The Distribution Of Eimeria.Tenella DNA Vaccine PVAX1.0-TA4-IL-2in Organic Tissues And Its Release To The Environment

To evaluate the biosafety of the E. tenella DNA vaccines pVAX1.0-TA4-IL-2, the following trials have been fulfilled.The biosafety of the DNA vaccine was evaluated basing on Rules of Management on Agricultural Transgenic organision. The first rule is about that the biosafety evaluation of a certain immunization on the youngest target animal by different routes. As the require of the first rule, the experimental groups were immunized only once with25μg of the vaccine pVAΧ1.0-TA4-IL-2at the age of one day by the following routes, i.e. intranasal—ular administration, oral administration, wing-intravenous injection, neck-subcutaneous injection, intramuscular injection and the negative control group was injected with PBS. The second rule is about that the biosafty evaluation of multi-immunization on the target animal with the DNA vaccine with a certain dose. As the require of the second rule, the chickens were divided into two groups:the control group and the experimental group. The experimental group was muscularly injected with25μg per chicken in leg at7,14and21days and the control group was injected with PBS. The third rule is about that the biosafety evaluation of single immunization on the target animal respectively with three batches of DNA vaccine with the dose three times than normal in the target animal. As the request of the third rule, the DNA vaccine was injected with75μg per chicken by intramuscular injection at14days each batch. The fourth rule is about that the biosafety evaluation of single immunization on the non-target animal with the DNA vaccine. So as the request of the fourth rule,ten white mouse were injected with25μg by intramuscular injection at30days.The biosafety was evaluated by means of observation on the development of immune responses in immunized chickens, such as mental status, foraging, drinking water, intestinal discharge and the death of chickens. One pair of primers were designed and synthesized according to the sequences of TA4gene, to monitor the transfer of the Kanamycin-resistant gene and TA4gene, detect the distributions and the persistence of plasmid in main organs and whether the plasmid had integrated with chromosome DNA, the injected tissues as well as the gonads to analyze the influences of vaccine on their puppies. The pathological effect in the main organs was observed.The results showed that the DNA vaccine did not induce any abnormal clinical responses in the immunized chickens. The Kanamycin-resistant gene and TA4didn’t transfer into the normal intestinal flora, showing that this DNA vaccine didn’t excrete into the surroundings through intestinal flora, feces. The plasmid could exist in heart，spleen, kidney and the injected tissues at least for two months, while it could not be detected in the liver. The vaccine had no influence on their puppies because it was not detected in the gonads. The plasmid DNA didn’t integrate with chromosome DNA. Also, there was no pathological effect observed in the main organs.In one word, the DNA vaccine had no problems on biosafety, according to the requirement of Rules of Management on Agricultural Transgenic organision.