Microbial interplay between ZIKA virus and the native microbiome in mosquitoes

项目摘要。 蚊子的原生微生物群深刻地影响了蚊子生物学的许多方面，包括蚊子的繁殖能力。 蚊子传播虫媒病毒病原体。蚊子微生物组的扰动，无论是通过补充 抗生素或通过口服接种细菌分离物进入蚊子肠道可以干扰黄病毒，如登革热 病毒（DENV）和西尼罗河病毒以及甲病毒如基孔温亚病毒（CHIKV）。大多数研究表明， 微生物对病原体具有拮抗作用，但特定的细菌促进蚊子中的病毒感染。此外，我们 开始意识到蚊子中的虫媒病毒感染可以改变细菌微生物组。综合来看， 很明显，蚊子的肠道微生物之间存在着复杂的相互作用，无论是通过直接还是间接的机制， 和入侵虫媒病毒，这些相互作用对载体的能力有着深远的影响。在这里，我们将描述 伊蚊肠道微生物和寨卡病毒之间的相互作用，以确定这些相互作用如何影响ZIKV 蚊子的媒介能力。我们的工作将集中在埃及伊蚊和伊蚊的细菌和真菌类群 白纹伊蚊虽然细菌共生体改变载体能力的现象正在出现，但我们对细菌共生体改变载体能力的理解还很有限。 真菌微生物对蚊子生物学和媒介能力的作用。我们的工作将利用高通量测序 技术，以获得细菌和真菌微生物组的培养独立的定量表征。 重要的是，实验将使用实验室饲养的蚊子和野外收集的蚊子， 对该领域蚊子的媒介能力的微生物组了解甚少。在具体目标1中，我们将确定 通过高通量测序比较干扰的和 传统饲养的蚊子。使用野外收集的蚊子，ZIKV载体能力将与以下相关联： 微生物丰度和组成。由于微生物组的组成是可变的， 特定的共生体可以解释虫媒病毒载体能力的变化。ZIKV载体感受态测定将在 完成了对具有再感染分离株的知菌蚊系的功能验证， 我们的测序实验。在具体的目标2中，我们将研究ZIKV感染如何改变细菌和真菌感染。 使用高通量测序的微生物组。我们将确定ZIKV抑制的微生物是否具有抗病毒特性 并检查ZIKV介导的微生物组调节是否在世代之间转移。最后，我们将 确定微生物组的这种转代改变是否影响后代的载体能力， ZIKV。这项工作将为蚊子体内微生物的相互作用提供见解，这是一个研究不足但重要的领域 这是一种载体生物学，在控制ZIKV和其他蚊媒病毒疾病方面具有很好的应用前景。这个项目 将为休斯实验室未来的工作奠定基础，这是开发应用新型微生物为基础的 适用于同时控制多种虫媒病毒如ZIKV、DENV、CHIKV和黄热病的策略 病毒

Project summary. The native microbiota of mosquitoes profoundly influences many aspects of mosquito biology, including the ability of mosquitoes to transmit arboviral pathogens. Perturbation of the mosquito microbiome, either by supplementation of antibiotics or by oral inoculation of bacterial isolates into the mosquito gut can interfere with flaviruses such as Dengue virus (DENV) and West Nile Virus as well as alphaviruses like Chikungyunya (CHIKV). Most studies indicate that gut microbes are antagonistic to pathogens, but particular bacteria facilitate viral infection in mosquitoes. Additionally, we are beginning to appreciate that arbovirus infection in mosquitoes can alter the bacterial microbiome. Taken together, it is clear that there is complex interplay, either by direct or indirect mechanisms, between native gut microbes of mosquitoes and invading arbovirus, and these interactions have a profound effect on vector competence. Here we will characterize the interplay between gut microbes of Aedes mosquitoes and Zika virus to determine how these interactions influence ZIKV vector competence in mosquitoes. Our work will focus on both bacterial and fungal taxa of Aedes aegypti and Aedes albopictus. While it is emerging that bacterial symbionts alter vector competence, we have a poor understanding of the role of fungal microbes on mosquito biology and vector competence. Our work will exploit high throughput sequencing technologies to obtain a culture-independent quantitative characterization of the bacterial and fungal microbiome. Importantly, experiments will use both lab-reared and field-collected mosquitoes as our appreciation of the role of the microbiome of on vector competence of mosquitoes in the field is poorly understood. In specific aim 1, we will identify bacterial and fungal taxa that alter vector competence by high throughput sequencing comparing perturbed and conventionally reared mosquitoes. Using field-collected mosquitoes, ZIKV vector competence will be correlated to microbial abundance and composition. As the composition of the microbiome is variable, the presence or abundance of specific symbionts may account for variation in arbovirus vector competence. ZIKV vector competence assays will be completed on gnotobiotic mosquito lines that have had isolates reinfected, functionally validating candidate taxa identified from our sequencing experiments. In specific aim 2, we will examine how ZIKV infection alters the bacterial and fungal microbiome using high throughput sequence. We will determine if microbes that ZIKV inhibits have anti-viral properties and examine if ZIKV-mediated modulation of the microbiome is transferred between generations. Lastly, we will determine if such transgeneration alteration of the micobiome affects vector competence of subsequent generations to ZIKV. This work will provide insights in microbial interactions within mosquito, a poorly understudied but important area of vector biology that has promising applications to control ZIKV and other mosquito-borne viral disease. This project will lay the foundation for future work in the Hughes laboratory, which is to develop applied novel microbial-based strategies suitable for simultaneously controlling multiple arboviruses such as ZIKV, DENV, CHIKV, and yellow fever virus.