Detection of plasmid-mediated quinolone resistance genes in clinical isolates of Enterobacter spp. in Spain

Abstract

We have studied by PCR and DNA sequencing the presence of the qnrA, qnrB, qnrS, aac(6')-Ib-cr, qepA, intI1, and ISCR1 genes in 200 clinical isolates of Enterobacter cloacae (n = 153) and E. aerogenes (n = 47) consecutively collected between January 2004 and October 2005 in two hospitals located in Santander (northern Spain) and Seville (southern Spain). Mutations in the quinolone resistance-determining region of gyrA and parC also were investigated in organisms containing plasmid-mediated quinolone resistance genes. The isolates had different resistant phenotypes, including AmpC hyperproduction, extended-spectrum beta-lactamase production, resistance or decreased susceptibility to quinolones, and/or resistance to aminoglycosides. Among the 116 E. cloacae isolates from Santander, qnrS1, qnrB5, qnrB2, and aac(6')-Ib-cr were detected in 22 (19%), 1 (0.9%), 1 (0.9%), and 3 (2.6%) isolates, respectively. Twenty-one, 17, and 2 qnrS1-positive isolates also contained bla(LAP-1), intI1, and ISCR1, respectively. A qnrB7-like gene was detected in one E. aerogenes isolate from Santander. No plasmid-mediated quinolone resistance gene was detected in the isolates from Seville. The qnrS1-containing isolates corresponded to four pulsed-field gel electrophoresis patterns and showed various levels of resistance to quinolones. Six isolates were susceptible to nalidixic acid and presented reduced susceptibility to ciprofloxacin. The qnrS1 gene was contained in a conjugative plasmid of ca. 110 kb, and when the plasmid was transferred to recipient strains that did not have a specific mechanism of quinolone resistance, the ciprofloxacin MICs ranged from 0.047 to 0.125 microg/ml.

FIG. 1.

Analysis of plasmid DNA from E. cloacae parental isolates and derived transconjugants by pulsed-field agarose gel electrophoresis (A) and Southern blotting (B). Lanes: 1, molecular marker; 2, E. cloacae 248; 3, E. cloacae 3138; 4, qnrS1-positive transconjugant from E. cloacae 3138; 5, E. cloacae 3164; 6, E. cloacae 2531; 7, transconjugant from E. cloacae 2531 lacking qnrS1; 8, qnrS1-positive transconjugant from E. cloacae 2531; 9, E. cloacae 3249; 10, E. cloacae 3049; 11, E. cloacae 3210; 12, transconjugant from E. cloacae 3210; 13, E. cloacae 1640; 14, E. cloacae 3714.