Antiviral Pathways in Disease Vector Mosquitoes

描述（由申请方提供）：节肢动物传播的病毒（虫媒病毒）对公共卫生构成了持续的全球性挑战。蚊媒病毒在自然界中的维持需要一个生物传播周期，该周期涉及易感脊椎动物和蚊子宿主中的交替病毒复制。对于自然界中的虫媒病毒，与蚊子感染相关的适应度成本非常小是必要的。我们最近证明了RNA干扰（RNAi）在甲病毒在蚊子载体中建立持久的非致病性感染的机制中的重要性，表明在没有基于RNAi的调节的情况下，蚊子无法在虫媒病毒感染中存活。然而，对蚊子RNAi基因进行的直接实验很少。我们已经描述了埃及伊蚊的转基因株的产生和验证，该转基因株“感测”RNAi途径的状态，通过该转基因株，我们已经表明基因Dcr-2和Ago-2，但不是Ago-3，对于Ae中的RNAi是关键的。埃及人。我们假设RNAi途径的遗传变异直接影响蚊子被虫媒病毒感染和传播的能力。在具体目标1中，我们将使用我们的转基因“传感器菌株”来确定蚊子基因在RNAi的调控和执行中的参与。在具体目标2中，我们将比较伊蚊属内各种蚊子基因的一级核苷酸序列，并确定是否有任何进化选择压力作用于参与RNAi的基因。在具体目标3中，我们将确定RNAi基因中的遗传变异性对伊蚊对医学上重要的虫媒病毒的载体能力的作用。对蚊子RNAi基因的详细了解，以及这些基因中遗传变异对病毒传播的作用，将有助于更好地评估基于RNAi的遗传控制策略的可行性，并使我们能够更准确地确定新的和新兴的虫媒病毒传播到新地区的风险，特别是在美国。

DESCRIPTION (provided by applicant): Arthropod-borne viruses (arboviruses) constitute a persistent and worldwide challenge to public health. Maintenance of mosquito-borne viruses in nature requires a biological transmission cycle that involves alternating virus replication in a susceptible vertebrate and mosquito host. For arboviruses in nature, it is imperative that very little fitness cost be associated with infection of the mosquito. We recently demonstrated the importance of RNA interference (RNAi) in the mechanism by which alphaviruses establish a persistent, nonpathogenic infection in the mosquito vector, showing that in the absence of RNAi-based modulation, mosquitoes do not survive arboviral infection. However, very little direct experimentation has been done on mosquito RNAi genes. We have described the generation and validation of a transgenic strain of Aedes aegypti that "senses" the status of the RNAi pathway, through which we have shown that the genes Dcr-2 and Ago-2, but not Ago-3, are critical for RNAi in Ae. aegypti. We hypothesize that genetic variability in the RNAi pathway directly affects the ability of mosquitoes to become infected by, and transmit, arboviruses. In specific aim 1, we will use our transgenic "sensor strain" to determine the involvement of mosquito genes in the regulation and execution of RNAi. In specific aim 2, we will compare primary nucleotide sequences of various mosquito genes within the genus Aedes, and determine whether any evolutionary selective pressures are acting on genes involved in RNAi. In specific aim 3, we will determine the role of genetic variability in RNAi genes on the vector competence of Aedes mosquitoes for medically important arboviruses. A detailed understanding of mosquito RNAi genes, and the role played by genetic variation in those genes on virus transmission, will facilitate better evaluations of the feasibility of RNAi-based genetic control strategies, and allow us to more accurately determine the risks of new and emerging arboviruses spreading to new areas, particularly within the U.S.