Prokaryotically Express Virulence Associated Proteins Of Streptococcus Suis Type 2 And Evaluation Of Immunogenicity And Protective Capacity Of Recombination Proteins In Vitro

Streptococcus suis type 2 is an important zoonotic pathogenic bacteria. The invasion mechanism of Streptococcus suis type 2 is still indistinct. Therefore, it is crucial to the efficient preventation and controllability of S. suis type 2 infections whether we could develop efficacious vaccine and diagnostic reagent. Nowadays, there are three main types of vaccine: live attenuated vaccine; well-known virulence factors; immunogenic proteins of CW/CM.Our team, use to establish an antigenic profile of OMPs and extracellular proteins of the strain 98-149, high virulent isolate from the outbreak in Jiangsu Province in 1998, was established by immunoproteomics analysis and several immunoreactive proteins unpublished were discovered.The genes encoding the 16 immunogenic proteins, which were screened by immunoproteome, were amplified from genomic DNA of Streptococcus suis type 2 strain 98-149 by polymerase chain reaction (PCR) . Then the amplified fragments encoding the putative antigenic epitopes were cloned in the proper orientation into pET32a ( + ) via restriction endonucleases. The recombinant plasmid was verified by restriction endonuclease analysis and nucleotide sequencing, and them transformed into host E. coli strain BL21, recombinant proteins was highly expressed after induced by ITPG and purified by Ni-nitrilotriacetic acid affinity chromatography. Western blot analysis showed that the recombinant proteins were reactive to with the serum from the rat infected with strain 98-149. The antibody titer in blood serum reached 10~5 after immunization with the purified recombinant proteins five times. ELISA was performed to assess the presence of specific antibodies to recombinant proteins in serum from humans and post swines infected by S.suis type 2. The results clearly indicated that post immunization serum strongly immunoreactive with recombinant antigens. The bactericidal activity of human blood opsonized with immune serum from rat was assessed by a direct bactericidal activity. This assay demonstrated that some recombinant proteins were protective antigen just like MRP, known as a high virulence factors. This work provides a very important theorical evidence for the development and further research on the new protective antigen. Meanwhile, it also provides the basis for the search of the virulence-associated antigen and the research and exploitation of this candidate vaccine.