West Nile Virus Genotypic Affect on Host Competence

描述(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禽类体内病毒滴度高，以及与传播有关的广泛的蚊子媒介。尽管使用禽类死亡率作为西尼罗河病毒活动的前哨，但西尼罗河病毒在禽类中致病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储存库和昆虫媒介复制的基础上生成基本数据，将填补我们对西尼罗河病毒传播知识的关键空白，并有助于改进现有的监测战略和预防人类和兽医疾病的预测紧急情况模型。单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，随着西尼罗河病毒进入新的生态位，可能会选择额外的基因变化，因此迫切需要对其进行基因监测和毒力鉴定。我们关于入侵病毒必须进化和改变毒力以适应新的生态环境的总体假设将通过以下三个具体目标来解决：(1)确定引入的NY99基因与1999年以来从北美鸟类和蚊子以及澳大利亚、肯尼亚和南非分离的其他WNV毒株之间的WNV适应性差异。高灵敏度的病毒适合度分析将被用作通过体内分析检测标准化高、中、低敏感禽类和蚊子宿主表型差异的标记(2)通过将基因变化改造成北美引入后的早期西尼罗河病毒分离株产生的具有感染性的cDNA来表征已确定的用于宿主适应的基因替代的特定作用(3)确定在鸟类或蚊子宿主中长期存在是否会导致在春季终止后改变病毒移除潜力的一致的基因变化。这些数据可以帮助解释为什么西尼罗河病毒疫情在引入新的地理区域后的第二年似乎最广泛。从慢性感染的禽类中分离出的病毒或从受感染的蚊子中垂直传播的病毒将被检查与避免允许持久性的宿主免疫系统相关的基因变化。这些具体目标的完成将为调节病毒出现的选择性机制提供新的见解，并具体描述与鸟类/蚊子感染性相关的病毒基因变化，这些变化促进了西尼罗河病毒在北美的跨洲流行。

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.