Transgenic Resistance of Aedes aegypti to the Four Serotypes of Dengue Virus

埃及伊蚊对登革病毒四种血清型的转基因抗性

• 批准号: 8748903
• 负责人: Alexander W E Franz
• 金额: $ 19.19万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-07 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8748903
• 关键词: Aedes; Antiviral Agents; Antiviral resistance; Applications Grants; Arboviruses; Complement; Culicidae; Data; Dengue; Dengue Virus; Development; Disease; Double-Stranded RNA; Engineering; Epidemic; Epithelium; Female; Flaviviridae; Flavivirus; Generations; Genes; Genetic; Genetic Engineering; Genetic Vectors; Genome; Genotype; Goals; Health; Human; Immune; Infection; Inherited; Insecticide Resistance; Insecticides; Intake; Laboratories; Link; Measures; Mediating; Midgut; Molecular Analysis; Oral; Outcome; Pathway interactions; Pharmaceutical Preparations; Phenotype; Population; Population Replacements; Principal Investigator; Publishing; RNA Interference; RNA Interference Pathway; Refractory; Research; Research Personnel; Resistance; Risk; Salivary Glands; Secondary to; Serotyping; System; Target Populations; Testing; Therapeutic; Tissues; Transgenes; Transgenic Organisms; Vaccines; Viral; Virus; Work; base; density; design; fitness; killings; mathematical model; novel; novel strategies; prevent; programs; public health relevance; resistance mechanism; transmission process; vector; vector control; vector mosquito; viral RNA

Program Director/Principal Investigator (Last, First, Middle): Franz, Alexander, W.E. Project summary The four serotypes of dengue viruses (DENV1-4; Flavivirus; Flaviviridae) are the most important mosquito- borne arboviruses infecting humans. In tropical regions of the world, DENV are hyper-endemic with approximately 2.5 billion people at risk for epidemic transmission. The principal vector of DENV is Aedes aegypti. Currently, DENV control efforts rely primarily on insecticide applications, since vaccines or antiviral drugs are not available for widespread use. Additionally, insecticide resistance is increasing among Ae. aegypti populations, requiring the development of alternative, genetic vector/virus control strategies. One such novel strategy is population replacement in which wild-type mosquitoes susceptible for DENV are replaced by genetically-modified, virus-resistant mosquitoes. The purpose of this grant application is to generate transgenic Ae. aegypti lines, which are refractory to all four serotypes of DENV. The mosquito acquires DENV from a human host by intake of a viremic bloodmeal. The midgut epithelium is the first mosquito tissue that becomes infected with DENV. In the midgut, DENV needs to establish infection foci before being able to disseminate to secondary tissues including the salivary glands, which have to be infected before the virus can be transmitted to a new human host. The RNA interference (RNAi) pathway in Ae. aegypti is the major antiviral immune pathway targeting arboviruses. We have shown that triggering RNAi against DENV2 in midgut tissue completely eliminates infection before the virus can establish infection foci. We recently generated transgenic Ae. aegypti that express an inverted-repeat (IR) dsRNA derived from the genome of DENV2 in midgut tissue of bloodfed mosquitoes as infection begins. One DENV2-refractory transgenic line (Carb109) has been maintained for more than 30 generations and the Carb109 transgene has been introgressed into wild-type Ae. aegypti converting these mosquitoes from a highly susceptible DENV2 phenotype to a completely refractory phenotype. Here, we propose to engineer RNAi-mediated resistance in Ae. aegypti to all four DENV serotypes using a novel design for the IR effector constructs. We will design 190 bp IR constructs each targeting a highly conserved region of viral RNA encoding DENV1-4 NS5. The antiviral effector genes will be critically important components for a population replacement-based control strategy of DENV in the field. The proposed research effort will further strengthen the concept of population replacement and complement ongoing studies by other research groups to move anti-viral effector constructs into wild-type mosquito populations. 0925-0001/0002 (Rev. 08/12) Page Continuation Format Page

项目负责人/主要研究者（最后、第一、中间）：Franz，亚历山大，W.E. 项目摘要 登革热病毒的四种血清型（DENV 1 -4;黄病毒;黄病毒科）是最重要的蚊子- 感染人类的虫媒病毒。在世界的热带地区，DENV是高度地方性的， 约25亿人面临疫情传播的风险。登革病毒的主要传播媒介是伊蚊 埃及人。目前，DENV控制工作主要依赖于杀虫剂的应用，因为疫苗或抗病毒药物， 药物无法广泛使用。此外，杀虫剂的耐药性正在增加Ae。aegypti 这是一个非常严重的问题，需要制定替代性的遗传病媒/病毒控制战略。这其中的一个新颖 策略是种群替换，其中对DENV易感的野生型蚊子被替换为 转基因抗病毒蚊子这项资助申请的目的是生产转基因的 AE.埃及系，其对DENV的所有四种血清型都是难治的。 蚊子通过摄入病毒血症血粉从人类宿主获得DENV。中肠上皮是 第一个感染登革病毒的蚊子组织。在中肠，DENV需要建立感染 在能够扩散到包括唾液腺在内的次级组织之前， 在病毒传染给新的人类宿主之前就被感染了。 Ae.中的RNA干扰（RNAi）途径。埃及伊蚊是主要的抗病毒免疫途径， 虫媒病毒我们已经证明，在中肠组织中触发针对DENV 2的RNAi完全消除了 在病毒能够建立感染灶之前感染。我们最近产生了转基因Ae。埃及， 在中肠组织中表达来自DENV 2基因组的反向重复序列（IR）dsRNA， 感染开始时有蚊子。一个DENV 2抗性转基因系（Carb 109）已经维持了10年。 Carb 109转基因已经渗入野生型Ae中。aegypti 将这些蚊子从高度易感的DENV 2表型转化为完全难治的DENV 2表型。 表型 在这里，我们建议在Ae中工程化RNAi介导的抗性。埃及伊蚊感染所有四种DENV血清型， 红外效应器结构的新颖设计。我们将设计190 bp的IR构建体，每个构建体靶向一个高表达的 编码DENV 1 -4 NS 5的病毒RNA的保守区。抗病毒效应基因将是至关重要的 在该领域中，DENV的基于种群扩散的控制策略的组成部分。拟议研究 这一努力将进一步加强人口更替的概念，并补充其他机构正在进行的研究。 研究小组将抗病毒效应子结构转移到野生型蚊子种群中。 0925-0001/0002（2012年8月修订版）续页格式页