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Non-target gene mutations in the development of fluoroquinolone resistance in Escherichia coli
W V Kern1, M Oethinger, A S Jellen-Ritter
1Section of Infectious Diseases and Clinical Immunology, Department of Medicine, University Hospital and Medical Center, D-89070 Ulm, Germany.winfried.kern@medizin.uni-ulm.de
Abstract:
Mutations in loci other than genes for the target topoisomerases of fluoroquinolones, gyrA and parC, may play a role in the development of fluoroquinolone resistance in Escherichia coli. A series of mutants with increasing resistance to ofloxacin was obtained from an E. coli K-12 strain and five clinical isolates. First-step mutants acquired a gyrA mutation. Second-step mutants reproducibly acquired a phenotype of multiple antibiotic resistance (Mar) and organic solvent tolerance and showed enhanced fluoroquinolone efflux. None of the second-step mutants showed additional topoisomerase mutations. All second-step mutants showed constitutive expression of marA and/or overexpressed soxS. In some third-step mutants, fluoroquinolone efflux was further enhanced compared to that for second-step mutants, even when the mutant had acquired additional topoisomerase mutations. Attempts to circumvent the second-step Mar mutation by induction of the mar locus with sodium salicylate and thus to select for pure topoisomerase mutants at the second step were not successful. At least in vitro, non-target gene mutations accumulate in second- and third-step mutants upon exposure to a fluoroquinolone and typically include, but do not appear to be limited to, mutations in the mar or sox regulons with consequent increased drug efflux.
Insights
Mutations outside of target genes contribute to fluoroquinolone resistance in E. coli. Non-target gene mutations, like those in the mar or sox regulons, increase drug efflux and antibiotic resistance.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Fluoroquinolone resistance in Escherichia coli is primarily associated with mutations in gyrA and parC genes.
- However, other genetic factors may contribute to the development of resistance.
Purpose of the Study:
- To investigate the role of non-target gene mutations in fluoroquinolone resistance in E. coli.
- To characterize the genetic and phenotypic changes associated with increasing fluoroquinolone resistance.
Main Methods:
- Generation of ofloxacin-resistant mutants from E. coli K-12 and clinical isolates.
- Analysis of topoisomerase mutations, multiple antibiotic resistance (Mar) phenotype, organic solvent tolerance, and gene expression (marA, soxS).
- Assessment of fluoroquinolone efflux in resistant mutants.
Main Results:
- First-step mutants acquired gyrA mutations.
- Second-step mutants exhibited a multiple antibiotic resistance (Mar) phenotype, organic solvent tolerance, enhanced fluoroquinolone efflux, and constitutive expression of marA and/or soxS, without additional topoisomerase mutations.
- Third-step mutants showed further enhanced efflux, sometimes with additional topoisomerase mutations.
- Attempts to select for pure topoisomerase mutants by inducing the mar locus were unsuccessful.
Conclusions:
- Non-target gene mutations, particularly in the mar and sox regulons, significantly contribute to fluoroquinolone resistance in E. coli by increasing drug efflux.
- These mutations can accumulate independently of or in conjunction with target topoisomerase mutations, leading to complex resistance mechanisms.