Towards the Use of Novel High Density Anopheles-Specific Wolbachia Strains for Anopheles Vector Control.

在最近的一份报告中，世界卫生组织估计全球有2.47亿例疟疾病例，其中61.9万人死亡。鉴于已确立的病媒控制方法对减少全世界疟疾负担作出的巨大贡献，最近报告的病媒控制方法效力下降的情况特别令人关切。亚洲疟蚊，即斯氏按蚊，一种适应性极强的城市病媒进入撒哈拉以南非洲，对消除疟疾造成了额外的障碍。事实证明，以沃尔巴克氏体为基础的干预措施在针对虫媒病毒的主要载体伊蚊时非常有效。在最近发表的一项聚类随机对照试验中，释放感染沃尔巴克氏体的蚊子与登革热发病率降低77%和登革热住院率降低86%相关，这不仅表明基于沃尔巴克氏体的病媒控制策略的可行性，而且还表明沃尔巴克氏体研究的转化性质。鉴于疟疾是绝大多数与蚊子传播的疾病相关的发病率和死亡率的罪魁祸首，人们对开发基于沃尔巴克氏体的疟疾干预措施非常感兴趣也就不足为奇了。可行的干预措施的主要障碍是难以产生感染沃尔巴克氏体的按蚊，已知唯一产生稳定感染的按蚊携带沉重的适应成本。直到最近，人们还认为野生按蚊种群没有沃尔巴克氏体感染，尽管许多种类的蚊子和50%的昆虫被认为感染了沃尔巴克氏体。最近首次发现了两种确认的按蚊特异性沃尔巴克氏体菌株，wAnD和wAnM在两个载体An。demeilloni和An。研究人员预计，使用按蚊特异性菌株传播沃尔巴克氏体的按蚊将会更成功。拟议的项目存在于这些菌株的发现和它们作为干预措施的使用之间的结合点。我将调查按蚊和沃尔巴克氏体之间的独特关系，旨在确定影响按蚊对沃尔巴克氏体易感性的一些因素，同时迈出利用这些菌株控制疟疾的第一步。目的1：识别和减轻体外沃尔巴克氏体感染的限制因素1.1建立机制模型框架，以更好地了解限制体外沃尔巴克氏体感染成功的因素。1.2在已开发的框架内使用高通量检测来确定限制细胞沃尔巴克氏体感染的因素。1.3比较wAnD、wNo和wAlbB感染伊蚊和按蚊细胞的能力1.4制定沃尔巴克氏体感染细胞系的优化方案1.5用感兴趣的沃尔巴克氏体菌株产生稳定的细胞系。目的2胚胎发育过程中宿主-沃尔巴克氏体相互作用的体内分析2.1胚胎早期发育过程中沃尔巴克氏体的嗜性特征表征2.2胚胎微注射后沃尔巴克氏体的嗜性特征表征2.3比较胚胎微注射与沃尔巴克氏体的宿主反应2.4比较不同来源沃尔巴克氏体微注射在蚊子体内的宿主反应。目的3研究自然感染按蚊种群中沃尔巴克氏体宿主的相互作用。3.2评估自然感染豚鼠后代的宿主反应。moucheti母亲

T1 - Basic Research T2 - Human/Clinical ResearchIn its most recent report, the World Health Organisation estimated 247 million cases of malaria, with 619,000 associated deaths worldwide. The recently reported reduction in efficacy of established vector control methods is of particular concern given the vast contribution they have made to the reduction in malaria burden worldwide. The introduction of the Asian Malaria Mosquito, Anopheles stephensi, a highly adaptable urban vector into sub-Saharan Africa presents an additional barrier to malaria elimination. Wolbachia-based interventions have proven highly effective when targeting Aedes mosquitoes, the prominent vectors of arboviruses. In a recently published cluster randomised control trial, the release of Wolbachia infected mosquitoes was associated with an 77% reduction in dengue incidence and an 86% reduction in dengue hospitalisations demonstrating not only the viability of a Wolbachia based vector control strategy but also the translational nature of Wolbachia research. Given that malaria is responsible for the vast majority of mosquito-borne disease related morbidity and mortality, it is unsurprising that there has been great interest in developing a Wolbachia based malaria intervention. The main barrier to a viable intervention has been the difficulty of producing a Wolbachia infected Anopheles mosquito, with the only stable transinfection generated, known to carry a heavy fitness cost. Until recently, wild populations of Anopheles mosquitoes were believed to be free of Wolbachia infections, despite many species of mosquito and 50% of all insect species thought to be infected. The recent discovery of the first two confirmed Anopheles-specific Wolbachia strains, wAnD and wAnM in two vectors An. demeilloni and An. moucheti respectively, has reinvigorated research, with the expectation that Anopheles mosquito Wolbachia transinfections using Anopheles-specific strains will prove more successful. The proposed project exists in the juncture between the discovery of these strains, and their use as an intervention. I will investigate the unique relationship between Anopheles mosquitoes and Wolbachia, aiming to determine some of the factors that influence the susceptibility of Anopheles to Wolbachia while taking the first steps towards the use of these strains for malaria control. Aim 1Identifying and mitigating factors limiting Wolbachia infection in-vitro1.1 Develop the framework for a mechanistic model to better understand factors limiting the success of in-vitro Wolbachia infection. 1.2 Use high throughput testing within the developed framework to identify the factors that limit Wolbachia infection of cells. 1.3 Compare the ability of wAnD, wNo and wAlbB to infect Aedes and Anopheles cells1.4 Develop an optimised protocol for infecting cell lines with Wolbachia 1.5 Generate cell lines stable infected with Wolbachia strains of interest.Aim 2In-vivo analysis of Host-Wolbachia Interactions During Embryonic Development 2.1 Characterising Wolbachia tropism during early embryonic development2.2 Characterising Wolbachia tropism following embryonic microinjection.2.3 Compare the host response upon embryonic microinjection with Wolbachia.2.4 Compare host response in mosquitoes upon microinjection of Wolbachia from different sources.Aim 3Characterise Wolbachia host interaction in naturally infected populations of Anopheles 3.1 Characterise Wolbachia tropism in wild-caught An. moucheti 3.2 Assess host response in offspring of naturally infected An. moucheti mothers