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

项目总监/首席研究员（最后、第一、中间）：Franz, Alexander, W.E. 项目概要 登革热病毒的四种血清型（DENV1-4；黄病毒；黄病毒科）是最重要的蚊子病毒。 携带虫媒病毒感染人类。在世界热带地区，DENV 高度流行， 大约25亿人面临流行病传播的风险。 DENV 的主要传播媒介是伊蚊 埃及伊蚊。目前，登革热病毒控制工作主要依靠杀虫剂的应用，因为疫苗或抗病毒药物 药物无法广泛使用。此外，伊蚊对杀虫剂的抗药性正在增强。埃及伊蚊 人口，需要制定替代的遗传载体/病毒控制策略。一本这样的小说 策略是种群替代，其中易受 DENV 影响的野生型蚊子被替换为 转基因、抗病毒的蚊子。该拨款申请的目的是产生转基因 艾。埃及伊蚊系，对所有四种登革热病毒血清型均具有耐药性。 蚊子通过摄入含病毒的血粉从人类宿主那里获得登革热病毒。中肠上皮是 第一个感染 DENV 的蚊子组织。 DENV 需要在中肠建立感染 病灶才能传播到包括唾液腺在内的次级组织，这些组织必须经过 在病毒传播到新的人类宿主之前就被感染。 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 DENV2 的 RNAi 可以完全消除 在病毒建立感染灶之前发生感染。我们最近生成了转基因 Ae。埃及伊蚊那个 在血喂养的中肠组织中表达源自 DENV2 基因组的反向重复 (IR) dsRNA 感染开始时有蚊子。一个 DENV2 难治性转基因品系 (Carb109) 已被维持了 超过 30 代，Carb109 转基因已渗入野生型 Ae 中。埃及伊蚊 将这些蚊子从高度易感的 DENV2 表型转变为完全难治的 表型。 在这里，我们建议在AE中设计RNAi介导的抗性。埃及伊蚊对所有四种 DENV 血清型使用 红外效应器结构的新颖设计。我们将设计 190 bp IR 构建体，每个构建体都针对高度 编码DENV1-4 NS5的病毒RNA的保守区域。抗病毒效应基因将至关重要 现场基于人口替代的 DENV 控制策略的组成部分。拟议的研究 努力将进一步强化人口更替的概念，并补充其他机构正在进行的研究 研究小组将抗病毒效应结构转移到野生型蚊子种群中。 0925-0001/0002（修订版 08/12） 页面延续 格式页面