Genotypic and Phenotypic Analysis of West Nile virus, Yucatan Peninsula of Mexico

描述（由申请人提供）：西尼罗病毒（WNV）在西半球的首次暴发于1999年在纽约发生，此后该病毒迅速蔓延到美国和加拿大，在那里它已造成严重的人类发病率和死亡率。相比之下，在墨西哥尤卡坦半岛（YP）没有人感染西尼罗河病毒病例，尽管有证据表明存在西尼罗河病毒活动，但也没有在该地区的马或鸟类中广泛存在西尼罗河病毒相关疾病的报告。事实上，我们和我们在梅里达的尤卡坦自治大学（DAY）的墨西哥同事在YP的马、鸟类和其他脊椎动物身上检测到了西尼罗河病毒抗体。然而，与这项建议最相关的是在科苏梅尔岛进行的一项血清调查的结果；144匹马中有75匹（52%）检测到西尼罗河病毒抗体，但没有一匹马有疾病迹象，也没有西尼罗河病毒样疾病史。因此，本提案的基本假设是，YP中的显性西尼罗河病毒变体在基因型和表型上与美国的显性西尼罗河病毒变体不同。在这项资助中，我们和way的同事将获得YP中的西尼罗河病毒分离株，并确定其基因型和细胞培养表型。在具体目标1中，将在YP中以前发现血清阳性脊椎动物的地点捕获蚊子。采用RT-PCR法和病毒分离法对蚊虫进行西尼罗河病毒检测。这些研究将确定YP中可能的西尼罗河病毒载体，从而为未来的媒介能力和宿主摄食行为研究奠定基础。在特定目标2中，西尼罗河病毒分离株将通过部分核苷酸测序和系统发育分析进行检查。WNV的显性变异将被完全测序。在特定目标3中，将比较来自日本和美国的西尼罗河病毒显性变异的细胞培养表型（即复制动力学和产量、斑块形态和温度敏感性）。这些研究将为今后脊椎动物宿主体内发病机制的研究奠定基础。相关性：在本提案中，将对YP中显性西尼罗河病毒变异的基因型和细胞培养表型进行表征，并确定该地区可能的西尼罗河病毒载体。这些信息将有助于确定为什么西尼罗河病毒没有在太平洋平原流行，并将确定与西尼罗河病毒疾病相关的危险因素。

DESCRIPTION (provided by applicant): The initial outbreak of West Nile virus (WNV) in the Western Hemisphere occurred in New York in 1999, and since then the virus has rapidly spread across the United States and Canada, where it has been responsible for significant human morbidity and mortality. In contrast, there have been no human cases of WNV in the Yucatan Peninsula (YP) of Mexico, nor have there been any reports of widespread WNV associated illness in horses or birds in this region, despite evidence of WNV activity. Indeed, we and our Mexican colleagues at the Universidad Autonoma de Yucatan (DAY) in Merida detected antibodies to WNV in horses, birds and other vertebrate animals in the YP. Most pertinent to this proposal, however, are the findings from a serosurvey conducted on Cozumel Island; antibodies to WNV were detected in 75 of 144 (52%) horses, but none of the horses had signs of illness and none had a history of WNV-like illness. Thus, the underlying hypothesis in this proposal is that the dominant WNV variants in the YP are genotypically and phenotypically distinct from the dominant WNV variants in the U.S. In this grant, we and our colleagues at the UAY will obtain WNV isolates in the YP, and define their genotypes and cell culture phenotypes. In specific aim 1, mosquitoes will be trapped at sites in the YP where seropositive vertebrates have previously been identified. Mosquitoes will be assayed for WNV by RT-PCR and virus isolation. These studies will identify likely vectors of WNV in the YP, thereby providing the groundwork for future vector competence and host-feeding behavior studies. In specific aim 2, WNV isolates will be examined by partial nucleotide sequencing and phylogenetic analysis. Dominant WNV variants will be completely sequenced. In specific aim 3, the cell culture phenotypes (i.e. replication kinetics and yields, plaque morphologies and temperature sensitivities) of dominant WNV variants from the YP and U.S. will be compared. These studies will provide the groundwork for future in vivo pathogenesis studies in vertebrate hosts. RELEVANCE: In this proposal, the genotypes and cell culture phenotypes of dominant WNV variants in the YP will be characterized, and likely vectors of WNV in this region will be identified. This information will help determine why there has not been an epidemic of WNV in the YP, and will identify risk factors associated with WNV disease.