Genetics of Cryptosporidium parvum

隐孢子虫是一种机会性肠道原生动物寄生虫，感染艾滋病患者， 其他免疫力低下的人。由于缺乏有效的药物，隐孢子虫病可能会持续存在， 特别是在免疫缺陷个体中。控制这种寄生虫毒力表型的基因是 未知的，但他们的鉴定是必要的，以指导药物的开发，以更好地了解 发病机制，并改善基因组的注释。本次修订的长期目标 应用是应用遗传学方法来定位控制特定表型的基因。与链接相反 分析中使用的第一个融资期，我们建议应用连锁集团选择，一个战略， 不需要克隆后代品系。相反，控制可选择性状的遗传基因座通过以下方式鉴定： 将两个表型不同的C.将后代群体暴露于一轮或多轮的 选择和批量定量基因分型后代群体。基于明确的表型 亲本系之间的差异，我们将选择早期卵囊分化，卵囊抗性， 漂白剂和寄生虫对干扰素的感染性基因敲除小鼠在选择过程中，携带等位基因的寄生虫 将从后代中消除所选择的表型。由于减数分裂 来自敏感亲本的等位基因的比例将随着与基因座的距离而增加。 在选择下，产生一个选择谷。这些谷将通过定量基因分型来鉴定， 选择的后代群体和对照后代群体具有一组单核苷酸多态性。本研究将 与疟疾遗传学家Paul Hunt博士合作进行，他开发并成功应用了 连锁群选择在疟疾抗药性研究中的应用。 具体目标： 1.将表型不同的微小隐孢子虫分离株进行杂交，并使用连锁群选择， 确定控制特定表型的染色体区域。未克隆重组体群体 来自表型不同的亲本系之间杂交的寄生虫将经受 选择压力，并使用一组100个SNP标记进行定量基因分型。采空区 选择将通过迭代重组和选择逐渐解决。 2.识别选择谷内控制选择下表型的基因，并识别突变 对表型负责。位于选择谷内的一组紧密间隔的SNP将被 按比例测序以鉴定选择下的基因和突变。突变将被映射到 C. hominis，C. muris和更远的相关apicomplexan来识别那些最有可能的 影响蛋白质功能或转录调控。

Cryptosporidium parvum is an opportunistic intestinal protozoan parasite which infect AIDS patients and other immune-compromised individuals. For lack of effective drugs, cryptosporidiosis may become persistent, particularly in immunodefficient individuals. Genes controlling virulence phenotypes in this parasite are unknown, but their identification is needed to guide the development of drugs, to better understand pathogenesis, and to improve the annotation of the genome. The long-term objective of this revised application is to apply genetic methods to map genes controlling specific phenotypes. In contrast to linkage analysis used in the first funding period, we propose to apply Linkage Group Selection, a strategy which does not require the cloning of progeny lines. Instead, genetic loci controlling selectable traits are identified by crossing two phenotypically distinct C. parvum lines, exposing progeny populations to one or multiple rounds of selection, and quantitatively genotyping progeny populations in bulk. Based on well-defined phenotypic differences between the parental lines, we will select for early oocyst differentiation, oocyst resistance to bleach, and parasite infectivity to interferon-? knock-out mice. During selection, parasites carrying alleles conferring the phenotype selected against will be eliminated from the progeny. As a result of meiotic recombination the proportion of alleles from the sensitive parent will increase with distance from the locus under selection, giving rise to a ¿selection valley¿. Such valleys will be identified by quantitatively genotyping a selected and a control progeny population with a panel of single nucleotide polymorphisms. This study will be performed in collaboration with Dr Paul Hunt, a malaria geneticist who has developed and successfully applied Linkage Group Selection to the study of malaria drug resistance. Specific Aims: 1. Cross phenotypically distinct isolates of Cryptosporidium parvum and use Linkage Group Selection to identify chromosomal regions controlling specific phenotypes. Populations of uncloned recombinant parasites derived from crosses between phenotypically distinct parental lines will be subjected to selective pressure and quantitatively genotyped using a set of 100 SNP markers. Regions under selection will be progressively resolved by iterative recombination and selection. 2. Identify genes within a selection valley controlling the phenotype under selection and identify the mutations responsible for the phenotype. A set of closely spaced SNPs located within the selection valley will be proportionally sequenced to identify genes and mutations under selection. Mutations will be mapped to the genome of C. hominis, C. muris and more distantly related apicomplexan to identify those most likely to affect protein function or transcriptional regulation.