Determinants of Usutu virus bird-to-mosquito transmission

项目摘要 西尼罗河病毒（WNV）、圣路易斯脑炎病毒（SLEV）和西尼罗河病毒（USUV）密切相关 蚊子传播的病毒，导致人类神经侵入性疾病。这三种病毒都保存在 重叠的鸟类/蚊子周期，并蔓延到人类。1999年西尼罗河病毒进入美国 由于鸟类中的交叉反应性免疫应答，快速取代了先前传播的SLEV。但 SLEV的新基因型最近在美国出现，导致了2015年的爆发，我们发现， 鸟类中的西尼罗河病毒免疫应答在体外不能中和新的SLEV基因型。USUV最近出现了 在欧洲，它至少三次从非洲引进。我们发现家雀 （Passer acuticus）和致倦库蚊（Culex quinquefasciatus），它们对WNV和SLEV有能力， 也容易受到USUV的影响。这个项目的长期目标是了解影响小说的因素 黄病毒的出现，以预测在人类中的爆发。本研究的目的是确定 主管北美鸟类和蚊子物种的USUV，并确定西尼罗河病毒免疫的作用 在鸟类的SLEV和USUV出现。假设新的SLEV基因型和USUV具有 已建立或有可能建立北美的传输周期，因为有能力 本地雀鸟和蚊子，以及雀鸟逃避对西尼罗河病毒的免疫力， 人类的神经侵袭性疾病两个具体的目标将解决这个假设：1）确定是否北 美国的雀形目和蚊子对USUV有能力; 2）评估以前的影响 暴露于西尼罗河病毒对鸟类SLEV和USUV感染的影响。在第一个目标中，我们将捕捉三种野生鸟类 以确定当地鸟类对USUV的敏感性。我们将量化USUV的感染阈值， 通过在受感染的鸟类上喂食蚊子，然后测量鸟类中的传染性病毒， 受感染蚊子的吐出物。在第二个目标中，WNV预暴露对随后的 将在鸟类中评估新的SLEV基因型和USUV的感染。接种过的鸟的血清将 测试了对所有三种病毒的体外中和活性，以确定诊断 用血清学鉴定黄病毒感染。本研究具有创新性，因为它 研究了一种新的SLEV基因型和USUV，这是被忽视的，新兴的病毒，使用一种新的禽流感病毒， 实验模型在成功完成拟议的研究，预期的贡献，这 工作将是确定USUV水库和媒介物种和影响地方性流行病的因素， SLEV和USUV的传输。预计这一贡献将是重要的，因为了解 新出现的黄病毒的传播周期将导致预测人类疾病的能力。

Project Summary West Nile virus (WNV), St. Louis encephalitis virus (SLEV), and Usutu virus (USUV) are closely-related mosquito-borne viruses that cause neuroinvasive disease in humans. All three viruses are maintained in overlapping bird/mosquito cycles, with spillover to humans. The introduction of WNV into the U.S. in 1999 quickly displaced the previously-circulating SLEV due to cross-reactive immune responses in birds. However, a novel genotype of SLEV recently emerged in the U.S., causing an outbreak in 2015, and we have found that WNV immune responses in birds do not neutralize the novel SLEV genotype in vitro. USUV recently emerged in Europe, where it has been introduced at least three times from Africa. We have found that House sparrows (Passer domesticus) and Culex quinquefasciatus mosquitoes, which are competent for WNV and SLEV, are also susceptible to USUV. The long-term goal of this project is to understand the factors that influence novel flavivirus emergence in order to predict outbreaks in humans. The objectives of this study are to identify competent North American avian and mosquito species for USUV and to determine the role of WNV immunity in birds on SLEV and USUV emergence. The hypothesis is that the novel SLEV genotype and USUV have established or have the potential to establish transmission cycles in North America due to the competence of local birds and mosquitoes and the evasion of WNV immunity in birds, thus leading to an increased risk of neuroinvasive disease in humans. Two specific aims will address this hypothesis: 1) Determine whether North American passerines and mosquitoes are competent for USUV; and 2) Assess the influence of previous exposure to WNV on SLEV and USUV infection in birds. In the first aim, we will trap three species of wild birds to determine the susceptibility of local birds to USUV. We will quantify the infectious threshold of USUV for bird-to-mosquito transmission by feeding mosquitoes on infected birds and then measure infectious virus in the expectorate of infected mosquitoes. In the second aim, the effect of WNV pre-exposure on subsequent infection by the novel SLEV genotype and USUV will be evaluated in birds. Sera from inoculated birds will be tested for in vitro neutralizing activity against all three viruses to determine the capacity for diagnostically identifying flavivirus infections using serology. The research proposed here is innovative because it investigates a novel SLEV genotype and USUV, which are neglected, emerging viruses, using a novel avian experimental model. Upon successful completion of the proposed research, the anticipated contribution of this work will be the identification of USUV reservoir and vector species and factors that influence enzootic transmission of SLEV and USUV. This contribution is expected to be significant because understanding the transmission cycle of emerging flaviviruses will lead to the ability to predict disease in humans.