Wolbachia as an Agent for Population Replacement in Anopheles Gambiae

描述（由申请人提供）：控制疟疾的传统方法的失败刺激了创造用于控制疟疾的转基因蚊子的努力。为了使转基因策略获得成功，必须积极地将转基因以高频率驱入自然按蚊种群。沃尔巴克氏体是与细胞质不相容性（CI）相关的母系遗传的内共生体，即，当受感染的雄性与未受感染的雌性交配时，减少卵孵化。受感染雌性的交配是有生育能力的，无论雄性的感染状况如何。CI赋予受感染的雌性生殖优势，并允许沃尔巴克氏体在昆虫种群中迅速传播到高频率。与沃尔巴克氏体紧密相关的转基因性状有望通过遗传搭便车进入种群，用一个对寄生虫传播难治的种群取代自然种群。虽然沃尔巴克氏体感染在蚊子中很常见，但从未在任何种类的按蚊中观察到。因此，基于沃尔巴克氏体的疟疾控制策略需要人工将感染转移到负责寄生虫传播的按蚊物种中。人工Wolbachia转染已经在几种医学上重要的伊蚊物种中成功，但尚未在按蚊中成功。我们的体外研究表明，冈比亚按蚊的遗传背景是有能力窝藏至少3种不同的沃尔巴克氏体感染，因此，以前不成功的按蚊转染尝试可能是由于技术，而不是一个内在的遗传块感染。我们将使用最近开发的胚胎和成人注射技术，以建立Wolbachia感染冈比亚按蚊，并检查Wolbachia和按蚊之间的相互作用在表型，遗传和人口水平。在拟议的研究结束时，我们将1）用沃尔巴克氏体人工感染冈比亚按蚊，2）确定沃尔巴克氏体感染对先前通过细胞系实验鉴定的候选按蚊基因表达的影响，3）评估沃尔巴克氏体驱动母系遗传转基因进入冈比亚按蚊笼群的潜力。这些结果将为成功部署转基因蚊子控制疟疾奠定基础。疟疾控制的转基因策略需要在自然按蚊种群中高频率地主动传播引入的转基因。沃尔巴克氏体内共生体理论上可以作为一种机制，将基因传播到种群中。我们建议评估转移沃尔巴克氏体感染冈比亚按蚊和评估共生体作为一个候选的转基因驱动系统疟疾控制。

DESCRIPTION (provided by applicant): Failure of traditional methods to control malaria has stimulated efforts to create transgenic mosquitoes for malaria control. Transgenes must be actively driven into natural Anopheles populations to high frequency for transgenic strategies to be successful. Wolbachia are maternally inherited endosymbionts associated with cytoplasmic incompatibility (CI) i.e., reduced egg hatch when an infected male mates with an uninfected female. Matings of infected females are fertile regardless of the infection status of the male. CI confers a reproductive advantage to infected females and allows Wolbachia to spread rapidly through insect populations to high frequency. Transgenic traits tightly linked to Wolbachia are expected to be driven into the population by genetic hitchhiking, replacing the natural population with one that is refractory to parasite transmission. While Wolbachia infections are common in mosquitoes, they have never been observed in any species of Anopheles. Thus, Wolbachia-based strategies for malaria control require the artificial transfer of infection into Anopheles species responsible for parasite transmission. Artificial Wolbachia transfections have succeeded in several medically-important Aedes species but have not yet succeeded in Anopheline mosquitoes. Our in vitro studies show that the Anopheles gambiae genetic background is competent to harbor at least 3 diverse Wolbachia infections, and thus, previous unsuccessful Anopheles transfection attempts are likely due to technique rather than an intrinsic genetic block to infection. We will use recently-developed embryonic and adult injection techniques to establish Wolbachia infections in Anopheles gambiae and examine the interaction between Wolbachia and Anopheles at the phenotypic, genetic and population levels. By the end of the proposed research, we will 1) artificially transfect Anopheles gambiae with Wolbachia, 2) determine the effect of Wolbachia infection on expression of candidate Anopheles genes previously identified by cell line experiments and 3) evaluate the potential for Wolbachia to drive maternally-inherited transgenes into Anopheles gambiae cage populations. These results will lay the foundation for the successful deployment of genetically-modified mosquitoes for malaria control. Transgenic strategies for malaria control require the active spread of introduced transgenes to high frequency in natural Anopheles populations. Wolbachia endosymbionts can theoretically act as a mechanism to spread genes into populations. We propose to evaluate transfer Wolbachia infection into Anopheles gambiae and evaluate the symbiont as a candidate transgene drive system for malaria control.