Induction of alpha toxin production and reduction of time of cultivation of Clostridium novyi for vaccine production

Introduction and Objectives: Clostridium novyi is an anaerobic bacterium and has a longer growth period than other Clostridum strains, which itself produces long-time toxin production and inactivates and hydrolyzes toxin over a long period of time. The aim of this work was reduction of time of growth and high toxin production with medium optimization and growth conditions. Materials and Methods: in this study, based on the characteristics of the bacterium and its metabolism, different media with various source of carbons and nitrogen and their percentage in the media were analyzed. In this step, growth of C. novyi, time of growth and toxin production were controlled. The optimized medium and the old one was used for alpha toxin production in class bottle (16 litters) for final vaccine production. Also, the optimized medium was used for growth of C. novyi in fermenter condition. Final results were analyzed for the best medium and growth condition for vaccine production. Results: The bacterial growth conditions decreased from 72 hours to 24 hours, with quantitative and qualitative changes in the culture medium and the use of appropriate amounts as well as material displacement. Alpha toxin increased significantly due to the high growth of the bacterium as well as the reduction of growth time. In these changes, carbon source displacement has created favorable conditions for the growth and production of toxin. Also, removal of liver extract as an unnecessary capacity in the culture medium reduced the cost of consumable raw materials. Adding small quantities of vitamin and mineral compounds stimulated more and better growth and was effective in the rate of toxin production. Comparison of production conditions in class bottle (16 litters) culture with two old and new culture medium formulations showed that the new formulation culture medium has higher growth rates and more toxin production than the old formulation culture medium. The growth rate has dropped to about one-third and the amount of toxin has increased significantly. The evaluation of the presence of toxin by SDS-PAGE and ELISA control methods indicated that the presence of toxin levels was approximately 3 times higher in new culture. The results of this study showed that growth of the bacterium in the fermenter with new medium formulation was effective in producing 40-50% more of the toxin compare with other methods and the production process is much easier and more voluminous. Conclusion: These changes in media and growth conditions reduce production costs by at least 30 percent in the consumption of materials and also more than 100 percent in the cost of the production process. This achievement is currently underway at Razi vaccine and serum research Institute of Mashhad as a production process for the production of black disease vaccine.