West Nile Virus Genotypic Affect on Host Competence

描述（由申请人提供）：西尼罗河病毒（WNV）已成为美国虫媒病毒性脑炎的主要原因。与西尼罗河病毒迅速出现相关的最显著的流行病学因素是鸟类中病毒滴度高以及与传播有关的蚊虫媒介范围广泛。尽管使用鸟类死亡率作为西尼罗河病毒活动的哨兵，但西尼罗河病毒在鸟类中致病性的潜在病毒遗传基础以及媒介能力增强在西尼罗河病毒传播中的作用尚不清楚。在西尼罗河病毒禽类宿主和昆虫媒介内复制的基础上生成基础数据将填补我们在西尼罗河病毒传播知识方面的关键空白，并有助于改进现有的监测战略和预测出现模型，以预防人类和兽医疾病。单个基因改变可以影响西尼罗河病毒的传播潜力，这一事实表明，在西尼罗河病毒进入可能选择额外遗传改变的新生态位时，迫切需要对其进行基因型监测和毒力表征。我们的总体假设是，入侵病毒必须进化并改变其毒力以适应新的生态位，这将通过以下三个具体目标来解决：(1)确定引入的NY99基因型与1999年以来从北美鸟类和蚊子中分离的毒株以及来自澳大利亚、肯尼亚和南非的其他西尼罗河病毒毒株之间的西尼罗河病毒适应度差异。高度敏感的病毒适应度测定将被用作检测标准化高、(2)在北美引进的早期西尼罗河病毒分离物中，通过将遗传变化工程转化为感染性cDNA，鉴定了遗传替代对宿主适应的具体作用。(3)确定在鸟类或蚊子宿主越冬期间的长期持续存在是否会导致一致的遗传变化，从而在春季终止后改变致病毒潜能。这些数据可以帮助解释为什么西尼罗河病毒在进入新的地理区域后的一年似乎传播最广。将检查从慢性感染的鸟类中分离的病毒或从受感染的蚊子中垂直传播的病毒，以寻找与避开宿主免疫系统相关的基因变化，从而使病毒得以持续存在。完成这些具体目标将为调节病毒出现的选择性机制提供新的见解，并具体描述与鸟/蚊子传染性相关的病毒遗传变化，这些变化促进了西尼罗病毒在北美的跨大陆流行。

Description (provided by applicant): West Nile virus (WNV) has become the leading cause of arboviral encephalitis in the United States. The most notable epidemiological factors associated with the rapid emergence of WNV have been the development of high viral titers within birds and the wide range of mosquito vectors implicated with transmission. Despite the use of bird mortality as a sentinel for WNV activity, the underlying viral genetic basis for WNV pathogenicity in birds and the role of increased vector competence in the transmission of WNV are poorly understood. Generation of fundamental data on the basis of replication within the WNV avian reservoir and insect vector will fill critical gaps in our knowledge of WNV transmission and aid in the improvement of existing surveillance strategies and predictive emergence models for the prevention of human and veterinary disease. The fact that a single gene alteration can impact on the transmission potential of WNV indicates the strong need for genotypic monitoring and virulence characterization of WNV as it moves into new ecological niches that may select for additional genetic change. Our overall hypothesis that that an invading virus must evolve and alter its virulence to adapt to new ecological niches will be addressed by the following three specific aims: (1) Identify WNV fitness differences between the introduced NY99 genotype and strains isolated from North American birds and mosquitoes since 1999 as well as other WNV strains from Australia, Kenya and South Africa. Highly sensitive viral fitness assays will be employed as a marker for the detection of phenotypic differences in standardized high, medium and low susceptible avian and mosquito hosts using in vivo assays (2) Characterize the specific role of genetic substitutions identified for host adaptation by engineering the genetic changes into an infectious cDNA made from an early WNV isolate following North American introduction (3) Determine if long term persistence in avian or mosquito hosts during the overwintering period results in consistent genetic change that alter viremogenic potential after vernal termination. These data could help explain why WNV epidemics seem most extensive the year following introduction into new geographical areas. Virus isolated from chronically infected birds or passed vertically from infected mosquitoes will be examined for genetic changes associated with the avoidance of the host immune systems that allow for persistence. Completion of these specific aims will provide new insight into the selective mechanisms that modulate viral emergence and specifically delineate Iviral genetic changes associated with avian/mosquito infectivity that have facilitated the transcontinental epidemic of West Nile virus in North America.