Molecular Aspects of Tsetse and Trypanosome Transmission

这项申请是关于预防非洲锥虫病，这是非洲最被忽视的疾病之一 由采采蝇传播的寄生非洲锥虫引起。缺乏有效的工具来遏制 哺乳动物中的感染和动物宿主的存在使得有必要进行病媒控制以对抗疾病。 我们将研究影响传播动力学的采采蝇-锥虫相互作用。用于传输到 锥虫首先在中肠（MG）建立感染，然后移动到苍蝇的口器， 进入唾液腺（SG）并定植。主要的障碍，消除寄生虫从大多数 苍蝇发生在MG。我们已经表明，寄生虫介导的载体的生理操纵过程 在感染早期短暂降低中肠屏障的完整性，使寄生虫绕过围食 基质（PM）屏障。在这个操纵过程的核心是哺乳动物寄生虫表面蛋白，变体 表面糖蛋白（VSG），在感染过程的早期脱落到肠腔中， 采采蝇的PM合成通过microRNA（miR-275）起作用。通过人为操作， 这一过程再次使感染MG的寄生虫重新进入管腔以定殖SG。我们将使用 跨学科研究计划调查： 1.降低PM功效的机制和寄生虫VSG蛋白的不同组分， 在感染过程的早期负责这种干扰。我们还将研究寄生虫 当寄生虫从MG迁移到SG时， 传输我们将进行载体和寄生虫转录组学分析，以发现潜在的介导因子， 生物体内部的对话 2.舌蝇microRNA（miR 275）在PM合成中的作用（通过鉴定其下游分子靶点） 通过转录组和Riboseq分析，并通过使用双荧光素酶测定在一个 S2细胞系，并通过免疫共沉淀试验。 3.采采蝇-寄生虫在野外自然感染中的相互作用， 在实验室中观察，并确定PM修改对建立共同 感染多种寄生虫物种和菌株。利用实地苍蝇，我们将确定 寄生虫传播过程，以评估PM屏障的流行病学意义。 总的来说，我们的研究将提供适应性和操纵性过程的基础知识， 影响病媒能力和重要病媒的疾病传播，并将揭示潜在的目标 用于通过传播阻断策略或副转基因应用进行干扰以减少疾病。

This application is on prevention of African Trypanosomiasis, one of the most neglected diseases of Africa caused by parasitic African trypanosomes transmitted by tsetse. The absence of effective tools to curb infections in the mammal and the presence of animal reservoirs necessitate vector control to combat disease. We will investigate tsetse-trypanosome interactions that influence transmission dynamics. For transmission to occur, trypanosomes first establish infections in the midgut (MG) and then move to the fly's mouthparts to access and colonize the salivary glands (SG). The major barrier that eliminates parasites from the majority of flies occurs in the MG. We have shown that a parasite mediated manipulative process of vector's physiology transiently reduces midgut barrier integrity early in the infection to enable the parasites to bypass the peritrophic matrix (PM) barrier. At the core of this manipulative process is the mammalian parasite surface proteins, Variant Surface Glycoproteins (VSGs), shed into the gut lumen early in the infection process, which interfere with tsetse's PM synthesis acting through a microRNA (miR-275). Loss of PM integrity through a manipulative process again enables MG infecting parasites to re-enter into the lumen to colonize SG. We will use an interdisciplinary research plan to investigate: 1. The mechanisms that reduce PM efficacy and the different components of the parasite VSG protein that are responsible for this interference early in the infection process. We will also investigate the parasite components that enable PM reduction later in the infection process as parasites migrate from MG to SGs for transmission. We will perform vector and parasite transcriptomic profiling to discover potential mediators of the intra-organismal dialogue. 2. The role of the tsetse microRNA (miR275) in PM synthesis by identifying its downstream molecular targets through transcriptome and Riboseq profiling and by validating these targets using a dual-luciferase assay in a S2 cell line and through co-immunoprecipation assays. 3. Tsetse-parasite interactions in natural infections in the field to validate the parasite-vector dialogue we observe in the laboratory, and to determine the influence of PM modification on establishment of co- infections with multiple parasite species and strains. Using field flies, we will determine the course of parasite transmission processes to assess the epidemiological significance of PM barriers. Collectively, our studies will provide fundamental knowledge on adaptive and manipulative processes that influence vector competence and disease transmission in an important vector and will reveal potential targets for interference by transmission blocking strategies or paratransgenic applications to reduce disease.