Immunization Studies To The Factor B Of Surface Iron From Staphylococcus Aureus

Staphylococcus aureus is an important pathogenic bacteria, it caused diseases in animal and human. Its growth and reproduction are both dependent on the iron in host cell. IsdB protein is the essential hemoglobin receptor of S.—aureus for the uptake system of iron, it plays an important role in the process of infection, colonization and pathogenicity. IsdB gene is highly conserved, and found the pathogenicity will decline after knocking out IsdB gene from the strain. It has been reported that IsdB protein have good immunogenicity in mice and rhesus monkeys and its has more effective function of protection than other S. aureus cell surface proteins. Therefore we cloned and expressed IsdB protein, and compared with mouse immune model to analyze the immunogenicity of Staphylococcus aureus, we also researched on the IsdB protein vaccine for the protective effect of S. aureus infection, and provided the data for the relevant research and development support about vaccine.In this study, we cultured and detected59Staphylococcus aureus strains from24cow farm in Shandong and neighboring cattle farm. Designed and amplified IsdB gene sequence according to No.BA000033. Design IsdB gene primer with primer5.0and DNAstar. Extraction DNA of Staphylococcus aureus strains and amplifily IsdB gene, purify IsdB gene, then translated into E.coli DH5α. Digesting plamid extracted from E.coli DH5α and detected the sequence of the purifying production. Connecting purified IsdB gene and plamid pQE-32, translated into E.coli M15, induction express IsdB protein. Detecting recombine IsdB protein with western blotting, and purify IsdB. Immunization mice with IsdB and detected the immunopotency. To detecte the protect effect of immuned mouse with Staphylococcus aureus, record the result.We get the1938bp DNA fragment from59staphylococcus aureus strains, and the fragments have connected with pMD18-T cloning vectors. The pMD18-T-IsdB have translated into E.coli DH5a successfully. IsdB gene digested from pMD18-T-IsdB have connected with pQE-32and translated into E.coli M15successfully. The recombined pQE-32-IsdB have express72ku IsdB protein. The IsdB protein is36%in all extracted proteins. Used the purified recombinant IsdB protein to immune mice, the level of IgG in the serum of mice immuned two weeks before could reach1:64000. The toxicity test results showed that the protection rate of SARI11, SAR-12and SAR-16Staphylococcus aureus strains is80%,80%and50%by the IsdB protein in recombinant protein immunostaining groups. While the whole bacteria groups protection of SAR111was70%.In summary, the bovine source of Staphylococcus aureus IsdB gene is highly conserved, in the immune test about the expression of IsdB protein showed good immunogenicity and immunoprotection function, so the IsdB protein can be used as an antigen of the S. aureus vaccine.