Preparation Of The Vaccine Against Cervus Hemorrhagic Enteritis Of And Nucleotide Sequencing Of The Alpha-toxin Of Clostricium Perfringens

In this study, strains of Clostridium perfringens were isolated from viscus and intestinal contents of the dead cervus in Shanxi Province. 22 strains were identified as Clostridium perfringens type A from 28 cases caught hemorrhagic enteritis. This indicates that the pathogen of hemorrhagic enteritis was mainly Clostridium perfringens type A in cervus in Shanxi province. One strain with the most strong virulence was selected from 22 isolates and named J5 and evaluated the best conditions of producing toxin. The results suggested that the virulence of toxin of J5 which was cultured in 35℃, anaerobic beef pepsin digest broth and anaerobic environment was the most strong, amounting to 100MLD/ml. The strain was cultured in this condition, then was inactivated with 0.3% formaldehyde. Five days later, 10% Al(OH)3 was added into the qualified liquid and stood for 10d at 2 to 8℃, then moved away pure liquid of 50 percent of the gross volume, shook and divided into bottles. The results of safety, efficacy and field tests showed that the vaccine had better immunogenicity and could be used safely. The major toxin of Clostridium perfringens type A was alpha-toxin. In this study, the recombinant plasmid of alpha-toxin gene was made then alpha-toxin gene was cloned and nucleotide sequence was determined. The results obtained in this research was agreement with the standard gene sequence of alpha-toxin and lay a good foundation for the preparation of gene vaccine.