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ABSTRACTQuinolone susceptibility was analyzed in 17 clinical isolates of Corynebacterium striatumand 9 strains of Corynebacterium amycolatumby the E-test method in Mueller-Hinton agar plates. The C. striatumATCC 6940 strain was used as a control strain. The amplified quinolone resistance determining regions of the gyrAgenes of C. amycolatumand C. striatumwere characterized. Four in vitro quinolone-resistant mutants of C. amycolatumwere selected and analyzed. Both in vivo and in vitro quinolone-resistant strains of C. amycolatumshowed high levels of fluoroquinolone resistance in strains with a double mutation leading to an amino acid change in positions 87 and 91 or positions 87 and 88 (unusual mutation) of GyrA, whereas the same concomitant mutations at amino acid positions 87 and 91 in GyrA of C. striatumproduced high levels of resistance to ciprofloxacin and levofloxacin but only showed a moderate increase in the MIC of moxifloxacin, suggesting that other mechanism(s) of quinolone resistance could be involved in moxifloxacin resistance in C. amycolatum. Moreover, a PCR-RFLP-NcoI of the gyrAgene was developed to distinguish between C. amycolatumand C. striatumspecies.

ABSTRACTAlthough quinolone resistance results mostly from chromosomal mutations, it may also be mediated by a plasmid-encoded qnrgene in members of the family Enterobacteriaceae. Thus, 297 nalidixic-acid resistant strains of 2,700 Escherichia colistrains that had been isolated at the Bicêtre Hospital (Le Kremlin-Bicêtre, France) in 2003 were screened for qnrby PCR. A single E. coliisolate that carried a ca. 180-kb conjugative plasmid encoding a qnrdeterminant was identified. It conferred low-level resistance to quinolones and was associated with a chromosomal mutation in subunit A of the topoisomerase II gene. The qnrgene was located on a sul1-type class 1 integron just downstream of a conserved region (CR) element (CR1) comprising the Orf513 recombinase. Promoter sequences for qnrexpression overlapped the extremity of CR1, indicating the role of CR1 in the expression of antibiotic resistance genes. This integron was different from other qnr-positive sul1-type integrons identified in American and Chinese enterobacterial isolates. In addition, plasmid pQR1 carried another class 1 integron that was identical to In53from E. coli. The latter integron possessed a series of gene cassettes, including those coding for the extended-spectrum β-lactamase VEB-1, the rifampin ADP ribosyltransferase ARR-2, and several aminoglycoside resistance markers. This is the first report of plasmid-mediated quinolone resistance in Europe associated with an unknown level of plasmid-mediated multidrug resistance in Enterobacteriaceae.

ABSTRACTThis study assessed the molecular epidemiology, resistance mechanisms, and susceptibility profiles of a collection of 150 extensively drug-resistant (XDR) Pseudomonas aeruginosaclinical isolates obtained from a 2015 Spanish multicenter study, with a particular focus on resistome analysis in relation to ceftolozane-tazobactam susceptibility. Broth microdilution MICs revealed that nearly all (>95%) of the isolates were nonsusceptible to piperacillin-tazobactam, ceftazidime, cefepime, aztreonam, imipenem, meropenem, and ciprofloxacin. Most of them were also resistant to tobramycin (77%), whereas nonsusceptibility rates were lower for ceftolozane-tazobactam (31%), amikacin (7%), and colistin (2%). Pulsed-field gel electrophoresis–multilocus sequence typing (PFGE-MLST) analysis revealed that nearly all of the isolates belonged to previously described high-risk clones. Sequence type 175 (ST175) was detected in all 9 participating hospitals and accounted for 68% (n= 101) of the XDR isolates, distantly followed by ST244 (n= 16), ST253 (n= 12), ST235 (n= 8), and ST111 (n= 2), which were detected only in 1 to 2 hospitals. Through phenotypic and molecular methods, the presence of horizontally acquired carbapenemases was detected in 21% of the isolates, mostly VIM (17%) and GES enzymes (4%). At least two representative isolates from each clone and hospital (n= 44) were fully sequenced on an Illumina MiSeq. Classical mutational mechanisms, such as those leading to the overexpression of the β-lactamase AmpC or efflux pumps, OprD inactivation, and/or quinolone resistance-determining regions (QRDR) mutations, were confirmed in most isolates and correlated well with the resistance phenotypes in the absence of horizontally acquired determinants. Ceftolozane-tazobactam resistance was not detected in carbapenemase-negative isolates, in agreement with sequencing data showing the absence of ampCmutations. The unique set of mutations responsible for the XDR phenotype of ST175 clone documented 7 years earlier were found to be conserved, denoting the long-term persistence of this specific XDR lineage in Spanish hospitals. Finally, other potentially relevant mutations were evidenced, including those in penicillin-binding protein 3 (PBP3), which is involved in β-lactam (including ceftolozane-tazobactam) resistance, and FusA1, which is linked to aminoglycoside resistance.