Comparative mutant prevention concentration and mechanism of resistance to veterinary fluoroquinolones in Staphylococcus pseudintermedius

Abstract

Background: The problem of antibacterial drug resistance is increasing worldwide, in part due to the therapeutic concentrations currently used based on the minimal inhibitory concentration (MIC) as a measure of potency are often the very concentrations required to selectively enrich the resistant mutant portion of the population. A mutant prevention concentration (MPC)-based dosing strategy is suggested to improve the therapeutic outcome based on the MIC.

Objective: Our aim was to investigate the MPC and mechanism of resistance to various fluoroquinolones using recent Staphylococcus pseudintermedius isolates from canine pyoderma.

Methods: The broth microdilution method for MIC and a series of agar plates containing different concentrations of fluoroquinolones were inoculated with ∼10(10) colony-forming units of the bacterial culture for MPC were used. PCR was used to identify mutation in the resistant isolates.

Results: The rank order of potency based on MIC and MPC was ciprofloxacin = enrofloxacin ≥ marbofloxacin > difloxacin ≥ orbifloxacin. Integrating our data with reported pharmacokinetic data at the recommended dose ranges revealed that only high doses of ciprofloxacin, enrofloxacin and marbofloxacin could achieve a maximal plasma concentration (C(max)) greater than the MPC of 90% of isolates (C(max)/MPC(90)). The overall rank of potency against S. pseudintermedius, based on C(max)/MIC, C(max)/MPC, the area under concentration-time curve (AUC)/MIC and AUC/MPC values, was in decreasing order: enrofloxacin > ciprofloxacin ≥ marbofloxacin ≥ orbifloxacin = difloxacin. Sequencing of the quinolone resistant determining region of gyrA, gyrB, grlA and grlB of resistant strains showed a base-pair substitution in both gyrA and gyrB that resulted in Ser-84 to Leu and Ser-80 to Arg amino acid changes, respectively.

Conclusions and clinical importance: High doses of ciprofloxacin, enrofloxacin and marbofloxacin could minimize the selection of resistant mutants, whereas the possibility of selecting mutants with the conventional doses of difloxacin and orbifloxacin, and low clinical doses of all fluoroquinolones, seems high.