Effect of Powassan virus genetic variation on replication in human cell culture and transmission by ticks

项目摘要 Powassan病毒（POWV）是一种神经侵入性蜱传黄病毒，在自然界中由硬蜱维持。 在过去的十年里，人类病例的数量翻了两番，野生动物研究表明， 地方性传染的强度也在增加。POWV的特点是不同的传输周期， 疾病结果。该病毒由两个不同的系统发育分支组成， 在几种蜱和脊椎动物中观察到。POWV之间关系的实验研究 目前缺乏遗传多样性、蜱传播和疾病结果。本研究的目的是 体外测定POWV的表型多样性，鉴定潜在医学重要性的相关基因型， 并确定病毒在蜱中传播的效率。为了便于研究，我们收集了47 环境分离株跨越北美广泛的地理范围，从一些蜱和 脊椎动物来源，跨越60多年。我们假设POWV是一种表型多样的 在潜在疾病和传播能力方面可能不同的病毒。为了验证这一假设，我们确定复制 以及人和蜱细胞培养物中的细胞病变效应表型。与Broad Institute合作，我们 使用POWV全基因组序列鉴定与细胞中病毒表型相关的遗传变异 文化在我们的第二个目标中，我们使用一种创新的病毒条形码方法来调查病毒传播 在4种蜱传播途径中：经口、垂直、经食道和共同进食。这些信息对于 了解POWV如何在tick向量中持续存在。这项研究的结果有助于我们 目前缺乏对这种新出现的黄病毒的了解。建议的目标如下： 目的1：量化细胞培养物中的POWV表型多样性并鉴定感兴趣的遗传变体。 目的2：探讨不同传播途径对肩突硬蜱体内Powassan病毒种群的影响。

PROJECT SUMMARY Powassan virus (POWV) is a neuroinvasive tick-borne flavivirus that is maintained in nature by Ixodid ticks. Over the past decade, the number of human cases has quadrupled, and wildlife studies suggest that the intensity of enzootic transmission is also increasing. POWV is marked by diverse transmission cycles and disease outcomes. The virus consists of two distinct phylogenetic clades, and evidence of infection has been observed in several tick and vertebrate species. Experimental studies of the relationships between POWV genetic diversity, tick transmission, and disease outcome are currently lacking. The goal of this study is to determine phenotypic diversity of POWV in vitro, identify relevant genotypes of potential medical importance, and determine efficiency of virus dissemination in ticks. To facilitate this study, we have collected 47 environmental isolates spanning a wide geographic range in North America, from a number of tick and vertebrate sources, and spanning over 6 decades. We hypothesize that POWV is a phenotypically diverse virus that can differ in disease potential and ability to transmit. To test this hypothesis, we determine replication and cytopathic effect phenotypes in human and tick cell culture. In collaboration with the Broad Institute, we use POWV whole genome sequences to identify genetic variants that correlate with virus phenotype in cell culture. In our second aim, we use an innovative method of barcoding virus to investigate virus dissemination in 4 tick transmission routes: oral, vertical, transstadial, and co-feeding. This information is critical to understanding how POWV persists in the tick vector. The results of this proposed study contribute to our currently lacking knowledge of this emerging flavivirus. The following aims are proposed: Aim 1: Quantify POWV phenotypic diversity in cell culture and identify genetic variants of interest. Aim 2: Assess the effects of transmission route on Powassan virus population in Ixodes scapularis ticks.