Study On Regulatory Effects Of OMP And LPS On Fish Immune Defense Function

In the present study, OmpV and Omp from V. alginolyticus were prepared andlipopolysaccharide (LPS) was extracted from Aeromonas hydrophila by usingimmunological technology, microbiological molecular biology technology andproteinic technology. Both immune protection and generation regulation of antibodyof yellow croaker (Pseudosciaena crocea) inoculated with FKC, OmpV and Ompwere investigated respectively. The immune polysaccharide could greatly improveimmune reaction of ricefield eel to Aeromonas hydrophila.According to V. alginolyticus gene sequence from Genebank, the primer ofOmpV was designed and OmpV gene at a length of 720 bp was cloned by PCR.OmpV gene was recombined to plasmid pET30a, and then cloned in E. coli DH5a.Finally pET30a-OmpV was extracted and expressed in BL21. Analyzed byelectrophoresis method, the molecular weight of OmpV is 27kDa. The result ofwestern blotting analysis indicated that the protein we extracted was OmpV.Purified OmpV was used to immunize P. crocea. After injection of OmpV, fishappeared normal. The result indicated that 80% of P. crocea were protected by OmpV,which was considered as a perfect vaccine.Three kinds of bacterin including FKC, OmpV and Omp were prepared from V.alginolyticus, and then they were used to immunize P. crocea. Serum of fish wasseparated and analyzed from fourth week to eighth week of experiment. The antibodytiter of FKC, OmpV and Omp were measured by ELISA method, in order todetermine the optimum time for vaccination. At the fourth week or even earlier, theantibody peaked in FKC immunized group. At the fifth week, the antibody peaked inOmpV immunized group. At the seventh week, the antibody peaked in Ompimmunized group. The results indicated that four weeks before V. alginolyticusepidemics was the best time for vaccination of P. crocea.Aimed at evaluate the effects on improving the immune response, immunepolysaccharide (yeast cell wall) was fed at a fixed ratio of 100.0 mg/kg to ricefield eel which was injected LPS form A. hydrophila. The results showed that addingimmune polysaccharide into feed can not only enhance the immune response and theresistance of ricefield eel to A. hydrophila, but also improve their liver function andprompt their growth to some degrees.