Impact ofgyrAandparCMutations on Quinolone Resistance, Doubling Time, and Supercoiling Degree ofEscherichia coli

Abstract

Isogenic mutants derived from quinolone-susceptible isolate WT by introducing gyrA (S83L, D87G) and parC (S80I, E84K) mutations associated with quinolone resistance were characterized with respect to quinolone resistance, growth rate, and degree of global supercoiling. The latter was determined by use of a pair of reporter plasmids carrying supercoiling-dependent promoters pgyrA and ptopA, respectively, transcriptionally fused to the reporter gene bla coding for TEM-1 ␤-lactamase. The quotient (Qsc) of the ␤-lactamase specific activity determined for a mutant carrying either plasmid was taken as a measure of the degree of global supercoiling. These Qsc data were comparable to results obtained from the separation of topoisomers of plasmid pBR322 on chloroquine-containing agarose gels and indicate a reduced degree of negative supercoiling in resistant mutants relative to the parent, WT. The S83L mutation in gyrA had the strongest influence on quinolone resistance while leaving other parameters nearly unaffected. The gyrA double mutation (S83L plus D87G) had an effect on quinolone resistance similar to that of a single mutation. Phenotypic expression of the parC mutation (S80I) was dependent on the presence of at least one gyrA mutation. Expression of high-level fluoroquinolone resistance (ciprofloxacin MIC, >4 g/ml) required a combination of the gyrA double mutation and one parC mutation (S80I or E84K). Such mutants showed considerable alterations of growth rate, global supercoiling, or both. Introduction of a parC mutation affected neither the doubling time nor the degree of supercoiling, while the presence of the gyrA D87G mutation was associated with a significant reduction in the degree of DNA supercoiling.

Selection of Antibiotic‐Resistant Bacterial Mutants: Allelic Diversity among Fluoroquinolone‐Resistant Mutations

“…High-level mutants were not detected at low drug concentrations with sample sizes of 10-30; thus, under these conditions, low-level resistance mutants were much more abundant than high-level ones. Although selection of resistant pathogens is likely to be more complex in patients than in the laboratory [30], our data collectively suggest that the recovery of many types of M. tuberculosis mutants from patients [4,13] reflects treatment with low-to-moderate concentrations of fluoroquinolone.…”

Characterisation of antimicrobial resistance-associated integrons and mismatch repair gene mutations in Salmonella serotypes

“…The results correlated well with their quinolone/fluoroquinolone resistance profiles. Salmonella Typhimurium ST20751, S. Heidelberg 22396 and S. Enteritidis 17929R and 17929N that were resistant to nalidixic acid and ciprofloxacin and/or enrofloxacin all had an amino acid substitution at codon 87 (Asp87Asn), which has been widely reported in previous studies [25].…”

Fluoroquinolone-resistantEscherichia coliisolated from healthy broilers with previous exposure to fluoroquinolones: Is there a link?

“…The molecular investigations into the underlying quinolone resistance mechanisms revealed that all quinolone-resistant isolates possessed the typical mutations in the topoisomerase genes, gyrA and parC , reported by other studies (39,40). Although quinolone resistance results mostly from chromosomal mutations, it may also be mediated by a plasmid-encoded qnr gene in members of the family Enterobacteriaceae (41).…”