GENETIC DETERMINANTS OF EPIDEMIC ALPHAVIRUS TRANSMISSION

蚊子传播的病原体是人类和动物疾病的重要原因。从历史上看，破坏病原体传播周期的尝试依赖于通过使用杀虫剂或脊椎动物宿主免疫来减少病媒种群。不幸的是，杀虫剂耐药性的发展和病媒控制规划的崩溃需要制定控制病媒传播病原体的替代战略。通过在分子水平上阐明病毒/媒介相互作用的要素，旨在减少媒介容量的新方法可能是实现这一目标的最有效方法之一，从而控制节肢动物传播的病毒性疾病的传播。甲病毒是研究得最好的节肢动物传播的病毒属之一。由于这些病毒的基因组由单链阳性RNA分子组成，并且许多甲型病毒在分子水平上具有很好的特征，因此它们非常适合用于阐明其与蚊子载体相互作用机制的研究。本建议的重点是甲型病毒o’nyong nyong （ONN）和基孔肯雅（CHIK）。ONN在节肢动物传播的病毒中是独特的，因为它是由按蚊类蚊子传播的。相比之下，与ONN最密切相关的CHIK病毒是由伊蚊传播的。利用感染性cDNA克隆技术，可以检查这两种病毒之间的遗传差异，从而允许通过不同的载体宿主传播。该项目的具体目的是：(1)评估CHIK病毒和ONN病毒在冈比亚按蚊中的不同感染性，以对比在流行病学上具有重要意义的媒介中的感染、复制和传播模式。这将提供有关保持不同传播周期的密切相关病毒之间存在的生物学差异的信息；(2)构建含有ONN和一种密切相关的非洲CHIK病毒株部分的嵌合感染性克隆。这些亲代和嵌合克隆将被用来检验一个中心假设，即ONN病毒基因组的特定成分负责这种病毒感染按蚊物种蚊子并由其传播的独特能力；(3)利用CHIK病毒的传染性克隆进行适应性研究，以确定发生突变的可能性，从而导致细胞和载体特异性和毒力模式的变化。病毒与蚊子载体相互作用的分子动力学在很大程度上尚未得到研究。加深对这些相互作用的了解将为调查流行病学上重要的疾病媒介的分子生物学提供工具，并有助于设计控制媒介传播疾病的潜在新靶点。

Mosquito borne pathogens are important causes of disease in both humans and animals. Historically, attempts to disrupt pathogen transmission cycles have depended upon reducing vector populations through the use of insecticides or immunization of vertebrate hosts. Unfortunately, development of insecticide resistance and the collapse of vector control programs necessitate the development of alternate strategies for the control of vector borne pathogens. Novel approaches aimed at reducing vectorial capacity by elucidating elements of virus/vector interactions at the molecular level may be one of the most effective methods in attaining this goal and thus, controlling the spread of arthropod-borne viral disease. Alphaviruses are one of the best studied genera of arthropod borne viruses. Because the genome of these viruses consists of a single stranded, positive sense RNA molecule and many alphaviruses are well characterized at the molecular level, they are ideally suited for studies designed to elucidate mechanisms of interaction with their mosquito vectors. This proposal focuses on the alphaviruses o'nyong nyong (ONN) and chikungunya (CHIK). ONN is unique among arthropod-borne viruses in that it is transmitted by Anopheles species mosquitoes. In contrast, CHIK, the virus most closely related to ONN, is transmitted by Aedes mosquitoes. Using infectious cDNA clone technology, the genetic differences between these two viruses allowing transmission by such disparate vector hosts can be examined. The specific aims of this project are to: (1) Evaluate the differential infectivities of CHIK and ONN viruses in Anopheles gambiae mosquitoes to contrast the patterns of infection, replication, and dissemination within an epidemiologically significant vector. This will provide information on the biological differences that exist between closely related viruses maintaining distinct cycles of transmission; (2) Construct chimeric infectious clones containing portions of both ONN and a closely related African strain of CHIK virus. These parental and chimeric clones will be used to test the central hypothesis that specific components of the ONN viral genome are responsible for the unique ability of this virus to infect and be transmitted by Anopheles species mosquitoes; (3) Perform adaptation studies using an infectious clone of CHIK virus to determine the potential for development of mutations that lead to changes in cell and vector specificity and virulence patterns. The molecular dynamics involved in virus interactions with their mosquito vectors have gone largely unstudied. Developing a greater understanding of these interactions will provide tools for investigating the molecular biology of epidemiologically important disease vectors as well as aid in the design of potential new targets for the control of vector borne diseases.