Diapause in Culex Mosquitoes

描述（由申请人提供）：蚊媒疾病是大多数发展中国家的主要健康问题，并已成为发达国家流行病爆发的重大威胁。使这些问题恶化的是蚊子种群中杀虫剂耐药性的出现、寄生虫种群中的耐药性、疫苗开发进展的延迟、对蚊子控制计划的支持缺乏以及全球旅行的增加。尖音库蚊复合群中的蚊子是淋巴丝虫样线虫寄生虫、班氏吴策线虫和几种虫媒病毒的主要载体。这包括最近在北美引入的西尼罗河病毒。拟议研究的长期目标是确定、分离和表征与滞育相关的遗传因素，滞育是一种强烈影响尖音库蚊复合群内物种地理分布和生存的特征。目前，对库蚊滞育的分子决定因素还没有明确的认识。本申请的目的是基于一般假设，即与滞育表型相关的基因可以通过定位克隆技术鉴定和分离。确定滞育的基因和相关的生物化学途径的详细了解的知识可能有几个潜在的应用，包括：转基因结构，促进改变和疾病限制行为后，人口更替，有针对性的生物农药的发展，或生物相关的拮抗剂，破坏行为的病原体传播或生存的关键。该项目的具体目标是：1）精细绘制被确定为含有决定滞育表型的基因的基因组区域，2）确定决定滞育的候选基因，以及3）对决定滞育的候选基因进行功能分析。来自遗传杂交的信息将被利用来对抗Cx。quinquefasciatus基因组计划，以确定离散基因组区域内的候选基因。候选基因将使用微阵列进行评估，然后进行定量真实的时间PCR测定，以确认和扩展有希望的基因的微阵列结果。将使用重组辛德比斯转导病毒，使用RNA干扰介导的基因敲除或基因过表达来检测阳性候选基因的生物相关性。然后将在代表Cx的自然种群中评价实验室实验的结果。派皮恩斯公司所以Cx。致倦肌公共卫生相关性：尖音库蚊复合群中的蚊子是人类淋巴丝虫病和包括西尼罗河病毒在内的几种虫媒病毒的主要传播媒介。没有可用的疫苗，疾病预防在很大程度上取决于避免接触受感染的蚊子。这项拟议中的研究将为蚊子的生物学提供有价值的新信息，这些信息与成年雌性蚊子在滞育状态下度过温带气候冬季的能力有关。

DESCRIPTION (provided by applicant): Mosquito-borne diseases represent major health problems in most developing countries and have emerged as significant threats for epidemic outbreaks in developed countries. Exacerbating these problems are the emergence of insecticide resistance in mosquito populations, drug resistance among parasite populations, delayed progress in vaccine development, lack of support for mosquito control programs, and increased global travel. Mosquitoes in the Culex pipiens complex are primary vectors for the lymphatic filarioid nematode parasite, Wuchereria bancrofti and several arboviruses. This includes the recently introduced West Nile virus in North America. The long- term objectives of the proposed research are to identify, isolate and characterize genetic factors associated with diapause, a trait that strongly impacts the geographic distribution and survival of species within the Culex pipiens complex. At present, no knowledge exists on the molecular determinants of diapause in Culex. The objectives of this application are based on the general hypothesis that genes associated with the diapause phenotype can be identified and isolated by positional cloning techniques. Identification of genes determining diapause and the knowledge from the detailed understanding of the associated biochemical pathways could have several potential applications including: transgene constructs that promote altered and disease limiting behaviors following population replacement, development of targeted biopesticides, or identification of biologically relevant antagonists that disrupt behaviors critical to pathogen transmission or survival. The specific aims of the project are to: 1) fine-scale map genome regions identified as containing genes that determine the diapause phenotype, 2) identify candidate genes determining diapause, and 3) conduct functional analysis of candidate genes determining diapause. Information from genetic crosses will be leveraged against the Cx. quinquefasciatus genome project to identify candidate genes within discrete genome regions. Candidate genes will be evaluated using microarrays, followed by quantitative real time PCR assays to confirm and expand microarray results for promising genes. Positive candidate genes will be assayed for biological relevance using RNA interference mediated gene knockdowns or gene overexpression using recombinant Sindbis transducing viruses. Results from laboratory experiments will then be evaluated among natural populations representing Cx. pipiens s.s. and Cx. quinquefasciatus. PUBLIC HEALTH RELEVANCE: Mosquitoes in the Culex pipiens complex are primary vectors for human lymphatic filariasis and several arboviruses, including the West Nile virus. No vaccines are available and disease prevention is largely dependent on avoiding contact with infected mosquitoes. The proposed research will provide valuable new information on the biology of the mosquito relating to the ability of adult females to survive temperate climate winter in a diapause state.