Performance Of Pneumococcal LytA And CbpA For Diagnosis In A Rat Model Of Pneumococcal Infection

Objective:Streptococcus pneumoniae is a kind of conditioned pathogens to estoblish pharynx nasalis of 40%～50% healthy crowd, and lead to pneumonia, meningitis diseases,otitis media and bacteremia, especially in infants and elderly people. The World Health Organization estimates that at least 1.2 million children less than 5 years of age die each year in developing countries from pneumococcal pneumonia.Streptococcus pneumoniae rapid diagnostic tests were extremely important significance for early symptomatic treatment ,to reduce the overuse of antibiotics and reduce morbidity and mortality.To obtain pneumococcal LytA and CbpA by prokaryotic expression system and investigate their diagnosis for infection caused by Streptococcus pneumoniae.Methods:1.Prokaryotic Expression and Purification for lytA and cbpA :The specific primers were designed according to lytA and cbpA of Streptococcus pneumoniae gene sequence. lytA and cbpA were amplified by PCR form the pneumococcus genome.After IPTG inducing ,the recombinant proteins were purified by electroeluting of bag filter,detected by SDS-PAGE and Western-blot.2.Infected animal models:By considering infection group and infected control group≥1:160 fold increase in antibody concentration as qualified serum.3.ELISA: Serum IgG and IgM antibodies accordingly of BALB / c mice infected with Streptococcus pneumoniae were detected by ELISA .Results:1.The recombinant plasmid pET-32a(+)-lytA and pET-32a(+)-cbpA were constructed successfully. Fusion proteins LytA and CbpA were expressed and displayed expected antigenicity.2.Infected animal models were constructed successfully. To determine the accuracy of the source of infection in serum samples.3.IgM and IgG antibodies level anti LytA were significantly higher than the control group (Infections with B Streptococcus group and healthy mice)(P<0.05),but antibodies level anti CbpA did not increase as compared with group infected with B Streptococcus(P>0.05) . Diagnostic sensitivity of CbpA was 83.3%( IgG)and 75%(IgM). Diagnostic specificity of LytA was100% (IgG and IgM).Conclusion:1.It were confirmed some antibodies stimulated by Streptococcus pneumoniae would appear in serum and can be used for serological diagnosis.2.The recombinant plasmid pET-32a(+)/cbpA and pET-32a(+)/lytA were constructed successfully.The expressed and purified recombinant proteins were proved with immunogenicity and can be used for serological test.3.The synergistic use of specificity of LytA and sensitivity of CbpA may be worthy of serological diagnosis for Streptococcus pneumoniae infection, and may be used for further clinical test .