title: Evaluation of the contribution of gyrA mutation and efflux pumps to fluoroquinolone and multidrug resistance in pathogenic Escherichia coli isolates from dogs and cats.
abstract: OBJECTIVE
  To investigate the contribution of gyrA mutation and efflux pumps to fluoroquinolone resistance and multidrug resistance among Escherichia coli isolates from dogs and cats.
  
  
  METHODS
  536 clinical isolates of E coli.
  
  
  METHODS
  Minimum inhibitory concentrations (MICs) were determined for enrofloxacin and 6 other drug classes by use of broth microdilution techniques. Real-time PCR assay was used to determine the mutation in gyrA; Phe-Arg-β-naphthylamide, an efflux pump inhibitor, was used to examine the contribution of efflux pump overexpression.
  
  
  RESULTS
  The MIC for fluoroquinolones increased in a stepwise fashion and was lowest in the absence of mutations, higher with a single point mutation, and highest with 2 point mutations. Level of resistance in the latter category was high (8 times the breakpoint), but this was associated with expression of the AcrAB efflux pump. Inhibition of the efflux pump resulted in a reduction in the MIC to less than the susceptible breakpoint for isolates with an MIC ≤ 4 mg/L, regardless of the presence of a mutation. The greatest magnitude in MIC decrease (MIC was decreased by a factor of > 67 fold) was for isolates with a single mutation but the greatest absolute decrease in MIC (124 mg/L) was for isolates with 2 mutations. Inhibition of the AcrAB efflux pump in isolates characterized by multidrug resistance decreased the MIC of drugs structurally unrelated to fluoroquinolone.
  
  
  CONCLUSIONS
  Fluoroquinolone resistance in E coli appeared to be a stepwise phenomenon, with MIC increasing as the number of point mutations in gyrA increased, but high-level resistance and multidrug resistance associated with fluoroquinolone resistance reflected overexpression of the AcrAB efflux pump.
authors.affiliation: Department of Anatomy, Physiology and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849, USA.
authors.forename: Bashar W
authors.initials: BW
authors.lastname: Shaheen
authors.forename: Dawn M
authors.initials: DM
authors.lastname: Boothe
authors.forename: Omar A
authors.initials: OA
authors.lastname: Oyarzabal
authors.forename: Chengming
authors.initials: C
authors.lastname: Wang
authors.forename: Calvin M
authors.initials: CM
authors.lastname: Johnson
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002352:Carrier Proteins / Q000235:genetics*; D002371:Cat Diseases / Q000382:microbiology; D002415:Cats; D027081:DNA Gyrase / Q000235:genetics* / Q000378:metabolism; D004283:Dog Diseases / Q000382:microbiology; D004285:Dogs; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics*; D004927:Escherichia coli Infections / Q000382:microbiology / Q000662:veterinary; D024841:Fluoroquinolones / Q000494:pharmacology*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D008826:Microbial Sensitivity Tests
chemical_list: D000900:Anti-Bacterial Agents; D002352:Carrier Proteins; D024841:Fluoroquinolones; D027081:DNA Gyrase