Seminal Fluid Adipokinetic Hormone Increases Insemination Refractoriness in Female <i>Aedes aegypti</i>

Mating often changes behavior and physiology of female insects. In many species, these changes have been attributed to receipt of seminal fluid molecules (SFMs). SFMs influence phenotypes including feeding, egg production, and response to male courtship and insemination attempts. These same phenotypes are potential targets for management of insect pests. <i>Aedes aegypti</i> mosquitoes are the primary vector of several pathogens including dengue, Zika, and chikungunya. Within an hour after an initial insemination, female <i>Ae. aegypti</i> are generally refractory to subsequent inseminations, a response attributed to SFMs. However, the specific molecules involved in inducing long-term insemination refractoriness have not been identified. In a previous study, we identified adipokinetic hormone (AKH) precursor protein as an SFM in <i>Ae. albopictus.</i> AKH is a well-studied insect neuropeptide that impacts phenotypes including those related to metabolism, locomotion, and reproduction. In this study, we investigated whether AKH is an SFM in <i>Ae. aegypti</i> and whether it impacts female re-insemination patterns. We first established that AKH is produced in the male reproductive tract and transferred to females during mating, and is, therefore, an SFM. We then created an AKH-null line which allowed us to demonstrate that seminal fluid AKH contributes to long-term insemination refractoriness of females. Together, these results have established a novel expression pattern for AKH and identified AKH as a contributor to <i>Ae. aegypti</i> insemination refractoriness, laying the groundwork for understanding the evolution and mode of action of novel seminal fluid proteins as well as for investigating novel pathways or approaches for mosquito control.