MOLECULAR GENETICS OF LEISHMANIA

描述（由申请人提供）：锥虫原虫属利什曼原虫是导致一系列热带疾病的寄生虫，全世界有1000多万人受到这些疾病的折磨，根据特定的物种和感染者的免疫状态，可能是严重的或致命的。在巴西、非洲和地中海盆地，利什曼病是艾滋病患者常见的机会性感染，现在驻扎在中东的许多美军士兵都被感染。我们的研究目标是开发和应用新的方法来鉴定这种寄生虫用于进行其感染周期的基因。我们的前提是，强大的分子遗传工具的可用性，以及对寄生虫毒力重要基因的鉴定，将从根本上提高我们制定改进控制策略的能力。在以前的研究中，人们成功地开发了强大的新方法，以各种方式操纵寄生虫基因组，并应用了现代“基因组方法”，如表达谱分析和比较基因组杂交。这些研究相当成功，但发现的令人兴奋的基因数量远远超过了我们处理它们的方法的能力。幸运的是，我们取得了两个突破。首先，我们发现RNA干扰途径在巴西利什曼原虫的物种复合体中是有效的，尽管它在其他利什曼原虫物种中丢失了。我们发现，RNAi途径可以方便、快速地适应于寄生虫内基因功能的功能评估。这种方法的特点使我们能够迅速扩大和测试许多基因，并确定它们在毒性中的作用。在一些利什曼原虫物种中这一途径的缺失是具有挑战性的，我们的研究表明，这与与人类疾病密切相关的感染周期的重要变化有关。我们的第二个突破是证明有可能通过实验产生利什曼原虫的杂交。这些新方法将使我们能够更好地描述利什曼原虫基因组的功能组织，并通过研究影响动物毒力的性状的遗传，以以前不可能的方式确定与人类疾病相关的基因。

DESCRIPTION (provided by applicant): Parasites of the trypanosomatid protozoan genus Leishmania are responsible for a spectrum of tropical diseases that afflict more than 10 million people worldwide, and depending on the specific species and immune status of the infected person, can be severe or fatal. In Brasil, Africa and the Mediterranean basin, leishmaniasis is a common opportunistic infection in AIDS patients, and many US soldiers now stationed in the Middle East have been infected. The goal of our research is to develop and apply new methods for the identification of genes used by this parasite to carry out its infectious cycle. Our premise is that the availability of powerful molecular genetic tools, and identification of genes important to parasite virulence, will radically advance our ability to develop improved control strategies. In previous studies powerful new methods were successfully developed for manipulating the parasite genome in a variety of ways, and modern 'genomic approaches' such as expression profiling and comparative genome hybridization applied. These studies were quite successful, but the number of exciting genes identified far exceeded the ability of our methods to tackle them. Fortunately we achieved two breakthroughs. First we showed that the RNA interference pathway was functional in species belong to the L. braziliensis species complex, despite its loss in other Leishmania species. We showed that the RNAi pathway could be used to conveniently and rapidly adapted to functionally assess gene function within the parasite. The features of this methodology promise to allow us to rapidly scale up and test many genes, and determine their role in virulence. The loss of this pathway in some Leishmania species is provocative and our studies suggest this is linked to important changes in the infectious cycle of great relevance to human disease. Our second breakthrough was the demonstration that it was possible to generate crosses of Leishmania experimentally. These new approaches will allow us to better describe the functional organization of the Leishmania genome, and by studying the inheritance of traits affecting animal virulence, to identify genes relevant to human disease in ways not previously possible.