Qβ Virus-like Particle As Carrier To Conjugate With Streptococcus Suis Serotype Ⅱ Capsular Polysaccharide

Streptococcus suis（S.suis）serotype 2,the most common pathogenic of swine origin,is a zoonotic pathogen which can cause meningitis,arthritis,endocarditis,septicemia and other diseases in humans or pigs after infection.Patients even appear sequelae such as deafness.S.suis capsular polysaccharide is the main virulence factor,but also an important protective antigen,one of the important targets for vaccine development.Due to its carbohydrate properties,the immunogenicity of capsular polysaccharide is lacked,we decided to improve its immunogenicity by conjugating it with Qβvirus-like particles（VLPs）.In this study,after chemical coupling between the polysaccharide and the nanoparticle carrier,we successfully prepared conjugates of S.suis serotype 2 capsular polysaccharide and QβVLPs.It provides an important basis for the research and the development of S.suis vaccines and glycoconjugate vaccines.The methods and results are as follows:1.The expression system of Escherichia coli for the preparation of QβVLPs was established.After purified by sucrose density gradient centrifugation and size exclusion chromatography（SEC）,a plethora of QβVLPs with high purity were obtained,and substantial VLPs with a diameter of about 28 nm were observed by transmission electron microscopy（TEM）.In addition,QβVLPs specific antibody from immunized rabbits was successfully prepared by using purified QβVLPs as an antigen.Indirect immunofluorescence assay showed that QβVLPs were internalized into PK15,3D4/2,Marc145,Vero and 293FT.In vitro thermal stability experiments showed that the morphology of QβVLPs particles remained intact and stable after 4 months of storage at-80℃,-20℃,4℃,37℃or room temperature,respectively.2.S.suis serotype 2 capsular polysaccharide（CPS2）was activated by the coupling agent1-cyano-4-dimethylaminopyridine tetrafluoroborate（CDAP）,and then was successfully conjugated with QβVLPs at p H 9.0,which was determined by SEC.The preliminary experiment of immunizing mice showed that CPS2 specific antibodies could be elicited in mice after immunizing glycoconjugates（QβVLPs-CPS2）,which was significantly higher than non-conjugated group（QβVLPs+CPS2）on 42^nd day after immunization.And challenge test results showed that the survival rate of mice in QβVLPs-CPS2 group was 80%（4/5）.The survival rate of QβVLPs+CPS2 group was 20%（1/5）,while the survival rate of PBS and QβVLPs group was 0（0/5）,suggesting that QβVLPs-CPS2 could provide better protection for mice.Conclusion:After conjugation of S.suis serotype 2 capsular polysaccharide as hapten with QβVLPs,the immunoassay results showed that antibodies against S.suis serotype 2capsular polysaccharide were produced in mice which immunized with glycoconjugates,indicating that conjugation of polysaccharides with QβVLPs can effectively improve the immunogenicity of polysaccharides.QβVLPs can be used as an effective nanoparticles carrier to improve the immunogenicity of hapten.These results provide an effective approach for the study of polysaccharide vaccine,which laying a foundation for the development of S.suis serotype 2 vaccine.