Elucidation of mosquito-malaria parasite interaction using genetically engineered mosquitoes

疟疾的子孢子侵入蚊子的唾液腺，并在唾液管中等待，直到下一次吸血。唾液腺侵袭的过程和分子的机制仍然很大程度上不清楚。为了在斯氏按蚊中建立唾液腺特异的转基因表达系统，我们获得了一个唾液腺特异的启动子，用于编码斯氏按蚊抗血小板聚集蛋白(AAPP)的基因。AAPP启动子是一种女性唾液腺特异的、血液诱导的强启动子。利用该启动子，我们获得了表达丰富的盘状瘤菌的转基因斯氏按蚊。红色荧光蛋白(DsRed)在腺体的远外侧叶中，子孢子优先入侵那里。这些结果为阐明唾液腺-寄生虫的相互作用和培育对寄生虫具有抵抗力的转基因蚊子提供了可能性。最近在按蚊基因工程方面的进展为将其用作…带来了希望这为疟疾控制提供了更多新的战略，并为研究蚊虫与寄生虫的相互作用提供了一个强有力的工具。我们利用转基因蚊子阻断传播的研究主要集中在寄生虫的中肠阶段，干扰动细胞入侵或卵囊分化。我们在转基因蚊子中引入的分子是从刺参(海洋无脊椎动物)中分离出来的CEL-III。作为一种凝集素，CEL-III具有很强的溶血活性和凝血活性。Cel-III基因的表达受中肠特异性启动子--羧肽酶启动子的调控。正如预期的那样，CEL-III在血餐后6h在中肠被诱导表达。我们将对照蚊子和转基因蚊子喂给同一只感染的大鼠，并测量卵囊形成的数量。与对照蚊子相比，转基因蚊子的传播阻断效果为91%。因此，CEL-III在蚊子中肠中的表达通过抑制疟原虫的发育而严重降低了媒介能力。由于CEL-III针对的是红细胞而不是寄生虫，我们预计这种CEL-III转基因蚊子将能够破坏所有疟原虫物种的寄生虫发育。较少

Malaria sporozoites invade the mosquito salivary glands and wait in the salivary duct until the next blood feeding. The mechanisms of the process and molecules involved in the salivary gland invasion remain largely unknown. To establish a robust salivary gland-specific transgene expression in Anopheles stephensi, we obtained a salivary gland-specific promoter for a gene encoding Anopheles anti-platelet aggregation protein (AAPP). The aapp promoter is a female salivary gland-specific and blood meal-inducible strong promoter. Using this promoter, we generated a transgenic An.stephensi expressing abundant Discosoma sp. red fluorescent protein (DsRed) in the distal-lateral lobes of the glands, where the sporozoites invade preferentially. These results open up the possibilities of elucidating salivary gland-parasite interactions and generating transgenic mosquitoes refractory to parasites.Recent advances in the genetic engineering of anopheline mosquitoes have raised hopes for their use as … More new strategies for malaria control and provided a powerful tool to investigate mosquito-parasite interactions. Our research on transmission blockade using transgenic mosquitoes has been focused on the midgut stage of the parasites, interfering with ookinete invasion or oocyst differentiation. The molecule that we have introduced into a transgenic mosquito is CEL-III isolated from Cucumaria echinata (Marine invertebrate). CEL-III has strong hemolytic activity as well as haemaggrutination activity as a lectin. CEL-III gene expression was driven by the carboxypeptidase promoter, midgut specific promoter. As expected, CEL-III expression was induced in the midgut at 6h after a blood meal. We fed control and transgenic mosquitoes on the same infected rat and measure the numbers of oocysts formed. The transmission blocking efficacy of the transgenic mosquitoes was 91%, as compared with control mosquito. Thus, expression of CEL-III in the mosquito midgut severely reduced vector competency by inhibiting Plasmodium development. Because CEL-III targets to red blood cells but not the parasites, we anticipate that this CEL-III transgenic mosquito will be able to impair parasite development of all Plasmodium species. Less