金黄色葡萄球菌IsdA的B细胞免疫显性表位的鉴定及免疫保护效果的评价

Fine-mapping of immunodominant linear B-cell epitopes of the Staphylococcus aureus Isd A antigen using short overlapping peptides

目的系统筛选鉴定金黄色葡萄球菌Isd A的B细胞免疫显性表位,并评价其免疫保护效果。方法采用氨基酸步移策略,合成18 mer氨基酸重叠肽段。ELISA系统筛选鉴定Isd A的B细胞免疫显性表位肽段。免疫显性表位肽与KLH偶联后免疫Balb/c小鼠。分离免疫小鼠血清,ELISA检测免疫显性表位肽诱导的Ig G抗体水平,并测定表位肽诱导产生抗体所介导的调理吞噬作用。末次免疫后2周,经尾静脉感染S.aureus Newman,监测感染攻毒后各组的小鼠的生存率以及检测主要靶器官肾脏的细菌定植量。结果 Isd A49-66aa、Isd A91-108aa、Isd A217-234aa、Isd A259-276aa均能与Isd A抗血清产生强烈Ig G抗体反应。抗Isd A49-66aa、抗Isd A91-108aa、抗Isd A217-234aa和抗Isd A259-276aa血清均能与重组的Isd A产生较强的抗原抗体反应。与Alum组相比,免疫Isd A49-66aa、Isd A91-108aa、Isd A217-234aa、Isd A259-276aa后血清中的抗体能够显著增强抗体介导的中性粒细胞的调理吞噬作用(P<0.05),同时均显著提高了小鼠的生存率,并在感染后的第3天均显著降低肾脏的细菌定植(P<0.05)。结论成功鉴定出Isd A49-66aa、Isd A91-108aa、Isd A217-234aa、Isd A259-276aa为Isd A的B细胞免疫显性表位,显性表位肽Isd A49-66aa、Isd A91-108aa、Isd A217-234aa、Isd A259-276aa具有良好的免疫原性和免疫保护作用,可以作为金葡菌疫苗研究的重要候选表位疫苗组分。

This study aimed to map the immunodominant linear B-cell epitopes of the Staphylococcus aureus Isd A antigen and evaluate its immunoprotective effect in a mouse sepsis model. Synthetic overlapping peptideELISA was used to identify B-cell immunodominant Isd A epitopes, then Balb/c mice were immunized with theimmunodominant Isd A epitopes conjugated with KLH plus alum. The titers for immunodominant peptide antiserumreaction with Isd A were detected by ELISA; the opsonophagolytosis mediated by immunodominant Isd Aepitopes-induced antibody was detected. The survival rate and bacterial burden were analyzed after the micechallenged with S. aureus Newman. Data showed that immunodominant peptides Isd A49-66 aa, Isd A91-108 aa, Isd A217-234 aa,Isd A259-276 aainduced strong Ig G response to Isd A antiserum and the immunodominant peptide antiserum inducedstrong Ig G response to Isd A. Antiserum from the immunodominant peptides group significantly enhanced opsonizePimmunized with immunodominant peptides displayedhigher survival rates than the mice immunized withalum adjuvant, and the bacterial burdens in the kidneysof immunodominant peptides groups were significantlylower than those in the control group. In summary, thelinear B-cell epitopes of the Isd A in this study induce humor immunity and provide strong immune protectionagainst S. aureus infection, and could therefore be a promising vaccine candidate against S. aureus infection.