Assessing the roles of viral mutations and host factors in the transmission of Mayaro virus and other alphaviruses by urban mosquitoes

评估病毒突变和宿主因素在城市蚊子传播马亚罗病毒和其他甲病毒中的作用

• 批准号: 10663696
• 负责人: Rafael Kroon Campos
• 金额: $ 11.93万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-03 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10663696
• 关键词: Acute Disease; Aedes; Affect; Alphavirus; Antiviral Response; Arboviruses; Arthralgia; Binding Proteins; Biological Assay; Blood; Brazil; Chikungunya fever; Chikungunya virus; Chills; Chimera organism; Culicidae; Development; Disease; Disease Outbreaks; Dissection; Distant; Dose; Double-Stranded RNA; Drug Targeting; Elements; Environment; Epidemic; Evolution; Fever; Financial Hardship; Future; Genes; Genetic; Genome; Genomics; Genotype; Geography; Goals; Headache; Healthcare Systems; Human; Immune Evasion; Immune response; Immune system; Immunity; Infection; Injections; Integration Host Factors; Invertebrates; K-Series Research Career Programs; Knowledge; Laboratories; Laboratory Study; Link; Mayaro virus; Methylation; Methyltransferase; Midgut; Minority; Modeling; Modification; Monitor; Mutagenesis; Mutation; Mutation Fixation; Myalgia; Nonstructural Protein; North America; Pathway interactions; Phosphorylation; Phosphorylation Site; Phylogenetic Analysis; Play; Principal Investigator; Proteins; Publishing; RNA; RNA chemical synthesis; RNA methylation; Reader; Regulation; Research Personnel; Risk; Role; Saliva; Salivary Glands; Series; Single Nucleotide Polymorphism; Testing; Titrations; Urbanization; Vaccines; Variant; Vertebral column; Viral; Viral Genome; Viremia; Virus; Virus Activation; Virus Diseases; Virus Replication; Writing; career; chikungunya; design; experimental study; fitness; forest; genome-wide analysis; genomic RNA; global health; human pathogen; immune activation; immunoregulation; improved; knock-down; mortality; next generation sequencing; novel; novel strategies; oral infection; peri-urban; prospective; skills; surveillance strategy; transmission process; vector; vector competence; viral RNA; viral fitness; viral transmission; virus host interaction

Project Summary/Abstract Alphaviruses are human pathogens that represent a global health threat. Mayaro virus (MAYV) and chikungunya virus (CHIKV) are alphaviruses that can cause acute disease with fever, headache, myalgia, chills, and long- term debilitating arthralgia. Adaptation of CHIKV lineages to the urban mosquitoes Aedes aegypti and Aedes albopictus has contributed to its worldwide distribution and led to large outbreaks. MAYV is thought to be restricted to transmission by sylvatic mosquitoes of the genus Haemagogus, but laboratory studies show that MAYV can also be transmitted by Aedes mosquitoes, suggesting the need for improved surveillance and countermeasures. I found that MAYV can infect Aedes mosquitoes from Salvador (Brazil) or Galveston when present at titers found in viremia in humans. This project aims to understand how adaptive mutations and interactions with hosts may lead to the emergence of alphaviruses outbreaks. My central hypothesis is that mutations in the MAYV nsP3 gene can lead to efficient transmission by Aedes mosquitoes through promoting viral-host interactions. I will address this hypothesis through three specific aims. In specific aim 1, I will determine existing and prospective mutations in the genome of MAYV that function in vector competence of Aedes mosquitoes. I have performed next generation sequencing of salivary glands of infected Aedes mosquitoes and found 17 putatively adaptive mutations. I will find and validate MAYV minority variants that arise upon mosquito infection using competition assays, 50% oral infectious dose experiments and dual host models. As a proof-of- principle, I discovered an adaptive mutation in the virus non-structural protein 3 (nsP3). I will then determine at which step of mosquito infection they are important through a series of dissections followed by titrations. In specific aim 2, I will uncover the roles of nsP3 in mosquito infections and vector competence. I will assemble nsP3 chimeras of different MAYV strains to identify regions that are necessary and sufficient for increased fitness in Aedes mosquitoes and study a natural insertion that has evolved at least twice in MAYV strains, suggesting it is adaptive, and its proposed changes in protein phosphorylation. In specific aim 3, I will assess how m6A modifications on MAYV and CHIKV RNA may promote immune evasion and how these modifications in cellular RNAs modulate the immune system. These modifications are thought to have key functions in immune system regulation and immune evasion. I showed that MAYV has m6A modifications on its RNA which are concentrated in the sub genomic RNA and promote viral replication. Completion of this project will have a major impact in the control of alphaviruses by spurring novel surveillance strategies and countermeasures targeting virus-host interactions. This project and career development award aligns well with my current skills and career goals to become an independent principal investigator. It will help me fill gaps in my knowledge of vectors and to develop key administrative and writing skills required to become an independent researcher.

项目概要/摘要 甲病毒是对全球健康构成威胁的人类病原体。马亚罗病毒 (MAYV) 和基孔肯雅热 病毒 (CHIKV) 是甲病毒，可引起急性疾病，包括发烧、头痛、肌痛、寒战和长期感染。 术语“衰弱性关节痛”。 CHIKV 谱系对城市蚊子埃及伊蚊和伊蚊的适应 白纹伊蚊促进了其在全球范围内的传播并导致了大规模的疫情爆发。 MAYV被认为是 仅限于刺血蚊属的森林蚊子传播，但实验室研究表明 MAYV 也可以通过伊蚊传播，这表明需要加强监测和 对策。我发现 MAYV 可以感染来自萨尔瓦多（巴西）或加尔维斯顿的伊蚊 存在于人类病毒血症中发现的滴度。该项目旨在了解适应性突变和 与宿主的相互作用可能导致甲病毒爆发的出现。我的中心假设是 MAYV nsP3 基因突变可通过促进伊蚊有效传播 病毒与宿主的相互作用。我将通过三个具体目标来阐述这一假设。在具体目标1中，我将确定 MAYV 基因组中现有的和预期的突变对伊蚊的载体能力起作用 蚊子。我对受感染的伊蚊的唾液腺进行了下一代测序 发现了 17 个假定的适应性突变。我将找到并验证蚊子身上出现的 MAYV 少数变体 使用竞争测定、50%口服感染剂量实验和双宿主模型进行感染。作为证明—— 原理上，我发现了病毒非结构蛋白3（nsP3）的适应性突变。然后我将确定 通过一系列解剖和滴定，它们对蚊子感染的哪一步很重要。在 具体目标 2，我将揭示 nsP3 在蚊子感染和媒介能力中的作用。我会集合 不同 MAYV 毒株的 nsP3 嵌合体，用于识别增强适应性所必需和充分的区域 在伊蚊中，并研究了在 MAYV 菌株中至少进化了两次的自然插入，表明它 是适应性的，其提出了蛋白质磷酸化的变化。在具体目标 3 中，我将评估 m6A 如何 MAYV 和 CHIKV RNA 的修饰可能会促进免疫逃避，以及这些修饰如何在细胞中发挥作用 RNA 调节免疫系统。这些修饰被认为在免疫系统中具有关键功能 调节和免疫逃避。我证明 MAYV 在其 RNA 上有 m6A 修饰，这些修饰集中在 存在于亚基因组 RNA 中，促进病毒复制。该项目的完成将对 通过刺激针对病毒宿主的新型监测策略和对策来控制甲病毒 互动。该项目和职业发展奖与我当前的技能和职业目标非常吻合 成为独立首席研究员。它将帮助我填补向量知识的空白并发展 成为独立研究员所需的关键管理和写作技能。