Genetic Analysis of Host Specificity in Schistosoma mansoni

描述(由申请人提供)：寄生虫对特定宿主表现出强烈的特异性，但这种宿主特异性的遗传基础尚不清楚。曼氏血吸虫是感染人类的三个医学上重要的血吸虫物种之一，出色的实验工作证明，蜗牛中间宿主的化学识别和蜗牛内寄生虫的生存都遵循简单的孟德尔遗传模式。虽然埃及寄生虫对共域蜗牛Biomphalaria Alexandrina表现出强烈的化学识别能力，但巴西寄生虫没有表现出特异性，甚至会被非媒介钉螺宿主所吸引。同样，虽然埃及和巴西的寄生虫都会在共生的蜗牛宿主中感染和繁殖，但F1杂交种只在南美光滑蜗牛中发育。新的分子工具使我们能够确定决定宿主-寄生虫系统中宿主特异性的寄生虫基因，为了解寄生虫和钉螺媒介之间的关键分子相互作用提供了一种手段。利用R21的资助，我们(A)建立了曼氏血吸虫5 cM连锁图谱，(B)通过鉴定一个强QTL(LOD=21)，证明了连锁图谱的有效性。我们现在建议利用该遗传图谱，与最近公布的曼氏血吸虫基因组序列一起，识别曼氏血吸虫-Biomphalaria系统中宿主专一性的基因组区域(S)。宿主特异性涉及两个部分：(A)毛虫对钉螺的化学定位；(B)血吸虫幼虫在钉螺体内的穿透和克隆性增殖。对于这两个特征，我们将在巴西和埃及曼氏沙门氏菌之间进行遗传杂交，使用带有单一毛虫的蜗牛感染在蜗牛中产生单一基因感染。我们将量化单个F2毛虫对光面芽孢杆菌和亚历山大毛虫的化学识别行为，然后使用曼氏丝虫基因组上间隔约4 cM(2Mb)的SNPs对单个毛虫进行基因分型，以确定这一特征的QTL。使用经典的连锁作图方法不可能对钉螺宿主内的生存和克隆增殖进行遗传作图，因为不能在钉螺内生长的寄生虫不能进行基因分型。因此，我们将使用由疟疾和酵母菌遗传学家开发的极端QTL(X-QTL)方法来检测从光肩牛或亚历山大钉螺中出现的F2尾蚴的等位基因频率。在致病基因座上，我们预计来自埃及曼氏血吸虫亲本的等位基因在亚历山大芽孢杆菌产生的F2尾蚴中相对于从光滑假单胞菌产生的等位基因过多。因此，座位特异性偏离正常的孟德尔分离可以进行QTL定位。有了这两个性状的精细定位的QTL区域，我们将使用基因功能的RNAi干扰或基于逆转录病毒的转染来辅助鉴定致病基因座。了解曼氏血吸虫-Biomphalaria系统中宿主特异性的遗传和分子基础对于旨在扰乱寄生虫生命周期这一阶段的控制工作至关重要，并为了解最重要的人类寄生虫宿主特异性的进化提供了关键。

DESCRIPTION (provided by applicant): Parasites characteristically show strong specificity to particular hosts, but the genetic basis for this host specificity is poorly understood. In Schistosoma mansoni, one of three medically important schistosome species infecting humans, elegant experimental work demonstrates that both chemical recognition of the intermediate snail host and survival of parasites within snails follow a simple Mendelian pattern of inheritance. While Egyptian parasites show strong chemical recognition of the sympatric snail Biomphalaria alexandrina, Brazilian parasites show no specificity and are attracted to even non-vector snail hosts. Similarly, while both Egyptian and Brazilian parasites infect and proliferate in their sympatric snail hosts, F1 hybrids develop only in S. American B. glabrata snails. New molecular tools allow us to determine the parasite genes that determine host specificity in this host-parasite system, providing a means to understand key molecular interactions between parasites and snail vector. Using R21 funding we have (a) developed a 5 cM linkage map for S. mansoni, and (b) demonstrated the utility of linkage mapping by identifying a strong QTL (LOD = 21) for oxamniquine resistance to a short region of chr 6. We now propose to exploit the genetic map, together with the recently published genome sequence of S. mansoni identify the genome region(s) that underlie host specificity in the S. mansoni - Biomphalaria system. Host specificity involves two components (a) chemical location of snails by miracidia and (b) penetration and clonal proliferation of schistosome larvae within snails. For both traits, we will conduct genetic crosses between Brazilian and Egyptian S. mansoni, using snail infections with single miracidia to generate single genotype infections in snails. We will quantify chemical recognition behavior of single F2 miracidia to both B. glabrata and B. alexandrina, and then genotype individual miracidia using SNPs spaced at ~4 cM (2Mb) intervals across the S. mansoni genome to identify QTLs for this trait. Genetic mapping of survival and clonal proliferation within the snail host is not possible using classical linkage mapping methods, because parasites that do not grow within snails cannot be genotyped. We will therefore use extreme QTL (X-QTL) methods, developed by malaria and yeast geneticists, to examine allele frequencies of F2 cercariae emerging from either B. glabrata or B. alexandrina snails. At the causative loci, we expect alleles from the Egyptian S. mansoni parent to be overrepresented in F2 cercariae emerging from B. alexandrina relative to those emerging from B. glabrata. Hence locus specific deviation from normal Mendelian segregation allows QTL location. Having fine mapped QTL regions for both traits, we will use RNAi disruption of gene function or retroviral based transfection to aid identification of causative loci. Understanding the genetic and molecular basis of host specificity in the S. mansoni - Biomphalaria system is critical for control efforts that aim to disrupt this step in the parasite lifecycle and provides a key to understanding evolution of host specificity in the most important of the human helminth parasites.