and resistant. Results indicate that cefazolin, ceftazidime, polymyxin B, and vancomycin activate the Rcs system to varying degrees in an RcsB-dependent manner, whereas ciprofloxacin and tobramycin activated the promoter fusions, but in an Rcs-independent manner. Although minimum inhibitory concentration (MIC) analysis demonstrated resistance of the test bacteria to polymyxin B and vancomycin, the Rcs system was activated by sub-inhibitory concentrations of these antibiotics. Together, these data indicate that a bacterial stress system that influences numerous pathogenic phenotypes and drug-tolerance is influenced by different classes of antibiotics despite the susceptibility status of the bacterium.

Highlights

Serratia marcescens is a Gram-negative pathogen from the order Enterobacterales that causes contact lens-associated keratitis in healthy patients [1,2,3] and a wide variety of nosocomial infections in the immune compromised, such as ventilator-associated pneumonia and sepsis in adults and neonates [4,5]
SMDB11_1637 is similar to osmotically inducible lipoprotein B, which is positively regulated by the Rcs system in Erwinia amylovora [24], E. coli [25,26], P. mirabilis [27], S. enterica [28], and Yersinia pseudotuberculosis [29]
Luminescence reporters for Rcs-stress system activation were generated for use in bacteria of the Enterobacterales order

Summary

Introduction

Serratia marcescens is a Gram-negative pathogen from the order Enterobacterales that causes contact lens-associated keratitis in healthy patients [1,2,3] and a wide variety of nosocomial infections in the immune compromised, such as ventilator-associated pneumonia and sepsis in adults and neonates [4,5]. S. marcescens isolates are typically resistant to antibiotics of the macrolide, tetracycline, β-lactam, and narrow spectrum cephalosporin classes due to expression of efflux pumps and β-lactamases [6]. They are generally susceptible to aminoglycoside, third generation cephalosporin, and fluoroquinolone antibiotics [6,7]. The Rcs stress response system has been found in bacteria from the Enterobacterales including, but not limited to, numerous pathogens, such as Escherichia coli, Klebsiella species, Proteus mirabilis, Salmonella enterica, and Yersinia pestis [8].

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