Evolution of the Transmission-Blocking Vaccine Candidates Pvs28 and Pvs25 in <i>Plasmodium vivax</i>: Geographic Differentiation and Evidence of Positive Selection

Transmission-blocking (TB) vaccines are considered an important tool for malaria control and elimination. Among all the antigens characterized as TB vaccines against Plasmodium vivax, the ookinete surface proteins Pvs28 and Pvs25 are leading candidates. These proteins likely originated by a gene duplication event that took place before the radiation of the known Plasmodium species to primates. We report an evolutionary genetic analysis of a worldwide sample of pvs28 and pvs25 alleles. Our results show that both genes display low levels of genetic polymorphism when compared to the merozoite surface antigens AMA-1 and MSP-1; however, both ookinete antigens can be as polymorphic as other merozoite antigens such as MSP-8 and MSP-10. We found that parasite populations in Asia and the Americas are geographically differentiated with comparable levels of genetic diversity and specific amino acid replacements found only in the Americas. Furthermore, the observed variation was mainly accumulated in the EGF2- and EGF3-like domains for P. vivax in both proteins. This pattern was shared by other closely related non-human primate parasites such as Plasmodium cynomolgi, suggesting that it could be functionally important. In addition, examination with a suite of evolutionary genetic analyses indicated that the observed patterns are consistent with positive natural selection acting on Pvs28 and Pvs25 polymorphisms. The geographic pattern of genetic differentiation and the evidence for positive selection strongly suggest that the functional consequences of the observed polymorphism should be evaluated during development of TBVs that include Pvs25 and Pvs28.传播阻断（transmission-blocking，TB）疫苗被视为疟疾防控与消除的重要工具。在已被鉴定为抗间日疟原虫（Plasmodium vivax）TB疫苗的各类抗原中，动合子表面蛋白Pvs28与Pvs25是极具潜力的候选靶点。这类蛋白大概率起源于已知疟原虫物种向灵长类动物辐射演化前发生的一次基因复制事件。本研究针对全球范围内的pvs28与pvs25等位基因样本开展进化遗传学分析。研究结果显示，相较于裂殖子表面抗原AMA-1与MSP-1，这两个基因的遗传多态性水平较低；但这两种动合子抗原的多态性程度，可与MSP-8、MSP-10等其他裂殖子抗原相当。研究发现，亚洲与美洲的疟原虫种群存在地理分化，二者遗传多样性水平相当，且仅在美洲种群中发现特异性氨基酸替换。此外，两种蛋白中的遗传变异主要富集在间日疟原虫的EGF2样与EGF3样结构域中。这一变异模式在食蟹猴疟原虫（Plasmodium cynomolgi）等其他亲缘关系较近的非人灵长类疟原虫中同样存在，提示该结构域可能具有重要的功能意义。此外，通过一系列进化遗传学分析验证，观测到的变异模式与作用于Pvs28和Pvs25多态性的正向自然选择相符。遗传分化的地理分布特征以及正向选择的相关证据，强烈提示在开发包含Pvs25与Pvs28的TB疫苗时，应当评估观测到的多态性所带来的功能影响。