title: Effect of the efflux pump QepA2 combined with chromosomally mediated mechanisms on quinolone resistance and bacterial fitness in Escherichia coli.
abstract: OBJECTIVE
  The aim of the study was to determine the interplay between the plasmid-mediated qepA2 gene and multiple chromosomally mediated fluoroquinolone resistance determinants in the development of fluoroquinolone resistance in Escherichia coli and its influence on bacterial fitness.
  
  
  METHODS
  E. coli ATCC 25922 and derived isogenic strains harbouring different chromosomally mediated fluoroquinolone resistance determinants were electroporated with pBK-CMV vector encoding QepA2. The MICs of fluoroquinolones were determined by standardized microdilution. The mutant prevention concentration (MPC) was evaluated. Bacterial fitness was analysed using ΔlacZ system competition assays.
  
  
  RESULTS
  The ciprofloxacin MIC for strains harbouring the qepA2 gene was 4- to 8-fold higher compared with strains without the qepA2 gene. The qepA2 gene also increased the MPC of ciprofloxacin 4- to 16-fold. Combination of the qepA2 gene plus two to three additional mechanisms conferred a clinically relevant resistance level. The presence of the qepA2 gene was associated with fitness costs in strains with mutations in the gyrA and/or parC genes, although the presence of an additional deletion of the marR gene compensated for this fitness cost by increasing bacterial fitness by 5%-23%.
  
  
  CONCLUSIONS
  The additive effect of chromosomally mediated fluoroquinolone resistance mechanisms and the qepA2 gene led to clinical levels of fluoroquinolone resistance. Under competitive conditions, the qepA2 gene had a biological cost in E. coli that was compensated for by the presence of an additional deletion in the marR gene.
authors.affiliation: Unidad de Enfermedades Infecciosas y Microbiología Clínica, Hospital Universitario Virgen Macarena, Sevilla, Spain jesmacbar@hotmail.com.
authors.forename: Jesús
authors.initials: J
authors.lastname: Machuca
authors.affiliation: Departamento de Microbiología, Universidad de Sevilla, Sevilla, Spain Spanish Network for Research in Infectious Diseases (REIPI RD12/0015), Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Alejandra
authors.initials: A
authors.lastname: Briales
authors.affiliation: Departamento de Microbiología, Universidad de Sevilla, Sevilla, Spain.
authors.forename: Paula
authors.initials: P
authors.lastname: Díaz-de-Alba
authors.affiliation: Spanish Network for Research in Infectious Diseases (REIPI RD12/0015), Instituto de Salud Carlos III, Madrid, Spain Hospital Universitario Marques de Valdecilla and Valdecilla Biomedical Research Institute (IDIVAL), Santander, Spain Departamento de Biología Molecular, Universidad de Cantabria, Santander, Spain.
authors.forename: Luis
authors.initials: L
authors.lastname: Martínez-Martínez
authors.affiliation: Unidad de Enfermedades Infecciosas y Microbiología Clínica, Hospital Universitario Virgen Macarena, Sevilla, Spain Departamento de Microbiología, Universidad de Sevilla, Sevilla, Spain Spanish Network for Research in Infectious Diseases (REIPI RD12/0015), Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Álvaro
authors.initials: Á
authors.lastname: Pascual
authors.affiliation: Departamento de Microbiología, Universidad de Sevilla, Sevilla, Spain Spanish Network for Research in Infectious Diseases (REIPI RD12/0015), Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: José-Manuel
authors.initials: JM
authors.lastname: Rodríguez-Martínez
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002876:Chromosomes, Bacterial; D024881:Drug Resistance, Bacterial; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics* / Q000502:physiology; D024841:Fluoroquinolones / Q000494:pharmacology*; D005798:Genes, Bacterial; D008826:Microbial Sensitivity Tests; D010957:Plasmids; D014774:Virulence
chemical_list: D000900:Anti-Bacterial Agents; D024841:Fluoroquinolones
keywords: E. coli; qepA; resistance mechanisms