Toxicity Of APP Grown Under Different Contents Of NAD Conditions And Preparation Of Polyvalent Inactivated Vaccine

Actinobacillus Pleuropneumoniae(APP) is the causative agent of Porcine Pleuropneumoniae, a worldwide disease causing tremendous economic loss to the swine industry. It is a common infectious disease which infects the respiratory tract of pigs. The symptoms of infected pigs induced depression, difficult respiratory and pneumonia. With the development of the raising cosmically, Porcine Pleuropneumoniae becomes one of the five primary disease that threaten mordern pig industry.The following two parts were included in the studies:Part I: The pathogenicity and immunoprotection of APP grown under different contents of NAD conditions in miceThe Actinobacillus Pleuropneumoniae(APP) serotype 5 were cultured respectively on two lauria broth ( LB ) agar which supplemented with 0.05% (V/V)and 0.001%(V/V)nicotinamide ademine dinucleotide (NAD), inoculate 30days old mice with these two APP by 3×10~9CFU per mice, After inoculation, observation the pathogenicity of APP serotype 5 grown under different contents of NAD conditions in mice .result,.a lot of mice which inoculated with APP serotype 5 grown under 0.001%(V/V)NAD condition shown severe lung lesions but few mice which inoculated with APP serotype 5 grown under 0.05%(V/V)NAD condition shown severe lung lesions, many mice shown light lung lesions.The two bacteria were emulsified in an equal volume of a mineral oil adjuvants, at the age of 3 and 6 weeks, the mice were vaccinated with the vaccines. Three weeks after the second vaccination, all mice were challenged with an APP serotype 5, Sera were collected before the second vaccination and before the challenge, and ELISA. Result,the protection to mice of the vaccines were made of APP serotype 5 grown under 0.001%(V/V)NAD condition and grown under 0.05%(V/V)NAD condition had no significant difference,but after vaccinated with vaccine which be made of APP serotype 5 grown under 0.001%(V/V)NAD,the mice shown light lung lesions when challenged with APP serotype 5 and after vaccinated with vaccine which be made of APP serotype 5 grown under 0.05%(V/V)NAD,the mice shown severe lung lesions when challenged with APP serotype 5. so ,the pathogenicity and immunoprotection were different when APP growend under different contents of NAD conditions.Part II: Preparation of polyvalent oil-adjuvan Inactivated vaccines into Actinobacillus Pleuropneumoniae and protection in miceAccording to the result which restriction NAD in medium can improve the immunoprotection of APP. We culture serotype 1,3,5,7 respectively on LB agar which supplemented with 0.001%(V/V)NAD and 5%(V/V)calf serum, the cultures were rinsed off from plates with PBS , suspensions from several plates were pooled. After inactivated with formalin, different serotype suspensions were pooled with equal doses, the bacteria were emulsified in a suitable volume of mineral oil adjuvants. at the age of 3 and 5 weeks, the mice were vaccinated 0.3mL per mice with the vaccines,3 weeks after the second vaccination, all mice were challenged with APP , Result,the protection to mice of the vaccines against APP 4 different serotype were 91.7%,100%,91.7% and 91.7%; Sera antibody was detected by ELISA, 3 weeks after the second vaccination, the sera antibody of mice achieved peak, the OD of 450nm near to 0.6,10 weeks after the first vaccination, the OD of 450nm were also above 0.4. The results suggest that the polyvalent oil-adjuvan inactivated vaccines had good protection and safety in mice, this study did the groundwork to prepare the inactivated vaccines of pigs.