Mutations in the gyrA gene of a highly fluoroquinolone-resistant clinical isolate of Escherichia coli

Abstract

We have determined the DNA sequence of the gyrA gene of the fluoroquinolone-resistant Escherichia coli isolate 205096 (MIC of ciprofloxacin, 128 micrograms/ml), which was recently demonstrated to be a gyrA mutant (P. Heisig and B. Wiedemann, Antimicrob. Agents Chemother. 35:2031-2036, 1991). Compared with the gyrA+ gene of E. coli K-12, 55 nucleotide changes were found. Three of these resulted in amino acid exchanges: Ser-83-->Leu, Asp-87-->Gly, and Asp-678-->Glu. A 0.7-kb DNA fragment containing two of these mutations (Ser-83-->Leu and Asp-87-->Gly) was isolated and fused in frame to the residual 3' coding region of gyrA+ in a plasmid to yield a chimeric gyrA gene (gyrA#). After introduction into E. coli 205096, this gyrA# gene does not increase the fluoroquinolone susceptibility of the resulting heterodiploid strain in a dominance test, while the gyrA+ gene does. The ciprofloxacin concentration necessary to inhibit by 90% (IC90) the supercoiling activity of gyrase isolated from E. coli 205096 is above 2,000 micrograms/ml. An identical result was found for gyrase reconstituted in vitro from the gyrB+ gene product and the chimeric gyrA# gene product. This is more than a 4,000-fold increase compared with the IC90 determined for gyrase from E. coli K-12 (gyrA+) (IC90, 0.5 microgram of ciprofloxacin per ml). No indications for the involvement of the gyrB gene or for alterations in quinolone permeation were found.

The Impact of Mutations in Topoisomerase Genes and the Plasmid-Mediated Quinolone Resistance (PMQR) Determinants on the Resistance to Fluoroquinolones in Klebsiella pneumoniae

“…The MIC of fluoroquinolones for isolates with two mutations in gyrA gene was higher than isolates with only one mutation that is similar to other studies' results (25)(26)(27)(28). Sequence analysis of QRDR in parC gene revealed two mutations with amino acid changes which have been reported by several previous studies (19,29).…”

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

“…The reasons were known to have another resistance mechanisms against fluoroquinolones, such as mutations in quinolone-resistance determining regions (QRDRs) of gyrA or efflux pumps. Alteration of the gyrA subunit of DNA gyrase has an important role in conferring high-level quinolone resistance in gram-negative bacteria such as E. coli and Neisseria gonorrhoeae (18). In E. coli, several mutations have been identified in the gyrA gene.…”

High-Level Ciprofloxacin Resistance from Point Mutations in gyrA and parC Confined to Global Hospital-Adapted Clonal Lineage CC17 of Enterococcus faecium

“…In these species, low-level ciprofloxacin resistance was reported in strains with mutations only in gyrA, while a higher level of resistance was discerned in strains carrying mutations in both gyrA and parC (1,19,30,41). Conversely, in gram-positive bacteria, such as Staphylococcus aureus and Streptococcus pneumoniae (16,52), parC appears to be the primary target, with mutations resulting in low-level resistance, which changes to high-level resistance with subsequent gyrA mutations.…”