Thetechnology And Clinical Trialsof Rabbit Haemorrhagic Disease Virus Baculovirus Vector Inactiveted Vaccine

Rabbit hemorrhagic disease(RHD)is caused by rabbit hemorrhagic disease virus(RHDV),which is characterized by acute necrotizing hepatitis and highly mortality for rabbits.RHD has been classified as a class B infectious disease by the World Organisation for Animal Health(OIE)while it has been issued as a secondary epidemic by Ministry of Agriculture of the People's Republic of China.Immunization with inactivated tissue vaccine is the most important measure to prevent and control the disease.However,as the source of the non-immunized rabbits for the produce of tissue vaccine is uncertain,and uncompletely inactivation causes the risk of infection,the novel available vaccine should be developed.The study carried out production technology research and produced 10 batches of vaccine in the pilot scale test for the Rabbit Haemorrhagic Disease Virus Baculovirus Vector Inactivated Vaccine which was prepared by the recombinant RHDV carried by baculovirus.With the approval of the Ministry of Agriculture,the vaccine clinical trail,the safety test,the clinical immune effect test and the biosafety test were conducted.Finally,the Rabbit Haemorrhagic Disease Virus Baculovirus Vector Inactivated Vaccine obtained the certification of the new veterinary drug.Comparison of process with roller bottles and suspension technology of recombinant rabbit hemorrhagic virus baculovirus were performed.During pilot scale production and large-scale production,the culture conditions should be controlled at 27?28?,pH value ranged from 6 to 6.2 and speed 15 r/min.When the cell concentration was about 2 × 106/mL,1%of the total volume was continuously received for inoculation and observed for 5 days.Cell cultures can be harvested when the cell lesion is more than 85%.The pilot scale test was carried out and produced 10 bathes of vaccine continuously in Nanjing Tianbang Bio-industy Co.LTD from February to July,2013,according to the "procedures of manufacture and infection ofRabbit Haemorrhagic Disease Virus Baculovirus Vector Inactivated Vaccine".Then we analyzed the traits,sterility,and the amount of residual formaldehyde,safety and efficiency of these vaccines according to the current the Veterinary Pharmacopoeia of the People's Republic of China.The results of the inspection showed these 10 batches of vaccines met the quality standard in sterility,the amount of residual formaldehyde,safety check and efficiency test.The study indicated that vaccines with consistent quality were commercially available by large scale production.The safety test of 3 batches vaccines were conducted on rabbits of different varieties and ages in 6 different rabbit farms.We chose 3 varieties rabbits,meat rabbit,Rex rabbit and wool rabbit,for the safety evaluation.The Rabbits were segmented into 4 groups,healthy and susceptible rabbits of 2 months old,35 days(the minimum suitable age),20-30 days(under suitable age)and pregnant 20-25days.Then,the rabbits were injected vaccines with single dose,single dose twice and two doses by subcutaneous injection respectively.The results showed all the rabbits were alive during the observation period(the first 14 days).There was no inflammatory reaction in the injection sites at days 30 and days 60 after vaccination,and the vaccines were well absorbed.The vaccines were safe for pregnant rabbit,not weaning rabbit and weaning rabbit,and also had no impact on the growth performance of the rabbit.Then,we detected the recombinant baculovirus in the tissues,organs,excreta and living environment of immunized rabbits.The result showed that there were no recombinant baculovirus residues in liver,lung,muscle,fur,faecesandurine.No recombinant baculovirus were detected in the environment,including the soil,the water and the air.The above studies indicated the inactivated vaccine was safe after being injected in meat rabbit,Rex rabbit and wool rabbit.The safety level of the vaccine is level ?,and no potential risk of bio-safety.We chose 3 batches of products testing the effect in 6 different rabbit farms.The challenge tests were carried out during the different immune stage.There was no RHD occurred in 6 farms during the tests.The results represented the effective antibody against RHDV lasted for 7 months.