Immune Response of Mosquitoes to Filarial Worms

蚊子对丝虫的免疫反应

• 批准号: 6659796
• 负责人: BRUCE MARTIN CHRISTENSEN
• 金额: $ 21.83万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-20 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6659796
• 关键词: Aedes; Culicidae; Filarioidae; arthropod borne communicable disease; arthropod genetics; arthropod nonpollutant control; biotechnology; cellular immunity; disease vectors; epizootiology; genetic library; genetically modified animals; host organism interaction; insect control; malaria; microarray technology; parasitism

DESCRIPTION (provided by applicant): The resurgence of mosquito-borne diseases is having a devastating impact on global health and an urgent need exists to develop new control strategies. A new strategy being actively investigated is the use of transgenics for the control of vector-borne diseases, i.e., to change vector competence by genetic transformation to reduce the ability of mosquitoes within a population to transmit a particular pathogen. However, if we are to evaluate the use of mosquito transformation as a viable disease control methodology it is imperative that we gain a better understanding of the mosquito's innate immune response that governs vector competence. Melanotic encapsulation is a cellular innate immune response that is responsible for the resistant phenotype of mosquitoes to filarial worms and malaria parasites. Through support from our parent grant (Immune Response of Mosquitoes to Filarial Worms) we are studying the biochemistry and genetic regulation of melanotic encapsulation, but during the course of these studies we have determined that detailed studies of gene expression in the circulating immune reactive cells (hemocytes) of mosquitoes is necessary if we are to clarify mechanisms of immune recognition and the initiation of defense responses. Very few hemocytes (<1,200) are present in an individual mosquito and they are not amenable to in vitro culture or even short-term in vitro maintenance; consequently, we have constructed cDNA libraries representing mRNA obtained from immune-activated hemocytes from two mosquito species. Generation of a limited number of ESTs from these libraries indicated a low percentage of ribosomal clones and revealed a number of matches to proteins with known innate immunity functions. These data convinced us that the systematic development of additional ESTs would facilitate the construction of informative microarrays that could be used to test specific hypotheses about the role hemocytes play in influencing vector competence. The budget of the parent grant was not designed to cover the costs required to generate the number of ESTs required, let alone the costs associated with oligonucleotide microarray construction; and, importantly, we could not have anticipated the feasibility of a comprehensive transcriptome approach to assess gene regulation in our model. The specific aims of the parent grant would be significantly enhanced by the multi-dimensional data from these additional tools. It is therefore appropriate to use the R21 mechanism for technology application to enhance our NIAID-funded research by (1) generating a large EST data set from immune-activated hemocyte cDNA libraries, (2) constructing oligonucleotide microarrays representing these ESTs, and (3) using these microarrays in data mining of hemocyte expression profiles to identify distinct patterns of transcription underlying the immune response of mosquitoes against filarial worms.

描述（由申请人提供）：蚊媒疾病的死灰复燃正在对全球健康产生毁灭性影响，迫切需要制定新的控制策略。正在积极研究的一项新策略是利用转基因来控制媒介传播的疾病，即通过遗传转化改变媒介能力，以降低种群内蚊子传播特定病原体的能力。然而，如果我们要评估蚊子转化作为一种​​可行的疾病控制方法的使用，我们就必须更好地了解蚊子控制媒介能力的先天免疫反应。黑色素包裹是一种细胞先天免疫反应，导致蚊子对丝虫和疟疾寄生虫产生抗药性表型。通过我们的家长资助（蚊子对丝虫的免疫反应）的支持，我们正在研究黑色素包裹的生物化学和遗传调控，但在这些研究过程中，我们已经确定对循环免疫反应细胞中基因表达的详细研究（如果我们要阐明免疫识别和防御反应启动的机制，蚊子的血细胞是必要的。蚊子个体中的血细胞很少（<1,200），并且不适合体外培养，甚至不适合短期体外维持；因此，我们构建了 cDNA 文库，代表从两种蚊子的免疫激活血细胞中获得的 mRNA。从这些文库中生成的有限数量的 EST 表明核糖体克隆的比例较低，并揭示了与具有已知先天免疫功能的蛋白质的许多匹配。这些数据使我们相信，其他 EST 的系统开发将有助于构建信息丰富的微阵列，该微阵列可用于测试有关血细胞在影响载体能力中所起作用的具体假设。母公司拨款的预算并非旨在支付生成所需数量的 EST 所需的费用，更不用说与寡核苷酸微阵列构建相关的费用了；而且，重要的是，我们无法预料到在我们的模型中采用全面的转录组方法来评估基因调控的可行性。这些附加工具的多维数据将显着增强母基金的具体目标。因此，使用 R21 机制进行技术应用来增强我们的 NIAID 资助的研究是适当的，方法是 (1) 从免疫激活的血细胞 cDNA 文库生成大型 EST 数据集，(2) 构建代表这些 EST 的寡核苷酸微阵列，以及 (3 ）使用这些微阵列对血细胞表达谱进行数据挖掘，以识别蚊子针对丝虫的免疫反应背后的不同转录模式。