Study On Anti-PEDV Plant-derived Drugs Screening And Their Antiviral Effects

Porcine epidemic diarrhea(PED)is a kind of intestinal infectious disease caused by porcine epidemic diarrhea virus(PEDV).At the time of the outbreak,the mortality of newborn piglets was as high as 100%.At present,there is no effective antiviral drug for controlling this disease.In this paper,we plan screening of plant-derived antiviral chemical components in vitro,such as triterpenoids,phenolic acids,flavonoids and alkaloids.We will select safe and efficient compounds according to their IC50,CC50 and SI values.The selected active compounds were used to detect their antiviral activity in different periods of infection,and the effects of the above active compounds on the proliferation of DR13att were investigated.The proliferation of virus was observed by rPEDV-ΔORF3-GFP infection,and the antiviral effect of these compounds in different period of infection was analyzed.The results of the experiments of acute toxicity in mice and the animal experiment of piglets were used to verify their antiviral effect of high activity compounds.The antiviral effect of piglet in vivo was evaluated by clinical symptom observation,pathological changes,viral load and piglet survival rate.The above research can provide possible drugs and theoretical basis for the prevention and treatment of PEDV.The specific results are as follows:Firstly,cepharanthine,tetrandrine,fangchinoline,lycorine hydrochloride,berberine hydrochloride,oxygenine hydrochloride,quercetin,hesperidin,neohesperidin,naringin,caffeic acid,chlorogenic acid,gallic acid,resveratrol and magnolol,were detected to get antiviral compounds by CPE of vero cells induced by PEDV-DR13att in vivo.It was found for the first time that epharanthine(CEP),tetrandrine(TET),fangchinoline(FAN),and magnolol(MAG)inhibited PEDV with EC50 of 2.53,3.50,6.69 and 28.21 μM,respectively.Their CC50 were 29.92,24.78,30.19 and 57.28 μM,respectively.Their SI values were 11.83,7.08,4.51 and 2.03,respectively.We further found that 10-20 μM CEP,TET,FAN and 20-30 μM MAG had a good effect on PEDV.20 μM CEP and 30 μM MAG could inhibit the proliferation of PEDV effectively in vitro.Secondly,different concentrations of CEP,TET,FAN and MAG were pretreated 24 hours before rPEDV-△ORF3-GFP infection,co-treated vero cells infected by PEDV,and post-treated 24 hours after infection to investigate their antiviral effect at different stages of PEDV infection,the content of virus in cells gradually decreased,and the antiviral effect gradually increased,showing a dose-dependent effect.After that,the mechanism of antiviral compounds against PEDV was discussed.20 μM CEP,TET and FAN could significantly affected the adsorption of PEDV into cells through three ways of administration,while 30 μM MAG significantly affected the adsorption of PEDV into cells and the replication of PEDV through three ways of administration.Finally,the results of acute toxicity and subchronic toxicity experiments in mice showed that 100 mg/kg CEP had no toxicity to mice,and the safe dose of CEP was.11.1 mg/kg.The results of piglets PED treatment experiment showed that the viral load of high-dose CEP treatment group was significantly lower than that of control group one day after administration(3 dpc),and the intestinal villi of treatment group were intact without shedding,which indicated that high-dose CEP had a certain inhibitory effect on PEDV in piglets.