Biosafety Evaluation of Canine Adenovirus type-2 Expressing Rabies Virus Glycoprotein in Dogs

To evaluate the biosafety in the enviroment release of the recombinant canine adenovirus type-2 expressing the rabies virus glycoprotein,500 unvaccinated dogs were split into two groups,250 in each group.Group 1 was intramuscularly injected with 1 ml(107.75 TCID50 /ml) and Group 2 was orally inoculated with 3 ml(107.75 TCID50 /ml),respectively.At the vaccination and after,the dog sera were collected weekly or biweekly and the following items were evaluated: the genetic stability of the recombinant vaccine in cell passage and in dogs;the existence and sustaining of the recombinant virus in the body and in the feces of the immunized dogs;the toxic effect of the recombinant virus on dogs immunized with overdoes;the sepcific antibody against the recombinant virus in the bodies of producers,the experimenters and animals around the experimental area;the contamination of the exogenous(i.e.,the glycoprotein) gene in the environment around the experimental area during the experiment and 6 months after the experiment;the neutralizing antibody level in dogs and the kinetics of the neutralizing antibody.Results showed that after 79 passages in MDCK cells and 11 passages in dogs,no deletion or recombination of the foreign gene in the recombinant virus was detected.And no specific nucleotide sequence of the recombinant virus was assayed in the bacteria isolated from the tissues of the immunized dogs.No contamination of the recombinant virus and the glycoprotein gene was found out in the environment.Increasing the amount of the recombinant virus did no harm to the dogs and no toxicity was observed.No specific antibody against the recombinant virus was assayed in the producers,experimenters and the animals around the experimental area except for several dogs naturally infected with the canine adenovirus.Neutralizing antibody was detectable monthly after the intramuscular and oral vaccination,respectively.The antibody could persist for 54 weeks with an effective level of protection.All the above demonstrated that the recombinant virus was safe to both the dogs and the environment.