Plant Production and Delivery System for Bacterial Toxins to Mosquity Larvae

描述(申请人提供)：蚊子是许多血液传播的人类疾病的昆虫媒介，这些疾病每年在全球范围内导致数百万人死亡。蚊子幼虫是滤食性动物，生活在安静的开阔水域环境中，包括沼泽、池塘和溪流和河流沿岸的漩涡。根据物种的不同，幼虫生活在水柱的不同深度，可以通过在水中添加杀虫剂来控制。苏云金芽孢杆菌以色列亚种的某些形式的细菌毒素(Bti毒素)具有很强的双翅目昆虫活性，并且对蚊子和相关苍蝇幼虫具有特异性。BTI-毒素被认为比所有化学杀虫剂更安全，因为它们对大多数其他昆虫物种以及人类和其他哺乳动物无害。BtBooster(BTB)蛋白已被发现能增强Bt相关毒素的毒性，并通过增加幼虫中肠的毒素摄取而发挥作用。需要一种新的和廉价的BTI和BTB生产和交付系统，以便在不同的水环境和不同的深度接触到不同的蚊子种类。我们的生物技术假设是：在富油作物的种子油体中表达的BTI和BTB可以作为碾磨的Bt面粉有效地输送，它将在水柱中漂浮在不同的水平，并杀死取食蚊子的幼虫。在这个第一阶段的应用中，我们将通过在模式植物拟南芥的富含油的种子中表达Bti和BTB来检验这一假设，拟南芥是油菜籽的近亲。这些种子将被碾磨成面粉，其颗粒大小接近于滤食性蚊子幼虫消耗的食物的大小。漂浮的富含bti和btb的面粉将分散在水中，并喂给各种蚊子物种的幼虫，这些蚊子物种充当疟疾、西尼罗河病毒和黄热病的媒介，并对它们的死亡率进行量化。该项目开发了一种更高效、更便宜、更环保的解决方案，用于生产和交付与Bti相关的毒素和用于控制蚊子的Bt增强剂。计划进行第二阶段的应用，其中bti和btb将用油菜(甘蓝型油菜)表达。表达Bti/BTB的油菜籽可以生产并以较低的成本碾磨成Bt-粉。与公共卫生有关的蚊子是多种血液传播疾病的媒介，包括疟疾、黄热病、西尼罗河病毒、丝虫病、日本脑炎和登革热，全世界每年造成数百万人死亡。该项目旨在开发一种新的、更高效、廉价和环境安全的解决方案，用于生产和传递与Bti相关的毒素和Bt增强剂，用于在幼虫阶段控制蚊子，到达各种水生环境中的不同蚊子物种。它应该通过更好地控制病媒传播的疾病和大量的商业机会，产生与公共卫生相关的重大利益。

DESCRIPTION (provided by applicant): Mosquitoes are the insect vectors for numerous blood-borne human diseases that cause millions of deaths each year worldwide. Mosquito larvae are filter-feeders that inhabit quiet open-water environments including marshes, ponds, and eddies along streams and rivers. Depending upon species, the larvae live at various depths in the water column and may be controlled by adding insecticides to the water. Some forms of bacterial toxins from Bacillus thuringiensis subspecies israelensis (Bti toxin) have strong dipteran activity, and are specific for mosquito and related fly larvae. Bti-toxins are considered safer than all chemical insecticides, because they are harmless to most other insect species and to humans and other mammals. BtBooster (BtB) proteins have been identified that enhance the toxicity of Bt-related toxins, and act by increasing toxin uptake in the larval midgut. A novel and inexpensive system for Bti and BtB production and delivery is needed that will reach diverse mosquito species in various aquatic environments and at various depths. Our biotechnical hypothesis is that: Bti and BtB expressed in seed oil bodies of oil-rich crop plants can be delivered effectively as a milled Bt- flour that will float at various levels in the water column and kill feeding mosquito larvae. In this Phase I application, we will test this hypothesis by expressing Bti and BtB in the oil-rich seeds of the model plant, Arabidopsis thaliana, a close relative of canola. The seeds will be milled into flour with a particle size approximating that of the food consumed by filter-feeding mosquito larvae. The buoyant Bti- and BtB-rich flour will be dispersed on water and fed to larvae of the various mosquito species that act as vectors for malaria, West Nile virus, and yellow fever and their mortality quantified. This project develops a more efficient, inexpensive, and environmentally safe solution to production and delivery of Bti-related toxins and Bt-Boosters for mosquito control. A phase II application is planned in which Bti and BtB will be expressed in canola (Brassica napus). The Bti/BtB expressing canola seeds can be produced and milled into Bt-flour at relatively low cost. PUBLIC HEALTH RELEVANCE Mosquitoes are vectors for numerous blood-born diseases, including malaria, yellow fever, West Nile virus, filariasis, Japanese encephalitis, and dengue fever and worldwide cause millions of deaths annually. This project is designed to develop a novel, more efficient, inexpensive, and environmentally safe solution to production and delivery of Bti-related toxins and Bt-Boosters for mosquito control at the larval stage, reaching diverse mosquito species in various aquatic environments. It should produce significant public health-related benefits through better control of vector-borne disease and substantial commercial opportunities.