| As an important freshwater economic fish,yellow catfish(Pelteobagrus fulvidraco)has been cultivated in many provinces of China,bringing huge economic benefits to the aquaculture industry in China.With the expansion of yellow catfish culture scale,the outbreak of bacterial diseases is becoming more and more frequent,which seriously threatens the healthy development of Yellow catfish culture industry in China.It has been reported all over the world that the use of vaccines in the process of aquaculture can effectively prevent the occurrence of fish diseases.In this study,we prepared a bivalent vaccine against two main bacteria of yellow catfish,and inoculated yellow catfish by immersion.Subsequently,the immunological effects of the vaccines were evaluated by detecting the changes of immunological indexes of yellow catfish after immunization.This research is divided into the following aspects.We used formaldehyde to inactivate Edwardsiella ictaluri and Aeromonas veronii to make a bivalent vaccine.The safety of the vaccine was tested by plate coating test and in vivo immune immersion test.The results showed that both the two uninactivated pathogens could infect healthy yellow catfish,and typical clinical symptoms were observed on the fish surface.In the inactivated pathogen soaking group,the yellow catfish remained normal and no bacterial growth was observed on the coated plate.These results indicated that the bivalent vaccine was safe and the live pathogen had high infectivity as an immunogen.To evaluate the immune effect of the vaccine,we inoculated healthy yellow catfish with the experimental vaccine by immersion immunization at 1.0×10~7cfu/m L.At 1,3,7,14 and 28 dpi,Serum,gill mucus and skin mucus were collected and their enzyme activities,immune-related gene expression levels and antibody titers were detected.The results showed that immune reactions occurred in the gill tissue and skin tissue after immunization,including thickening of skin epidermis,increasing number of mucous cells and widening of primary gill filaments.The results of enzyme activity detection showed that the activities of lysozyme,complement C3 and superoxide dismutase increased significantly in serum,gill mucus and skin mucus after immunization.Several toll-like receptor genes were found to be up-regulated in immuno-related gene expression levels.Among them,the expression level is highest in skin,followed by spleen.The expression of C-type lectin in spleen,gill and skin was significantly upregulated within 14 dpi,and the change was most rapid in gill tissue,which reached the maximum value at 1 dpi,while the expression peak in spleen and skin tissue appeared at 1 dpi.The expression levels of IL-1βgene were increased in both mucosal and systemic tissues,among which IL-1βgene was significantly increased in gill and skin mucosal tissues at 1 dpi.When aeromonas veroniae was used as the immunogen,it was found that the antibody titer could be detected in serum and gill mucus as early as 7 dpi and continued to increase,finally reaching a peak at 28 dpi.Antibody titers in skin mucus were detected only at 14and 28 dpi.The antibody titers in serum,gill mucus and skin mucus could be detected at 7,14 and 28 dpi with Edwardii catfish.The antibody titers in serum and skin reached the highest value at 14 dpi,and the antibody titers in gill mucus reached the highest value at28 dpi.After soaking and inoculation at 28 dpi,challenge infection experiment was carried out.Statistical results showed that the relative immune protection rates against Aeromonas veronii and Edwardsiella ictaluri were 71.11%、69.17%respectively.These results indicated that inoculation with dual inactivated vaccine could improve the immune capacity of yellow catfish and enhance the resistance to Aeromonas veronii and Edwardsiella ictaluri. |
