约氏疟原虫丝/苏氨酸磷酸酶6免疫血清对原虫有性阶段发育抑制作用的研究  

INHIBITION ON THE DEVELOPMENT OF SEXUAL PARASITES OF PLASMODIUM YOELII DUE TO SERINE-THREONINE PROTEIN PHOSPHATASE 6 ANTIBODY

作　　者：于园超 洪明阳 周丹 朱晓彤 YU Yuan-Chao;HONG Ming-Yang;ZHOU Dan;ZHU Xiao-Tong(Department of Immunology,College of Basic Medical Science,China Medical University,Shenyang 110122,Liaoning,China)

机构地区：[1]中国医科大学基础医学院免疫学教研室,辽宁沈阳110122

出　　处：《寄生虫与医学昆虫学报》2020年第2期65-72,共8页Acta Parasitologica et Medica Entomologica Sinica

基　　金：国家自然科学基金项目(No.81971961)。

摘　　要：为探讨约氏疟原虫丝/苏氨酸磷酸酶6(Plasmodium yoelii Protein Phosphatase 6,PyPP6)作为传播阻断疫苗候选抗原的可行性,采用PCR扩增PP6优势抗原表位并克隆入pET32a(+)载体,诱导PyPP6重组蛋白(rPyPP6)表达与纯化,免疫小鼠获取抗-PyPP6免疫血清(anti-PyPP6)。采用ELISA和Western Blot方法检测anti-PyPP6血清效价和特异性,IFA检测PyPP6蛋白在约氏疟原虫各期定位,体内外实验检测anti-PyPP6血清对雄配子出丝、动合子形成和卵囊发育的影响。结果显示,成功诱导rPyPP6表达,anti-PyPP6血清抗体滴度为1:128000,PyPP6部分定位于约氏疟原虫质膜。与对照组相比,anti-PyPP6免疫血清1∶5倍稀释可显著抑制61.5%配子体出丝,动合子数目和动合子转化率分别降低了75%和19.7%,卵囊形成数量减少了35%。结果表明,PyPP6重组蛋白具有良好的免疫原性和抗原性,抗-PP6免疫血清具有显著的传播阻断效果。To investigate the feasibility of Plasmodium yoelii serine/threonine phosphatase 6(PyPP6)as a candidate antigen for transmission blocking vaccine(TBV),PP6 dominant antigen epitopes were amplified by PCR and cloned into pET32 a(+)plasmid to induce the expression and purification of recombinant PP6 protein(rPyPP6),obtaining anti-PyPP6 immune serum(anti-PyPP6)in immune mice.ELISA and Western Blot methods were used to detect the titer and specificity of anti-PyPP6 serum,IFA was used to detect the localization of PyPP6 protein in each stage of P.yoelii,in vivo and in vitro experiments were carried out to detect the effect of anti-PyPP6 serum on the development of male gametes exflagellation,ookinete formation and oocyst development.The results showed that rPyPP6 expression was successfully induced,anti-PyPP6 serum antibody titer was 1∶128000,and the PyPP6 part was localized in the P.yoelii plasma membrane.Compared with the control group,1∶5 times dilution of anti-PyPP6 immune serum inhibited significantly 61.5%gametophyte exflagellation,the number and conversion rate of ookinete decreased by 75%and 19.7%respectively(P<0.01),and the number of oocyst formation decreased by 35%(P<0.001).The results suggested that rPyPP6 recombinant protein had good immunogenicity and antigenicity.The anti-PyPP6 immune sera showed a significant transmission blocking activity.

关 键 词：约氏疟原虫 丝/苏氨酸磷酸酶6 传播阻断疫苗 

分 类 号：R382.1[医药卫生—医学寄生虫学]