伯氏疟原虫丝/苏氨酸磷酸酶6抗血清对原虫有性阶段发育抑制作用的研究

目的:探讨伯氏疟原虫丝/苏氨酸磷酸酶6(PPP6)作为传播阻断疫苗候选抗原的可行性。方法:PCR扩增PPP6蛋白全长编码基因并克隆入pET32a(+)载体。IPTG诱导PPP6重组蛋白的表达,纯化后皮下免疫小鼠,收集抗PPP6免疫血清。ELISA和Westernblot检测抗PPP6免疫血清效价和特异性。实验观察免疫血清对配子体出丝、动合子和囊合子发育的影响。结果:成功诱导PPP6重组蛋白表达,抗PPP6免疫血清抗体效价为1∶3200;与对照组相比,抗PPP6免疫血清可显著抑制配子体出丝(P<0.0001);动合子数目和动合子转化率分别显著降低65.3%和42.07%(P<0.0001);抗PPP6血清1∶5倍稀释时,囊合子形成数量减少68.91%(P<0.0001)。结论:PPP6蛋白具有良好的免疫原性和抗原性。抗PPP6免疫血清具有明显的传播阻断效果。

Inhibitive effect of Plasmodium berghei serine/threonine protein phosphatase 6 on parasite sexual stage development

Objective: To determine the feasibility of selecting Plasmodium berghei serine/threonine phosphatase 6 (PPP6) as a transmission blocking vaccine candidate. Methods: The entire open reading frame of PPP6 was amplified by PCR and cloned into pET32a(+) vector.PPP6 recombinant protein (rPPP6) expression was induced by IPTG,purified,mice immunization,and anti-PPP6 serum was collected.The anti-PPP6 serum titer and specificity was detected by ELISA and Western blot.The inhibitory effect of anti-PPP6 sera on exflagellation,ookinete formation,conversion and oocysts formation was investigated. Results: The recombinant PPP6 protein was successfully expressed,with an antibody titer around 1 ∶3 200.Moreover,the exflagellation was significantly reduced on anti-PPP6 sera treatment ( P <0.000 1).Ookinete formation and conversion rate decreased 65.3% and 42.07% in anti-PPP6 sera treatment group,respectively ( P <0.000 1).Anti- PPP6 sera significantly inhibited oocysts formation by 68.82% at 1 ∶5 dilution ( P <0.000 1). Conclusion: PPP6 protein has immunogenicity and antigenicity.The anti-PPP6 immune serum had obvious transmission blocking activity.