A whole-genome screen identifies Salmonella enterica serovar Typhi genes involved in fluoroquinolone susceptibility.

A Keith Turner,Muhammud Yasir,John Wain,Julian Parkhill,Ian G Charles,Mark A Webber,Sabine E Eckert,Daniel J Turner

doi:10.1093/jac/dkaa204

Abstract

ObjectivesA whole-genome screen at sub-gene resolution was performed to identify candidate loci that contribute to enhanced or diminished ciprofloxacin susceptibility in Salmonella enterica serovar Typhi.MethodsA pool of over 1 million transposon insertion mutants of an S. Typhi Ty2 derivative were grown in a sub-MIC concentration of ciprofloxacin, or without ciprofloxacin. Transposon-directed insertion site sequencing (TraDIS) identified relative differences between the mutants that grew following the ciprofloxacin treatment compared with the untreated mutant pool, thereby indicating which mutations contribute to gain or loss of ciprofloxacin susceptibility.ResultsApproximately 88% of the S. Typhi strain’s 4895 annotated genes were assayed, and at least 116 were identified as contributing to gain or loss of ciprofloxacin susceptibility. Many of the identified genes are known to influence susceptibility to ciprofloxacin, thereby providing method validation. Genes were identified that were not known previously to be involved in susceptibility, and some of these had no previously known phenotype. Susceptibility to ciprofloxacin was enhanced by insertion mutations in genes coding for efflux, other surface-associated functions, DNA repair and expression regulation, including phoP, barA and marA. Insertion mutations that diminished susceptibility were predominantly in genes coding for surface polysaccharide biosynthesis and regulatory genes, including slyA, emrR, envZ and cpxR.ConclusionsA genomics approach has identified novel contributors to gain or loss of ciprofloxacin susceptibility in S. Typhi, expanding our understanding of the impact of fluoroquinolones on bacteria and of mechanisms that may contribute to resistance. The data also demonstrate the power of the TraDIS technology for antibacterial research.

Highlights

Salmonella enterica subsp. enterica serovar Typhi causes tens of millions of cases of typhoid fever, resulting in over 100 000 deaths annually.[1]
The emergence of MDR strains associated with the acquisition of plasmids[5] and the emergence of the H58 haplotype[6,7] and of XDR strains has led to typhoid fever that fails to respond to treatment with any of the antibiotics commonly used for treatment, including chloramphenicol, sulfamethoxazole/
The parent strain, WT26 pHCM1, possesses a GyrA Ser83Phe substitution conferring reduced susceptibility to fluoroquinolone antibiotics (MIC of ciprofloxacin 0.25 mg/L compared with 0.016 mg/L for the parent strain), and harbours the multiple antibiotic resistance plasmid pHCM1

Summary

Introduction

Salmonella enterica subsp. enterica serovar Typhi causes tens of millions of cases of typhoid fever, resulting in over 100 000 deaths annually.[1]. Enterica serovar Typhi causes tens of millions of cases of typhoid fever, resulting in over 100 000 deaths annually.[1] These are likely to be underestimates because of the predominance of typhoid fever in low- to middle-income countries where there is often a paucity of diagnostic facilities[2] capable of differentiating typhoid fever from other, clinically similar, febrile diseases.[3] Since the introduction of chloramphenicol for the treatment of typhoid, the spread of antibiotic-resistant S. Typhi strains has led to treatment failures.[4] The emergence of MDR strains associated with the acquisition of plasmids[5] and the emergence of the H58 haplotype[6,7] and of XDR strains has led to typhoid fever that fails to respond to treatment with any of the antibiotics commonly used for treatment, including chloramphenicol, sulfamethoxazole/.

Objectives

Methods

Results