Immunogenicity and protective role of antigenic regions from five outer membrane proteins of Flavobacterium columnare in grass carp Ctenopharyngodon idella

Flavobacterium columnare causes columnaris disease in freshwater fi sh. In the present study, the antigenic regions of fi ve outer membrane proteins(OMPs), including zinc metalloprotease, prolyl oligopeptidase, thermolysin, collagenase and chondroitin AC lyase, were bioinformatically analyzed, fused together, and then expressed as a recombinant fusion protein in Escherichia coli. The expressed protein of 95.6 k Da, as estimated by 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was consistent with the molecular weight deduced from the amino acid sequence. The purifi ed recombinant protein was used to vaccinate the grass carp, C tenopharyngodon idella. Following vaccination of the fi sh their Ig M antibody levels were examined, as was the expression of I g M, Ig D and Ig Z immunoglobulin genes and other genes such as MHC Iα and MHC I I β, which are also involved in adaptive immunity. Interleukin genes( IL), including I L- 1β, IL- 8 and I L- 10, and type I and type II interferon(I FN) genes were also examined. At 3 and 4 weeks post-vaccination(wpv), signifi cant increases in Ig M antibody levels were observed in the fi sh vaccinated with the recombinant fusion protein, and an increase in the expression levels of I g M, Ig D and Ig Z genes was also detected following the vaccinations, thus indicating that an adaptive immune response was induced by the vaccinations. Early increases in the expression levels of IL and IFN genes were also observed in the vaccinated fi sh. At four wpv, the fi sh were challenged with F. column a re, and the vaccinated fi sh showed a good level of protection against this pathogen, with 39% relative percent survival(RPS) compared with the control group. It can be concluded, therefore, that the fi ve OMPs, in the form of a recombinant fusion protein vaccine, induced an immune response in fi sh and protection against F. columnare.

Elementary study on the antigenicity of SARS-CoV Mc region

To test the antigenic activity of M protein （Mc protein） in the inner membrane of SARS-CoV, SARS-CoV Me protein＇s bases locating inside the membrane were cloned, the His-fusion protein was expressed in E. coli and analyzed for its antigenic activity. Among those 7 clinically diagnosed patients＇ sera, there were 5 positive and 2 negative in reaction with His-fusion protein. All of the 20 healthy persons＇ sera and rabbit anti-OC43 and 229E were of negative reaction with His-fusion protein. The animals immunized with His-fusion protein have produced muhi-clonal antibody. The His-fusion protein could specially react with clinically diagnosed SARS patients＇ sera and the animals immunized with His-fusion protein could produce specifically multi-clonal antibody, but it could not react with the sera of healthy persons and the rabbit anti-OC43 and 229E.