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Journal of Clinical Microbiology, September 2009, p. 2751-2758, Vol. 47, No. 9
0095-1137/09/$08.00+0 doi:10.1128/JCM.00456-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Evaluation of Quinolones for Use in Detection of Determinants of Acquired Quinolone Resistance, Including the New Transmissible Resistance Mechanisms qnrA, qnrB, qnrS, and aac(6')Ib-cr, in Escherichia coli and Salmonella enterica and Determinations of Wild-Type Distributions 
Research Group for Antimicrobial Resistance and Molecular Epidemiology, Department for Microbiology and Risk Assessment, National Food Institute, Technical University of Denmark, Bülowsvej 27, Copenhagen V DK-1790, Denmark
Received 4 March 2009/ Returned for modification 2 May 2009/ Accepted 25 June 2009
Fluoroquinolone resistance in members of the Enterobacteriaceae family is mostly due to mutations in the quinolone resistance-determining regions of the topoisomerase genes. However, transferable genes encoding quinolone resistance have recently been described. The current methods for susceptibility testing are not adapted to the detection of new resistance determinants, which confer low levels of resistance. The aim of this study was to compare the ability of the screening of the different quinolones by disk diffusion assays and MIC determinations to detect fluoroquinolone resistance. Sixty-nine Escherichia coli strains and 62 Salmonella strains, including strains fully susceptible to quinolones, nalidixic acid-resistant strains, strains with resistance to fluoroquinolones (resistant to nalidixic acid), and strains showing low-level resistance to fluoroquinolones conferred by transferable quinolone resistance genes, including qnrA, qnrB, qnrS, and aac(6')Ib-cr, were selected. Disk diffusion assays and MIC determinations by the agar dilution method were performed, according to CLSI standards, with nalidixic acid, flumequine, oxolinic acid, ciprofloxacin, enrofloxacin, marbofloxacin, norfloxacin, ofloxacin, and levofloxacin. The MIC of levofloxacin was determined by an Etest. The results showed a trimodal distribution of the MICs for both E. coli and Salmonella. The MIC distributions for the isolates varied with the compounds tested. Screening for nalidixic acid resistance by MIC testing or disk diffusion assay was not efficient for the detection of some of the isolates carrying qnr and aac(6')Ib-cr. Transferable resistance genes would best be detected by testing for the MIC of ciprofloxacin or norfloxacin, as testing for the MICs of the other compounds would fail to detect isolates carrying aac(6')Ib-cr because the enzyme produced is able to reduce the activities of these two compounds only due to their chemical structures. In conclusion, screening with nalidixic acid is efficient for the detection of mutants, but it is not so efficient for the detection of qnr and aac(6')Ib-cr. Detection would be maximized by screening with either ciprofloxacin or norfloxacin by both MIC determination and disk diffusion assays. Furthermore, a low concentration of ciprofloxacin (1 µg) in the disks seemed to increase the sensitivity of the disk diffusion assay.
* Corresponding author. Mailing address: Research Group for Antimicrobial Resistance and Molecular Epidemiology, Department for Microbiology and Risk Assessment, National Food Institute, Technical University of Denmark, Bülowsvej 27, Copenhagen V DK-1790, Denmark. Phone: 45 72 34 62 69. Fax: 45 72 34 63 41. E-mail: licav{at}food.dtu.dk
Published ahead of print on 1 July 2009.
Journal of Clinical Microbiology, September 2009, p. 2751-2758, Vol. 47, No. 9
0095-1137/09/$08.00+0 doi:10.1128/JCM.00456-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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