Construction Of Cbm-deficient Strain Of Streptococcus Suis Type 9 Mutant And Immunogenicity Characteristics Of Cbm

Streptococcus suis can cause a variety of diseases,including arthritis,meningitis and septicemia,which cause greater economic losses in pig farming.Type 2 and 9 are one of the predominant serotypes of S.suis in swine herds.At present,there is no effective vaccine to prevent S.suis.The study of the pathogenic mechanism is one of the bases for the prevention and control of S.suis.We isolated a S.suis of type 9,designated as DN13.After 10 passages in mice,it was found that the virulence of the passage bacterial(SS9-P10)was significantly enhanced in mice,we have sequenced the complete genome of DN13 and whole genome resequencing of SS9-P10.Bioinformatics analysis showed that the collagen-binding protein Cbm which containing the "EEPTTTTTT" repeat and the LPXTG motif,lots of site mutations occupied in the non-repetitive region of SS9-P10,compared to the parent strain DN13.To study the role of Cbm in S.suis type 9,we used the homologous recombination method to obtain the cbm gene deletion strain of SS9-P10.After the absence of Cbm in SS9-P10,the results showed that the growth rate and morphology of cbm-deficient strains did not change significantly when compared with SS9-P10.But the virulence of the bacteria decreased in mice.Compared with the parental strain SS9-P10,the mortality of the mice was reduced by 62.5% after the Cbm deletion,and the bacterial virulence was restored after the cbm gene was complemented to the deletion strain,but the mortality rate of the mice was 75%,which was lower than SS9-P10,in which the mortality rate of mice was 100%.The study indicates that cbm is one of the virulence-related genes of S.suis and may be involved in the pathogenicity of S.suis to the host.We expressed the Cbm protein of EEPPTTTTT-free repeat and the Cbm protein containing the repeat,respectively.The purified two Cbm proteins were added with aluminum hydroxide adjuvant to immunize the mice three times.The results showed that the mortality of mice immunized with repetitive sequence of Cbm protein was 100%,and with repeat-free sequence of Cbm protein was 75%,significantly higher than that of the control group(25%),suggesting that Cbm-induced antibodies enhance bacterial infection,anti-Cbm Antibodies may be beneficial for S.suis to evade host immune mechanisms.