In Vivo Pharmacokinetics/Pharmacodynamics Modeling Of Ceftiofur Against Mouse Pyoderma Due To Pseudointermediate Staphylococcus

Pyoderma caused by Staphylococcus pseudointermedia reported in pets is increasing.As a zoonotic pathogen,it poses a serious threat to animal health and public health safety.Ceftiofur,a third-generation of cephalosporin antibiotic for animals use only,shows excellent pharmacokinetic characteristics and broad-spectrum antibacterial activity.It is the first-line drug for the treatment of pyoderma caused by Staphylococcus pseudointermedia.The dosing regimen based on PK/PD research is the basis for rational drug use and the control of drug resistance.But there are few studies on PK/PD synchronization model of ceftiofur against Staphylococcus pseudointermediate.The purpose of this study is to establish the in vivo PK/PD model of ceftiofur in mice infected with Staphylococcus pseudointermedia.1.Isolation and identification of Staphylococcus pseudointermedia and establishment of mouse skin infection model cuased by Staphylococcus pseudointermediaTwelve strains of Staphylococcus pseudointermedia were isolated from 19 dogs suffering from pyoderma.It accounts for 55%of all bacteria.Including the 15 strains of Staphylococcus pseudointermediate preserved in the laboratory before,there are 27 strains in total.The sensitivity of enrofloxacin,danofloxacin,streptomycin,ceftiofur,florfenicol,vancomycin and amoxicillin,against 27 strains of Staphylococcus pseudointermediate were determined,the results showed that Staphylococcus pseudointermediate was resistant to most drugs except vancomycin,but the resistance rate to ceftiofur was only 4%.Therefore,ceftiofur was selected as the required drug for subsequent experiments.The MIC of 171107D1 strain was MIC50 and was used for the follow-up researchs.The skin at back area of 8-week-old Balb/c mice was injected subcutaneously with 1×106 CFU bacterial solution.After 24 hours of infection,the inoculation site appeared red and swollen,and the bacterial load of the infected skin tissue fluctuated relatively smoothly within 3 days.The bacteria loading was maintained at about 107 CFU.It can be used for the follow-up experiments.2.The pharmacokinetic study of ceftiofur in mice infected with Staphylococcus pseudointermediate150 eight-week-old infected mice were randomly divided into 3 groups.After intramuscular injection of 60,20,1 mg/kg b.w.ceftiofur solution,0.7 mL blood samples were collected from the orbit at 0,0.167,0.5,1,2,4,6,8,12,and 24 h.The concentration of ceftiofur in plasma was determined by high performance liquid chromatography(HPLC),and the pharmacokinetic parameters of ceftiofur in infected mice were determined by noncompartmental model fitting.The results showed that after intramuscular injection of 1,20,60 mg/kg b.w.ceftiofur sodium in mice,the peak time of the low,medium and high doses(0.6±0.224 h,0.433±0.149 h,0.367±0.182 h)were similar.The values of T1/2β(14.641±5.214 h,14.461±5.936 h,15.563±6.783 h)are similar too.The value of Cmax were 0.726±0.238 μg/mL,2.421 ±0.534μg/mL,4.403±1.608 μg/mL,respectively;The value of AUC0～24 h were 3.530±0.333 h·μg/mL,15.572±3.525 h·μg/mL,19.630±1.255 h·μg/mL,respectively;The value of AUC0～12 h were 3.533±0.333 h·μg/mL,9.403±1.966 h·μg/mL,11.697±1.934 h·μg/mL,respectively;The value of AUC0～8 h were 2.916±0.245 h·μg/mL,6.735±1.455 h·μg/mL,8.869±2.577 h·μg/mL,respectively.In the dose range of ceftiofur from 1 to 60 mg/kg,AUC0～24 h,AUC0～12 h,AUC0～8 h,Cmax have a linear relationship with the dose.3.Pharmacodynamics and PK/PD model of ceftiofur against Staphylococcus pseudointermediateThe MIC of Ceftiofur against Staphylococcus pseudointermediate was 0.25μg/ml,the MBC was 1μg/ml,and the MBC was 4 times the MIC.It can be seen from the static sterilization curve that when the initial bacterial count is 106 CFU/mL,the antibacterial effect can be achieved when the drug concentration reaches the MIC;the bactericidal effect can be achieved when the drug concentration reaches 2 × MIC and above.162 8-week-old infected mice were randomly divided into 27 groups,and 60,40,30,20,15,10,5,1,and 0 mg/kg b.w.ceftiofur with a dosing interval of 8,12,24 h were injected intramuscularly respectively,for 5 consecutive days.The mice were sacrificed 24 hours after the last administration,and the bacterial load of Staphylococcus pseudointermediate in the skin tissue was measured.The in vivo efficacy results showed that the bacteria in the blank control group stabilized at 106 CFU/mouse after 5 days.With the increase of the dose,the bacterial load decreased.When the drug increased to 10 mg/kg body weight,the bacteria loading could be reduced by 1× Log10 CFU/mouse.When the drug concentration reaches 30 mg/kg body weight or more,the bacterial load is not reduced significantly.Changing the dosing interval has little influence on the antibacterial effect of the drug.The Inhibitory Emax model was used to fit the correlation between PK/PD parameters and in vivo antibacterial effects.The results showed that the AUC0～24 h/MIC parameter of ceftiofur has a better fit with the antibacterial effect of the drug in vivo.When the AUC0～24h/MIC are 35.118 and 69.330 respectively,it can reduce Staphylococcus pseudointermediates by 1 × Log 10 CFU/mouse and 2 × Log 10 CFU/mouse respectively.