Pharmacokinetics Of Robenidine Hydrochloride In Carassius Auratus Gibelio And Its Immune Effect To Carassius Auratus Gibelio

ROBH was invented as an anti-coccidiosis agent to chicken and rabbit in1980s,it is an artificial synthesized drug.The mechanism of ROBH is affect metabolism of proteins,so that the coccidiosis was inhibited.The drug was listed as a standard fish drug to cure myxosporidiasis of fish in 2010.To date,the effects of ROBH to fish is less clear.What’s more,during the drug using whether the other diseases besides myxosporidiasis is affected that also unknow.This study was based on silver crucian carp as experiment subject,has investigated the pharmacokinetics of ROBH in the carp,and the immune function effects on the fish life activity of the drug and its inhibitory effects of virus disease and bacterial diseases.1.The pharmacokinetics of robenidine hydrochloride in Carassius auratus gibelioIn order to provide theoretical guidance for clinical rational administration in the breeding of silver crucian carp,pharmacokinetics of ROBH in the carp with body weight of（120±10）g was studied following oral administration of different doses（20mg/kg,30mg/kg and 40mg/kg）at water temperature of（25±1）℃.After administration,concentration-time curves of ROBH in tested tissues of silver crucian carp exhibited a“two-peak”pattern.The peak time,the maximum concentration and the area under the concentration-time curve of three doses in blood were as follows:20mg/kg,Tmax=2.3h,Cmax=0.55μg/kg,AUC=9.35μg·mL^-1·h^-1;30mg/kg,Tmax=2.1h,Cmax=0.73mg/kg,AUC=13.02μg·mL^-1·h^-1;40mg/kg,Tmax=2.2h,Cmax=1.17mg/kg,AUC=13.98μg·mL^-1·h^-1.Area under the concentration-time curve value from higher to lower in tested tissues were:kidney,liver,blood and muscle.The lowest rate of elimination was found in muscle,which was followed by blood.The fact that high ROBH concentration and long MRT were identified in kidney and liver suggested some kind of accumulative effect in both of these two tissues.According to the pharmacokinetics of ROBH in muscle,we suggest a withdrawal period of no less than 9 days in silver crucian carp.What’s more,we suggest the withdrawal period can be adjusted properly because it might be influenced significantly by administration dose and temperature.2.The effects of robenidine hydrochloride to cyprinid herpesvirus 2（CyHV-2）and Aeromonas hydrophila.In order to study robenidine hydrochloride prevents silver crucian carp from bacteria and virus infecting,body weight of the fish（body weight,120±10 g）were studied following oral administration with the dose of20mg/kg at water temperature（25±1）℃.After 24h,these fish were infected with Aeromonas hydrophila（AH10）at the concentration of 10⁵,10⁶,10⁷,10⁸,10⁹CFU/mL following intraperitoneal injection.The lethal concentration(LD₅₀)of AH10 was 6.98×10⁷ CFU/mL with ROBH oral administration to silver crucian carp was obtained via Bliss way.However,the LD₅₀ of the same concentration of AH10 to fish without drug used was 9.49×10⁶CFU/mL.At the same experimental condition,the death rate of silver crucian carp following 40 mg/kg,20 mg/kg and 0 mg/kg ROBH oral administration maintained 24h and subsequently infected with CyHV-2 was investigated.The mortality of these carp was observed maintain 7 days.The results showed that silver crucian carp dead at 3 days and the death numbers was higher at 4 days,and all of fish dead at 6 days except the control group,which were infected with CyHV-2.According to the results,ROBH can decrease the death rate of bacteria infected fish but have no inhibition effect to virus diseases.Thus,ROBH is suggested to cure the myxosporidiasis of silver crucian carp while also can inhibit bacterial diseases.3.Transcriptome analysis of immune effects of Carassius auratus gibelio after robenidine hydrochloride exposureRobenidine hydrochloride（ROBH）is a one of the drug to treated myxosporidiosis of fish in recent decade.But the immune function effects of the drug to fish is less clear.The previous study of pharmacokinetics of ROBH was obtained an effective concentration in silver crucian carp blood was 1.17μg/mL.The present study have investigated the differentially expressed genes（DEGs）of GiCF cell of silver crucian carp（Carassius auratus gibelio）after ROBH exposed to 1.17μg/mL ROBH at 24h and 48h via high-throughput sequencing.A total of 118364 unigenes were obtained after assembled with the ranging from 201 to 13990 bp.1976 DEGs at 24h with 1271 up-regulated genes and 705 down-regulated genes and 2568 DEGs at 48h with 2071 up-regulated genes and 497 down-regulated genes were gained,and a total of 336 DEGs were obtained between these two points.For Kyoto Encyclopedia of Genes and Genomes（KEGG）enrichment analysis,more than 20 immune-related pathways were obtained,and we selected 3 pathways of them with P value<0.05,which are JAK-STAT signaling pathway,Natural killer cell mediated cytotoxicity and Th1 and Th2cell differentiation.According to further KEGG analysis based on P value>0.01,9 immune-related gene were obtained,which were confirmed by qRT-PCR.