Evaluation Of Combined Immunization With Two Live Attenuated Vaccines And Oral Vaccines Development

Edwardsiella piscicida and Vibrio anguillarum are two common pathogens of marine fish,which infect many economic fish species and cause vibrosis and ewardsiellosis.In this study,we selected live attenuated E.tarda vaccine WED and V.anguillarum attenuated vaccine MVAV6203 that developed in our laboratory,and inoculated turbot with these two vaccines.By investigating the efficacy of the two vaccines,we analyzed the synergy or inhibition of the two vaccines in the course of combined immunization,and further explored the mechanism of immune synergy or inhibition.The results showed that the combined vaccines could significantly improve the immune protection of turbot against V.anguillarum compared with single vaccination.Specific antibodies increased significantly in serum.The expression of genes related to antigen recognition and presentation,such as TLR2,TLR5,TLR21,MHC ? and MHC ?,increased at the transcriptional level,which confirmed that the combined vaccination of the two vaccines had a synergistic immune effect.In addition,inactivated water-type and oil-type V.anguillarum vaccines were prepared,and the immune potential of the two kinds of vaccines as oral vaccines for turbot was analyzed.The results showed that sodium alginate concentration of 3%(w/v),vaccine concentration of 2.5 OD,emulsifier Sipan-80 dosage of 0.2%,water-oil ratio of 1:9,and rotational speed of 2000 r/min were the optimal conditions for preparation.However,it did not achieve good immune protection effect at all.Then food-grade fish oil was selected.Bacteria concentration of 5 OD,ratio of oil to water of 1:9,rotational speed of 4000 r/min and the amount of emulsifier of 3%(w/v)were the optimal conditions for preparation.Clinical data showed that the vaccine could improve the disease resistance of juvenile turbot.In summary,the combined immunization effect of two attenuated live vaccines for marine fish was evaluated and the synergistic mechanism was analyzed.On the other hand,the inactivated oil-type V.anguillarum vaccine was developed.Totally,our results provided some data for the development and evaluation of fish vaccines.