Molecular epidemiology of Plasmodium reichenowi

描述(申请人提供)：四种疟原虫导致人类疟疾，但恶性疟原虫是几乎所有人类疟疾死亡的罪魁祸首，每年导致100多万人死亡。然而，恶性疟原虫只与其他三种导致人类疟疾的疟原虫有较远的亲缘关系，而雷氏疟原虫是一种黑猩猩和大猩猩的寄生虫，在形态上与恶性疟原虫几乎没有区别。这种密切的亲缘关系已被分子系统学研究证实，赖氏疟原虫序列是恶性疟原虫遗传多样性研究中非常强大的外群，目的是在免疫选择压力下识别恶性疟原虫基因。这些生物之间的相似性对于了解恶性疟原虫致病机制的具体方面也很有意义。然而，尽管在20世纪初曾在野生黑猩猩的血液中多次观察到莱氏疟原虫，但科学界只获得了一个莱氏疟原虫分离物，而且CDC只保留了该分离物的少数冰冻试管。尽管可以获得一些有限的莱氏疟原虫基因组序列，这些基因组序列是用这些冷冻的种群产生的，但它主要由短片段组成，并被黑猩猩DNA严重污染。研究莱氏假单胞菌基因组的工作已经停止，直到找到莱氏假单胞菌DNA的新来源。因此，迫切需要新的莱氏肺孢子虫分离株来完成莱氏肺孢子虫基因组序列，以及用于比较遗传学和致病机理研究。为了获得新的莱氏并殖吸虫分离株，我们需要对发生雷氏并殖吸虫感染的野生黑猩猩种群进行鉴定，但还没有对野生黑猩猩中赖氏并殖吸虫感染的流行情况进行研究。黑猩猩的受保护状态对可用于此类研究的样本设置了严格的限制。这项应用的目的是使用一种独特的、非侵入性的和伦理敏感的检测方法来确定野生黑猩猩群落中莱氏肺吸虫感染的流行率。为了实现这一目标，我们开发了一种新的检测方法，使用尿样来检测疟原虫感染。我们建议将这一检测方法应用于一种独特的、预先存在的黑猩猩尿液和粪便样本的收集。我们的具体目标是：1.建立使用尿样检测既往疟原虫感染的敏感性。2.建立野生黑猩猩种群中莱氏肺吸虫的流行和分布情况。该项目有望首次确定野生黑猩猩中莱氏肺孢子虫感染的流行和分布。预计它还将直接导致对新的莱氏肺孢子虫分离株的鉴定，这些分离株将免费提供给疟疾研究界。这些分离株将是了解恶性疟原虫这一人类最重要的传染病杀手之一的进化和发病机制的杰出和独特的资源。恶性疟原虫每年导致100多万人死亡。为了了解恶性疟原虫的毒力和致病性，我们需要将其与近缘关系密切的寄生虫物种进行比较，但与恶性疟原虫关系最密切的寄生虫--一种名为雷氏疟原虫的黑猩猩寄生虫--只被分离过一次，而且只剩下有限的种群。这项应用的目的是确定野生黑猩猩中赖氏肺吸虫感染的流行率，并利用这一信息获得新的雷氏肺吸虫分离物，供疟疾研究界用于比较研究。

DESCRIPTION (provided by applicant): Four species of Plasmodium parasites cause malaria in humans, but Plasmodium falciparum is responsible for almost all human malaria mortality, killing more than 1 million people each year. However P. falciparum is only distantly related to the other three Plasmodium species that cause malaria in humans, whereas P. reichenowi, a parasite of chimpanzees and gorillas, is morphologically almost indistinguishable from P. falciparum. This remarkably close relationship has been confirmed by molecular phylogenetic studies and P. reichenowi sequences are therefore very powerful outgroups for P. falciparum genetic diversity studies aimed at identifying P. falciparum genes under immune selection pressure. The parallels between the organisms are also of great interest in understanding specific aspects of P. falciparum pathogenesis. However, although P. reichenowi was observed in the blood of wild chimpanzees on several occasions in the early 20th century, only a single P. reichenowi isolate has ever been obtained by the scientific community, and only a handful of frozen tubes of that isolate remain at CDC. Although some limited P. reichenowi genome sequence is available, generated using these frozen stocks, it consists largely of short fragments and is heavily contaminated with chimpanzee DNA. Work on the P. reichenowi genome has ceased until new sources of P. reichenowi DNA can be found. New P. reichenowi isolates are therefore urgently needed to complete the P. reichenowi genome sequence, as well as for comparative genetic and pathogenesis studies. In order to obtain new P. reichenowi isolates we need to identify wild chimpanzee populations in which P. reichenowi infections occur, but no prevalence study of P. reichenowi infection in wild chimpanzees has ever been carried out. The protected status of chimpanzees places strict limits on the samples that can be obtained for such studies. The objective of this application is to employ a unique, non-invasive and ethically sensitive assay to establish the prevalence of P. reichenowi infection in wild chimpanzee communities. To achieve this objective we have developed a novel assay that uses urine samples to detect Plasmodium infection. We propose to apply this assay to a unique and pre-existing collection of chimpanzee urine and fecal samples. Our specific aims are: 1. Establish the sensitivity of using urine samples to detect past Plasmodium infections. 2. Establish the prevalence and distribution of P. reichenowi in wild chimpanzee populations. This project is expected to define for the first time the prevalence and distribution of P. reichenowi infection in wild chimpanzees. It is also expected to lead directly to the identification of new P. reichenowi isolates that would be made freely available to the malaria research community. Such isolates would be an outstanding and unique resource for understanding of the evolution and pathogenesis of one of the most important human infectious disease killers, Plasmodium falciparum. Plasmodium falciparum parasites kill more than 1 million people each year. In order to understand P. falciparum virulence and pathogenicity we need to compare it with closely related parasite species, but the parasite most closely related to P. falciparum, a chimpanzee parasite called P. reichenowi, has only ever been isolated once and only limited stocks of that isolate remain. The objective of this application is to establish the prevalence of P. reichenowi infection in wild chimpanzees and to use that information to obtain new P. reichenowi isolates that would be made available to the malaria research community for comparative studies.