Paratransgenesis for malaria using Asaia sp. bacteria

使用 Asaia sp. 治疗疟疾的副转基因。

• 批准号: 8767309
• 负责人: DAVID J LAMPE
• 金额: $ 39.6万
• 依托单位: DUQUESNE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8767309
• 关键词: Adult; Anopheles Genus; Antimalarials; Bacteria; Biological Metamorphosis; Blood; Characteristics; Chromosomes; Complement; Contracts; Culicidae; Data; Disease; Ecology; Engineering; Ensure; Female; Gene Transfer Techniques; Genes; Genetic Screening; Genomics; Global Warming; Goals; Gonadal structure; Health; Human; Incidence; Insecta; Insecticides; Laboratories; Malaria; Methods; Midgut; Morbidity - disease rate; Pantoea agglomerans; Parasites; Pharmaceutical Preparations; Phenotype; Plasmids; Plasmodium; Population; Prevalence; Protein Secretion; Proteins; Research Project Grants; Resistance; Risk; Route; Salivary Glands; Signal Transduction; System; Testing; Time; Tissues; Transgenic Organisms; Vector-transmitted infectious disease; base; combat; design; fighting; global health; killings; meetings; microbial; mortality; offspring; prevent; promoter; public health relevance; tool; vector; vector mosquito

DESCRIPTION (provided by applicant): Malaria is a disease caused by protozoan parasites that are transmitted to humans by mosquitoes in the genus Anopheles. There are nearly 500 million new cases of malaria every year, and from 1-2 million people die from the disease while others are severely debilitated. About half the human population is at risk of contracting malaria, and its range may spread as global warming accelerates. The broad, long term objectives of this proposal are to create new methods of combating malaria to complement the current methods of control, namely insecticides to kill mosquito vectors and drugs to kill parasites in infected people. This project seeks to develop the means to create strains of bacteria that can interfere with the ability of mosquitoes to transmit malaria thus reducing its overall health burden and aiding in the goal of eradicating this disease. The specific aims of this research project are as follows: Aim 1: Creation of strains of Asaia SF2.1 that secrete anti-Plasmodium effector proteins using native secretion signals. Asaia SF2.1 is intimately associated with Anopheles mosquitoes in the field, colonizing the midgut, salivary glands, and gonads of these insects. Thus it has optimal microbial ecology on which to build a paratransgenesis system against malaria. We will develop native secretion systems using genomic data and a genetic screen for the secretion of anti-malarial effector proteins from Asaia SF2.1 to ensure efficient secretion of these proteins in mosquito midguts. Aim 2: Isolation of strong conditional promoters from Asaia SF2.1. Antimalarial effector proteins must be expressed when the parasite is present in the mosquito midgut and in sufficient quantities to be effective. We will isolate strong conditional promoters from Asaia SF2.1 that are active when bacteria are present during a blood meal using a genetic screen, genomic homology searches, and RNAseq. Aim 3: Creation of genetically stable strains of transgenic Asaia SF2.1. For eventual field use, paratransgenic strains of bacteria must be genetically stable. They cannot be based on laboratory plasmids that are maintained with drug selection. We will develop methods to create strains that contain genes inserted in the chromosome or borne on plasmids that need no drug selection.

描述(申请人提供)：疟疾是一种由原生动物寄生虫引起的疾病，这些寄生虫通过按蚊传播给人类。每年有近5亿新的疟疾病例，有100-200万人死于这种疾病，而其他人则严重虚弱。大约一半的人口面临感染疟疾的风险， 随着全球变暖的加速，其范围可能会扩大。这项提议的广泛和长期目标是创造新的防治疟疾的方法，以补充目前的控制方法，即杀死蚊媒的杀虫剂和杀死感染者寄生虫的药物。该项目寻求开发手段，创造能够干扰蚊子传播疟疾能力的细菌菌株，从而减轻蚊子的总体健康负担，并帮助实现根除这种疾病的目标。本研究项目的具体目标如下：目标1：利用天然分泌信号产生分泌抗疟原虫效应蛋白的Asaia SF2.1菌株。Asaia SF2.1与野外按蚊密切相关，在这些蚊子的中肠、唾液腺和性腺中定居。因此，它拥有最优的微生物生态，在此基础上建立抗击疟疾的副转基因系统。我们将利用基因组数据开发天然的分泌系统，并利用遗传筛选从Asia SF2.1中分泌抗疟疾效应蛋白，以确保这些蛋白在 蚊子的中肠。目的2：从Asia SF2.1中分离出强条件启动子。当寄生虫存在于蚊子中肠并且数量足够时，必须表达抗疟疾效应器蛋白才能起作用。我们将使用基因筛查、基因组同源性搜索和RNAseq从Asia SF2.1中分离出当血餐中存在细菌时具有活性的强条件启动子。目的3：建立转基因Asia SF2.1的遗传稳定株系。对于最终的现场使用，副转基因细菌菌株必须在遗传上稳定。它们不能基于在药物选择过程中保持的实验室质粒。我们将开发方法来创造包含插入到染色体中的基因或携带在不需要药物选择的质粒上的基因的菌株。