| In recent years, with the increasing development of aquaculture cultivation, fish disease has brought great loss. Abusing drugs and antibiotics gradually exposed a lot of shortcomings. Therefore the development and application of fish vaccines have become the primary way to solve the problems of healthy breeding. Now new pathogens continuously emerge and diverse types of diseases often occur in aquaculture. Vibro anguillarum and Edwardsiella tarda are two main pathogenic bacteria which caused vibrosis and edwardsiellosis on various kinds of fish in aquaculture cultivation. Also, vaccines in clinical application are usually single vaccines, effect of multiple vaccinations for several vaccines is not easily done and the workload is unavoidable in aquaculture. The use of combination vaccines can not only simplify the immunization program, but also reduce the immune costs and the damage of fish during the inoculation process. This study was based on two live attenuated vaccines V. anguillarum MVAV6203and E. tarda WED in our laboratory, and the combined immune efficacy was analyzed on zebrafish and turbot. In the meantime, immune efficacy of formalin-inactivated Vibro anguillarum with chitosan and chitosan oligosaccharide was analyzed.According to the vaccination strategy, we found that the immunoprotective effect of combination vaccines of two live attenuated vaccines was better than each single vaccine. On zebrafish and turbot, the relative protective survival (RPS) of combined immunization was not less than90%and70%to V. anguillarum and E. tarda after vaccination for one month, respectively. The immune response of the combination vaccines was analyzed with RT-qPCR analysis. It was found that Toll-like receptor signal pathway was activated and both MHC-I and and MHC-II pathways of antigen processing and presentation were aroused. The antibody levels of vaccinated turbot were also analyzed with ELISA. The results showed that the antibody levels of combination vaccines increased to some extent compared with single vaccine. Meanwhile, MHC-I and MHC-II pathways were both activated in vaccinated turbot, similar to the vaccinated zebrafish. In the analysis of immunoprotective effects about inactivated V. anguillarum, it was found that the RPS of vaccinated zebrafish without adjuvant for one month was about50%, but the RPS of the fish vaccinated with chitosan and chitosan oligosaccharide was up to85%, the similar immunoprotection as the veterinary oil adjuvant. Moreover the RPS of turbot vaccinated with inactivated V. anguillarum mixed with chitosan oligosaccharide for3months were not less than80%, which was significantly higher than the fish vaccinated with inactivated V. anguillarum alone. Furterhmore the antibody levels of sera from vaccinated turbot were analyzed with ELISA. The results indicated that chitosan oligosaccharide could enhance humoral immune response of inactivated V. anguillarum, and the sera could react with V. alginolyticus and V. harveyi as well. Therefore, chitosan oligosaccharide might be an effective adjuvant for fish vaccine.In a word, our work would promote the development of fish vaccine design and development. |
