Pharmacokinetic/Pharmacodynamic(PK/PD) Profiles Of Tulathromycin Against Haemophilus Parasuis In Experimental Intraperitoneal Infection Models

Tulathromycin,a semi-synthetic macrolide,showed a satisfactory pharmacokinetic feature in swine and an ideal external used drug in treating infection of Haemophilus parasuis(H.parasuis).However,few PK/PD model studies have reported about tulathromycin against H.parasuis.Therefore,the objective of this study was to characterize the PK/PD profiles of H.parasuis using a PK/PD model.MICs of tulathromycin aginst 94 H.parasuis strains and two standard strains of H.parasuis,serotype 5,13,were detected.The results showed that MIC90 of tulathromycin aginst 94 H.parasuis strains(2014?2015)was 0.5 ?g/mL,and MIC of the two standard strains of H.parasuis,serotype 5,13,were 4 ?g/mL,0.5 ?g/mL,respectively.And thus,H.parasuis,serotype 13(13R)was used in PK/PD model.An neutropenic guinea pig model was established by intraperitoneal infection cyclophosphamide.The results of blood index,which including white blood cell,lymphocyte,neutrophils,and serum physiological and biochemical index,which including AST,ALT,UREA,showed that 100 mg/kg b.w.,once a day,for 3 days,could make the guinea pig reach the state of immunosuppression.Through the mental state,morbidity and mortality of guinea pigs,indicated that it is optimal to apply a single inoculum of a 0.2 mL aliquot solution containing approximately 109 CFU/mL of the H.parasuis strain,which morbidity was 100%,and mortality was 20%,in an in vivo H.parasuis infection model.The infected,neutropenic guinea pigs were allocated to three groups and treated with tulathromycin at single IM doses of 1,10,or 20 mg/kg.Tulathromycin concentrations in serum and lung from different times were determined via a UPLC-MS/MS method.The PK profiles of tulathromycin were analyzed by the noncompartmental model using WinNonlin software.The results showed that,for health groups administration with 10 mg/kg,in serum,Tmax was 0.3 ± 0.1 h,Cmax was 1.99 ± 0.56 ?g/mL,T1/2? was 24.2 h,AUC168h was 40.68?g·h/mL.In lung tissue,Tmax was 0.5 ± 0.2 h,Cmax was 5.11 ± 2.55 ?g/g,T1/2? was 41.3 h,AUC168h was 125.94 ?g·h/g.For infected groups administration with 10 mg/kg,in serum,Tmax was 0.6 ± 0.2 h,Cmax was 3.61 ± 0.45 ?g/mL,T1/2? was 26.9 h,AUC168h was 57.18?g·h/mL.In lung tissue,Tmax was 0.5 ± 0.2 h,Cmax was 5.98 ± 1.91 ?g/g,T1/2? was 72.1 h,AUC168h was 262.78 ?g·h/g.These findings indicated that compared with the state of infection and health,the Tmax and T1/2? of tulathromycin were prolonged,but there was no significant difference(P>0.05),and AUC increased significantly(P<0.05).In addition,a linear relationship was found between three levels of dose(1,10,20 mg/kg)in guinea pig.Of the serum samples,significant correlations between dose treatment and AUC0-168h(R2 =0.9829)or Cmax(R2 =0.9889)were observed.Similarly for lung samples,significant correlations between dose treatment and AUC0-168h(R2 =0.9806)or Cmax(R2 =0.9986)were observed.PK/PD profiles of tulathromycin against H.parasuis in ex vivo were investigated in an experimental intraperitoneal H.parasuis infection model in neutropenic guinea pigs.All serum samples obtained from guinea pigs(0-168 h)after IM injection of tulathromycin were used to establish ex-vivo time-killing curves.The MIC of tulathromycin against H.parasuis 13R was determined in both CAMHB cultures and serum,which indicated that tulathromycin may have a large effect in serum.Minimal bactericidal concentration(MBC),mutant prevention concentration(MPC),post-antibiotic effects(PAEs)were also detected.Selection index(SI),MPC/MIC,indicated that the mutant selection windows(MSW)of tulathromycin against H.parasuis was narrow,which could inhibition of the selective mutation of H.parasuis to produce drug resistance.PAE of different concentrations of tulathromycin and exposure times showed tulathromycin had a long PAE.The in vitro PK/PD relationship of tulathromycin was described using a sigmoid inhibitory Emax model and integration of PK/PD for dose optimization of tulathromycin based on PK/PD indexes and MIC in serum,and an estimated dose range between 2.1 and 2.4 mg/kg would achieve the virtual eradication of H.parasuis.PK/PD profiles of tulathromycin against H.parasuis in vivo were investigated,and the AUC0-168h for any dose level from 1-20 mg/kg based on the linear relationship between our treatment levels of dose(1,10,20 mg/kg).Based on AUC0-168h for different doses from 1?20 mg/kg and MIC in serum,using a sigmoid inhibitory Emax model,the values of AUC0-168h/MIC serum for H.parasuis-3 log10 CFU/mL reduction and-4 logio CFU/mL reduction were 728.47 and 916.90,respectively.And,the values of AUC0-168h/MIC of lung tissue for H.parasuis-3 logio CFU/mL reduction and-4 log10 CFU/mL reduction were 2126.44 and 3462.62,respectively.The single dosage values to achieve a reduction of-4 logio CFU/lung were calculated from the ratios of AUC0-168h/MIC and were 4.4 mg/kg for serum and 5.0 mg/kg for lung tissue.According with a dose conversion coefficient(0.296),a converted dose of 1.3 mg/kg(serum)and 1.5 mg/kg(lung)were calculated for pig.The effectiveness of tulathromycin against H.parasuis were the first time evaluated using a in vitro or in vivo PK/PD model by a laboratory animal model.The drug concentr-ations in plasma and lung tissue were concurrently detected for the in vivo PK/PD model,used for predicting dose regimen of tulathromycin based on lung.The dose regimen were validated in clinical practice on swine,infected by the standard strain of H.parasuis 13R,and a clinical isolate(serotype was 13 and MIC was equal to 13R),which indicated the marked dose of tulathromycin may be optimal to treat H.parasuis infection.These investigations provide a solid foundation for the usage of tulathromycin in pigs for H.parasuis infections,and it provides a model reference for other macrolide antibiotics,with longer half-life or targeted drugs.