Construction And Immunologic Study Of The Recombinant Mycobacterium-Eg95 Vaccine Of Echinococcus Granulosus With Nonsecreting Type And Secreting Type

Objective To construct the mycobacterium-Eg95 vaccine of E.granulosus with nonsecreting type and secreting type,and to study the immunological effect of the recombinant vaccine.Methods BCG-Ag85B signal sequence with 120bp and Eg95 gene with 423bp were amplified from the genome BCG and PGEX-4T-Eg95 by PCR.Firstly,Eg95 gene fraction was cloned into E.coli- BCG Shuttle-vector pMV261 to get the recombinant nonsecreting plasmid pMEg95.Next,BCG-Ag85B signal sequence was cloned into pMEg95 to get the recombinant secreting plasmid pSMEg95.The recombinant plasmid was transformed into BCG and M.smegmatis respectively by electroporation.And humoral immune indexes were detected by ELISA.Results It was confirmed by restriction endonuclease digestion,PCR amplification and gene sequencing that Eg95 coding gene and BCG-Ag85B signal sequence coding gene were successfully cloned into pMV261 as well as the recombinant nonsecreting and secreting BCG-Eg95 and M.s-Eg95 vaccine of E.granulosus were successfully constructed.Mice were challenged by BCG-Eg95 and M.s-Eg95 vaccine,and antibody titer was detected by ELISA.The results showed that the BCG-Eg95 had a better effect than the M.s-Eg95.Conclusion The recombinant BCG-Eg95 and M.s-Eg95 vaccine of E.granulosus with both nonsecreting type and secreting type have been constructed.By analyzing and comparing the humoral immune indexes of the two type vaccines,the immunological effect of the recombinant BCG-Eg95 vaccine was better than M.s-Eg95.