Genome-wide studies to identify markers of artemisinin-resistant malaria

描述（由申请人提供）： 高效青蒿素类抗疟药物在全世界使用，为减轻许多地区的疟疾负担作出了贡献。柬埔寨西部出现了对青蒿素有抗药性的恶性疟原虫疟疾，其潜在的蔓延可能会使最近在防治疟疾方面取得的成果付诸东流，并使重新开展全球根除疟疾运动的计划流产。由于缺乏衡量其传播程度和方向的工具，目前遏制青蒿素抗药性疟疾的努力受到阻碍。目前，只有艰苦的临床试验才能可靠地测量耐药性。检测青蒿素耐药性标记的分子测定将是一种非常有价值的监测工具，但耐药性的遗传基础尚不清楚。我们提出了全基因组关联研究（GWAS），旨在确定可用于跟踪和遏制青蒿素耐药寄生虫在全球传播之前的分子标记。这项工作将在两个目标中完成。首先，我们将使用一组密集的单核苷酸多态性（SNP）来绘制与青蒿素耐药性相关的恶性疟原虫基因组区域。为了实现这一点，我们将使用已建立的SNP调用管道，从在青蒿素功效的已完成试验期间收集的寄生虫中的短读段测序数据中确定约421，000个SNP处的基因型，并且我们将估计寄生虫基因型与青蒿素治疗后将寄生虫血症减少一半所需的时间量（即寄生虫清除半衰期）之间的关联。将在两个独立的样本集中估计关联性，并将进行荟萃分析以验证与青蒿素耐药性相关的基因组区域。在第二个目标中，我们将识别和优先考虑目标1中确定的基因组区域内的候选基因，并使用下一代测序数据来识别高优先级基因内的多态性及其与寄生虫清除半衰期的关联。如果成功，该项目将导致开发一种快速测定法，在随后的候选基因关联研究中检测青蒿素耐药性的候选标志物，以及开始对候选基因进行功能表征的数据，以更好地了解耐药性的机制。

DESCRIPTION (provided by applicant): Highly effective artemisinin-based antimalarial drugs are used worldwide and have contributed to reductions in the malaria burden in many areas. Artemisinin-resistant Plasmodium falciparum malaria has emerged in western Cambodia and its potential spread threatens to reverse recent gains against malaria and to abort plans for a renewed global eradication campaign. Ongoing efforts to contain artemisinin-resistant malaria are hampered by the lack of tools to gauge the extent and direction of its spread. Presently, only laborious clinical trials can reliably measure resistance. A molecular assay to detect markers of artemisinin resistance would be a highly valuable surveillance tool, but the genetic basis of resistance is unknown. We propose genome-wide association studies (GWAS) that aim to identify molecular markers that can be used to track and contain artemisinin-resistant parasites before they spread globally. The work will be accomplished in two aims. First, we will use a dense panel of single nucleotide polymorphisms (SNPs) to map regions of the P. falciparum genome that are associated with artemisinin resistance. To accomplish this, we will use an established SNP- calling pipeline to determine genotypes at ~421,000 SNPs from short-read sequencing data in parasites collected during completed trials of artemisinin efficacy, and we will estimate the association between parasite genotypes and the amount of time it takes to reduce parasitemia by half following artemisinin treatment (i.e. parasite clearance half-life). Associations will be estimated in two independent sample sets, and meta-analysis will be performed to validate genomic regions associated with artemisinin resistance. In the second aim, we will identify and prioritize candidate genes within genomic regions identified in Aim 1 and use next-generation sequencing data to identify polymorphisms within high-priority genes and their association with parasite clearance half-life. If successful, this project will result in the development of a rapid assay to detect candidate markers of artemisinin resistance in subsequent candidate gene association studies, as well as data to begin the functional characterization of candidate genes to better understand the mechanisms underlying resistance.