Abstract
Although several emerging mosquito control technologies are dependent on mass releases of adult males, methods of sex-sorting that can be implemented globally have not yet been established. RNAi screens led to the discovery of siRNA, which targets gamma-glutamyl transpeptidase (GGT), a gene which is well conserved in multiple species of mosquitoes and located at the sex-determining M locus region in Aedes aegypti. Silencing the A. aegypti, Aedes albopictus, Anopheles gambiae, Culex pipiens, and Culex quinquefasciatus GGT genes resulted in female larval death, with no significant impact on male survival. Generation of yeast strains that permitted affordable expression and oral delivery of shRNA corresponding to mosquito GGT genes facilitated larval target gene silencing and generated significantly increased 5 males:1 female adult ratios in each species. Yeast targeting a conserved sequence in Culex GGT genes was incorporated into a larval mass-rearing diet, permitting the generation of fit adult male C. pipiens and C. quinquefasciatus, two species for which labor-intensive manual sex separation had previously been utilized. The results of this study indicate that female-specific yeast-based RNAi larvicides may facilitate global implementation of population-based control strategies that require releases of sterile or genetically modified adult males, and that yeast RNAi strategies can be utilized in various species of mosquitoes that have progressed to different stages of sex chromosome evolution.
Highlights
Mosquito-borne diseases lead to hundreds of thousands of human deaths annually [1]
Given evidence that the A. aegypti M/m locus is tightly linked to developmental genes that confer sex-specific effects [21,22,29], it was hypothesized that the A. aegypti gamma-glutamyl transpeptidase (GGT) gene, which is located on Chromosome 1 and flanks the sex determination M/m locus [17,18], functions as a sex-specific lethal gene
In support of this hypothesis, larval soaking assays performed with small interfering RNAs (siRNAs) #546, which corresponds to the GGT transcript, induced significant larval mortality, resulting in 25 ± 3% of expected adult females (p < 0.001), but had no significant impact on male survival
Summary
Mosquito control is critical for prevention of mosquito-borne illnesses, the emergence of insecticide resistance, concern for the unwanted negative effects of insecticides on non-target organisms, and a general lack of support for mosquito control programs threaten current schemes for managing mosquitoes worldwide [2]. These problems have led to increased interest in alternative control methods, such as the sterile insect technique (SIT), which was proposed decades ago [3] and entails the release of sterile adult males with the goal of reducing large populations of insects. The establishment of sex separation methods is a rate-limiting step in the global deployment of several emerging population-based mosquito control technologies [5]
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