title: Plasmid-mediated quinolone resistance determinants among Gram-negative bacteraemia isolates: a hidden threat.
abstract: OBJECTIVE
  The aim of the study was to investigate the prevalence of plasmid-mediated quinolone resistance (PMQR) genes in an unselected collection of bloodstream isolates recovered over an 18-month period in a laboratory affiliated to a university hospital in Athens, Greece, and to assess their impact on the in vitro activity of ciprofloxacin and levofloxacin.
  
  
  METHODS
  Eight PMQR genes were screened by PCR and sequencing. All PMQR-positive isolates were submitted to isoelectric focusing for β-lactamase detection, conjugation or transformation, time-kill assays, mutant prevention concentrationand inoculum effect evaluation. PCR and sequencing of gyrA and parC were performed for detection of chromosomal mutations.
  
  
  RESULTS
  Among 96 Gram-negative isolates, 7 (7.3 %) carried one or more PMQR genes. qnrS1 was the most prevalent (5.2 %), followed by aac(6')-Ib-cr (4.2 %) and their combination (2 %). Cloning was successful for three isolates. The presence of a single PMQR determinant without any target modification was not associated with quinolone resistance with one exception, Stenotrophomonasmaltophilia carrying qnrS1, which was resistant to norfloxacin and ciprofloxacin, but in this isolate, additional mechanisms of quinolone resistance cannot be excluded. All PMQR-positive isolates showed a significant inoculum effect. The mutant prevention concentrations of ciprofloxacin against the quinolone-susceptible clinical isolates ranged from 0.38 to 32 mg l-1 and those of levofloxacin from 1 to 32 mg l-1.
  
  
  CONCLUSIONS
  PMQRs compromised the bactericidal activity of ciprofloxacin and levofloxacin when expressed in Enterobactercloacae, S. maltophilia or Klebsiellapneumoniae and when more than one co-existed. PMQR determinants represent an unrecognized threat, capable to compromise the in vitro activity of quinolones if expressed in a favourable genetic environment and to favour selection of resistant mutants by widening the mutant selection window of these agents.
authors.affiliation: 4th Department of Internal Medicine, Infectious Diseases Laboratory, Molecular Biology Section, National and Kapodistrian University of Athens, School of Medicine, Athens, Greece.
authors.forename: Athanasios
authors.initials: A
authors.lastname: Margaritis
authors.affiliation: Present address: 4th Department of Internal Medicine, National and Kapodistrian University of Athens, School of Mecicine, Athens, Greece.|4th Department of Internal Medicine, Infectious Diseases Laboratory, Molecular Biology Section, National and Kapodistrian University of Athens, School of Medicine, Athens, Greece.
authors.forename: Irene
authors.initials: I
authors.lastname: Galani
authors.affiliation: 4th Department of Internal Medicine, Infectious Diseases Laboratory, Molecular Biology Section, National and Kapodistrian University of Athens, School of Medicine, Athens, Greece.
authors.forename: Marianthi
authors.initials: M
authors.lastname: Chatzikonstantinou
authors.affiliation: 4th Department of Internal Medicine, Infectious Diseases Laboratory, Molecular Biology Section, National and Kapodistrian University of Athens, School of Medicine, Athens, Greece.
authors.forename: George
authors.initials: G
authors.lastname: Petrikkos
authors.affiliation: 4th Department of Internal Medicine, Infectious Diseases Laboratory, Molecular Biology Section, National and Kapodistrian University of Athens, School of Medicine, Athens, Greece.|Present address: 4th Department of Internal Medicine, National and Kapodistrian University of Athens, School of Mecicine, Athens, Greece.
authors.forename: Maria
authors.initials: M
authors.lastname: Souli
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016470:Bacteremia / Q000382:microbiology*; D002939:Ciprofloxacin / Q000494:pharmacology; D027081:DNA Gyrase / Q000235:genetics; D027101:DNA Topoisomerase IV / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D004755:Enterobacteriaceae / Q000187:drug effects / Q000235:genetics*; D004756:Enterobacteriaceae Infections / Q000382:microbiology*; D004926:Escherichia coli / Q000235:genetics; D016905:Gram-Negative Bacterial Infections / Q000382:microbiology*; D006115:Greece; D006801:Humans; D064704:Levofloxacin / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D009154:Mutation; D015363:Quinolones / Q000494:pharmacology*; D011815:R Factors; D020615:Stenotrophomonas maltophilia / Q000187:drug effects / Q000235:genetics* / Q000276:immunology*; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D015363:Quinolones; D002939:Ciprofloxacin; D064704:Levofloxacin; D001618:beta-Lactamases; D027101:DNA Topoisomerase IV; D027081:DNA Gyrase