Development of a gene drive system for locally confined and reversible population replacement in Aedes aegypti using chromosomal translocations

埃及伊蚊是登革热和黄热病等毁灭性疾病的主要传播媒介，这些疾病每年造成数万人死亡，主要是儿童。在登革热的情况下，自20世纪70年代以来，感染率大约每十年翻一番，清楚地表明使用杀虫剂的经典方法未能抑制蚊子种群。因此，迫切需要新的策略来控制这种全球流行病，其中一种方法是使用基因驱动系统。它们可以用来取代伊蚊种群与菌株，阻止病毒的传播。目前为止，还没有一个单一的基因驱动系统已经建立在伊蚊，本文提出的研究目标。驱动系统，我将建立基于染色体易位。如果携带易位的蚊子在野外交配，一部分后代由于染色体不平衡而无法存活。这导致野生型染色体的逐渐丢失和易位等位基因的固定。在所有可用的驱动系统中，选择了基于易位的驱动，因为它是进化稳定的，它不会传播到非靶区域，并且它可以逆转。首先，易位构建体将随机整合到具有piggyBac转座酶的伊蚊基因组中，随后激活诱导双链DNA断裂的连接的归巢核酸内切酶，其在修复时可以形成易位。其次，新的CRISPR/Cas9系统将诱导易位。最后，将在适合度测定中评估所得菌株，并测试其驱动种群更替的潜力。

Aedes aegypti mosquitoes are the main vector for devastating diseases like dengue and yellow fever which are the cause for ten thousands of deaths each year, mainly among children. In the case of dengue fever the infection rates have approximately doubled every ten years since the 1970's, clearly showing that classic approaches using insecticides have failed to suppress the mosquito populations. Hence, novel strategies to control this global epidemic are in dire need.One approach to do this is with gene drive systems. They can be used to replace Aedes populations with strains that block the transmission of viruses. So far not a single gene drive system has been established in Aedes mosquitoes, the goal of the research proposed herein.The drive system I will build is based on chromosomal translocations. If translocation carrying mosquitoes mate into the wild, a proportion of the offspring is unviable due to chromosomal imbalance. This leads to a gradual loss of wildtype chromosomes and fixation of the translocation alleles. Among all the available drive systems, the translocation based drive was chosen because it is evolutionary stable, it does not spread to non-target areas and it can be reversed.To generate Aedes strains carrying translocations, two approaches will be pursued in parallel. First, translocation constructs will be integrated randomly into the Aedes genome with piggyBac transposase, followed by the activation of a linked homing endonuclease that induces double-stranded DNA breaks, which upon repair can form translocations. Second, translocations will be induced with the novel CRISPR/Cas9 system. Finally, the resulting strains will be evaluated in fitness assays and tested for their potential to drive population replacement.