The acquisition of full fluoroquinolone resistance in Salmonella Typhi by accumulation of point mutations in the topoisomerase targets

J Antimicrob Chemother. 2006 Oct;58(4):733-40. doi: 10.1093/jac/dkl333. Epub 2006 Aug 8.

Abstract

Objectives: To determine the contribution to fluoroquinolone resistance of point mutations in the gyrA and parC genes of Salmonella Typhi.

Methods: Point mutations that result in Ser-83-->Phe, Ser-83-->Tyr and Asp-87-->Asn amino acid substitutions in GyrA and Glu-84-->Lys in ParC were introduced into a quinolone-susceptible, attenuated strain of Salmonella Typhi using suicide vector technology. This is the first time that this approach has been used in Salmonella and abrogates the need for selection with quinolone antibacterials in the investigation of resistance mutations.

Results: A panel of mutants was created using this methodology and tested for quinolone resistance. The ParC substitution alone made no difference to quinolone susceptibility. Any single GyrA substitution resulted in resistance to nalidixic acid (MIC >or= 512 mg/L) and increased by up to 23-fold the MIC of the fluoroquinolones ofloxacin (MIC <or= 2 mg/L) ciprofloxacin (MIC <or= 1 mg/L) and gatifloxacin (MIC <or= 0.38 mg/L). Among the double substitution mutants, those with a substitution in ParC were less prone to killing with ciprofloxacin. The triple substitution mutants (Ser-83-->Phe or Tyr and Asp-87-->Asn in GyrA with Glu-84-->Lys in ParC) showed high levels of resistance to all the fluoroquinolones tested (MICs: gatifloxacin, 3-4 mg/L; ofloxacin, 32 mg/L; ciprofloxacin, 32-64 mg/L).

Conclusions: In Salmonella Typhi the fluoroquinolones tested act on GyrA and, at higher concentrations, on ParC. The point mutations conferred reduced susceptibility to ofloxacin and ciprofloxacin, and also reduced susceptibility to gatifloxacin. Three mutations conferred resistance to ofloxacin (32 mg/L), ciprofloxacin (32 mg/L) and to the more active fluoroquinolone gatifloxacin (MIC >or= 3 mg/L). These results predict that the use of ofloxacin or ciprofloxacin will select for resistance to gatifloxacin in nature.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Anti-Bacterial Agents / pharmacology*
  • Ciprofloxacin / pharmacology
  • DNA Gyrase / genetics
  • DNA Topoisomerase IV / genetics*
  • Drug Resistance, Bacterial / genetics*
  • Fluoroquinolones / pharmacology*
  • Humans
  • Microbial Sensitivity Tests
  • Nalidixic Acid / pharmacology
  • Point Mutation*
  • Salmonella typhi / drug effects*
  • Salmonella typhi / enzymology
  • Salmonella typhi / genetics
  • Salmonella typhi / growth & development

Substances

  • Anti-Bacterial Agents
  • Fluoroquinolones
  • Nalidixic Acid
  • Ciprofloxacin
  • DNA Topoisomerase IV
  • DNA Gyrase