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To evaluate the relationship between pharmacokinetic/pharmacodynamic (PK/PD) parameters and changes in susceptibility and resistance frequency of  Actinobacillus pleuropneumoniae  CVCC 259, a piglet tissue cage (TC) infection model was established. After  A. pleuropneumoniae  populations maintained at 10 8  CFU/mL in TCs, piglets were treated with various doses of danofloxacin once daily for 5 consecutive days by intramuscular injection. Both the concentrations of danofloxacin and the population of vial cells were determined. Changes in susceptibility and resistance frequency were monitored. Polymerase chain reaction (PCR) amplification of quinolone resistance-determining regions (QRDRs) and DNA sequencing were performed to identify point mutations in  gyrA ,  gyrB ,  parC , and  parE  genes. Furthermore, the susceptibility of mutants to danofloxacin and enrofloxacin was determined in the presence or absence of reserpine to assess whether the mutants were caused by efflux pumps. The MICs and resistant frequency of  A. pleuropneumoniae  both increased when danofloxacin concentrations fluctuated between MIC 99  (0.05 μg/mL) and MPC (mutant prevention concentration, 0.4 μg/mL). As for PK/PD parameters, the resistant mutants were selected and enriched when AUC 24h /MIC 99  ranged from 34.68 to 148.65 h or AUC 24h /MPC ranged from 4.33 to 18.58 h. Substitutions of Ser-83→Tyr or Ser-83→Phe in  gyrA  and Lys-53→Glu in  parC  were observed. The susceptibility of mutants obtained via danofloxacin treatment at 1.25 and 2.5 mg/kg were less affected by reserpine. These results demonstrate that maintaining the value of AUC 24h /MPC above 18.58 h may produce a desirable antibacterial effect and protect against  A. pleuropneumoniae  resistance to danofloxacin.

frontiers in microbiology

Pharmacokinetic/Pharmacodynamic Integration to Evaluate the Changes in Susceptibility of Actinobacillus pleuropneumoniae After Repeated Administration of Danofloxacin