Exploring the adaptive role of genomic instability in Trypanosoma cruzi.

克氏锥虫是一种引起恰加斯病的原生动物，恰加斯病是一种致命的寄生虫病，由一种名为接吻虫的昆虫媒介传播。在拉丁美洲，克鲁兹旋毛虫感染了700万人，其中1700万人被认为有感染的风险。感染者向欧洲和北美的迁移已经将恰加斯病传播到了其传统范围之外，在全球范围内造成了约50万例额外病例。在欧洲和美国，存在先天性和输血传播的风险，感染者对公共卫生服务构成重大挑战。恰加斯病是拉丁美洲最重要的寄生虫，每年导致1.2万人死亡。为了提供背景资料，该地区的疟疾死亡人数只占这个数字的一小部分(每年200-400人)。查加斯病患者感染克氏毛滴虫是终生的。药物治疗是有限的，在清除寄生虫感染方面往往无效，在缓解令人衰弱的慢性症状(心脏病、胃肠道异常)方面几乎总是无效。尽管恰加斯病对人类健康有影响，但与其他相关的人类寄生虫-布氏毛滴虫(昏睡病的病原体)和利什曼原虫(利什曼病的病原体)相比，人们对其生物学了解相对较少。关于克氏锥虫如何适应环境压力，存在着重要的知识空白。解决这些差距可以揭示这种寄生虫如何避免宿主免疫，从而在宿主中建立持续感染，以及它如何在药物治疗中幸存下来。我们联盟的成员和其他人对克鲁兹毛滴虫分离株的DNA测序显示，基因组处于不断重排的状态。克鲁兹毛滴虫染色体的数量、大小、拷贝数和组成在密切相关的分离株之间以及根据我们现在提供的试点数据在不同时间点采样的单个人类感染病例之间可能有很大差异。这种基因组重排的适应价值可能是理解和解决克氏锥虫生物学许多棘手方面的关键。在这项建议中，我们利用基因组学、基因操纵、动物疾病模型以及世界级研究团队的进展，了解克氏锥虫基因组重排如何支持哺乳动物宿主的长期生存以及寄生虫对一线和下一代药物的抗药性。利用单细胞基因组学，我们将把基因组重排与耐药性联系起来，然后通过基因操作，试图中断使这种重排发生的机制。然后，我们将进行一系列深刻的实验，将寄生虫基因组重排与小鼠模型在免疫压力下的存活联系起来，并通过对厄瓜多尔恰加斯病患者队列的观察来证实这一点。哺乳动物的实验将重点放在寄生虫细胞表面表达的基因家族之间的重新排列上。将跟踪具有免疫能力的宿主内寄生虫基因组变化的时间动态，并与没有正常运行的免疫系统的情况进行比较，以检测通过抗原转移而避免免疫的迹象。本研究计划中提出的实验是改进药物设计的重要基础科学先驱，也是未来托克氏锥虫疫苗开发的实验室基础，最终改善数百万受恰加斯病影响的人的健康结果。

Trypanosoma cruzi is a protozoan that causes Chagas disease, a fatal parasitic disease spread by an insect vector called a kissing bug. T. cruzi infects seven million people in Latin America and seventeen million are thought to be at risk of infection. Migration of infected individuals to Europe and North America has transferred Chagas disease outside its traditional range, accounting for c.500,000 additional cases globally. In Europe and America, a risk of congenital and transfusional transmission exists, and infected individuals represent a significant challenge for public health services. Chagas disease is the most important parasitic in Latin America, killing 12,000 people every year. To provide context, malaria in the region kills a fraction of that number (200-400 annually). Infection with T. cruzi in Chagas disease patients is life-long. Drug treatments are limited, often ineffective at clearing parasite infection, and almost always ineffective at alleviating debilitating chronic symptoms (heart disease, GI tract abnormalities). Despite the impact of Chagas disease on human health, relatively little is known about its biology by comparison to other related human parasites - T. brucei (agent of sleeping sickness) and Leishmania (agent of Leishmaniasis). Important knowledge gaps exist around how T. cruzi adapts to environmental stressors. Addressing theses gaps could shed light on how the parasite avoids host immunity to establish persistent infections in its host, as well as how it survives drug treatment. DNA sequencing of T. cruzi isolates by members of our consortium and others reveals a genome in a constant state of re-arrangement. The number, sizes, copy number and composition of T. cruzi chromosomes can vary substantially between closely related isolates, as well as, based on pilot data we now present, from individual human infections sampled at different time points. The adaptive value of such genomic re-arrangements may hold the key to understanding, and addressing, many intractable aspects of T. cruzi biology. In this proposal we leverage advances in genomics, genetic manipulation, animal disease models, as well as a world-class research team to understand how T. cruzi genomic re-arrangements may underpin long term survival in the mammalian host as well as parasite resistance to frontline and next generation drugs. Using single cell genomics, we will link genomic re-arrangements to drug resistance and then, via genetic manipulation, attempt to interrupt the machinery that enables such re-arrangements to occur. We will then undertake a series of incisive experiments to link parasite genomic re-arrangements to survival under immune pressure in mouse models and confirm these via observations in a cohort of Ecuadorian Chagas disease patients. Experiments in mammals will focus on re-arrangements among families of genes expressed on the parasite cell surface. The temporal dynamics of parasite genomic changes within immune-competent hosts will be followed and compared to the those in the absence of a functioning immune system to detect signatures of immune avoidance via antigenic shift.The experiments proposed in this research program are vital basic science precursors to improved drug design and the lab groundwork for future T. cruzi vaccine development, ultimately improving health outcomes for the millions affected by Chagas disease.