Project summary Tsetse flies (Diptera: Glossinidae) are the cyclical vectors of African trypanosomes (Trypanosoma brucei spp.) which are the causative agents of fatal Human African Trypanosomiasis (HAT), and Animal African Trypanosomiasis (AAT) in wild and domesticated animals. The taxonomically simple tsetse enteric microbiota is dominated by an obligate bacterial mutualist, Wigglesworthia. A monoculture of intracellular Wigglesworthia is found within a specialized organ, known as the bacteriome, located at the anterior midgut. Several critical roles towards tsetse biology, spanning from essential nutrient provisioning to immunological priming, have been characterized for Wigglesworthia. Importantly, the elimination (or functional disruption) of the tsetse- Wigglesworthia association results in lags to tsetse development and sizeable reductions in reproductive output. Our long-term objective is to expand our understanding of the molecular mechanisms that coordinate tsetse-Wigglesworthia activity. Here, the role of tsetse-generated microRNAs (miRNAs) towards facilitating the metabolic and developmental integration of Wigglesworthia-tsetse symbiosis will be examined. Our research design involves the identification of miRNAs that are differentially expressed within bacteriomes (relative to non-symbiont harboring organs) during tsetse sex-specific development and pregnancy using deep sequencing-based approaches. Additionally, the gene targets of these miRNAs will be initially identified using computational genomics. The effect(s) of disrupting miRNA-275, significantly upregulated within the bacteriomes of pregnant tsetse, towards tsetse fly life history traits, bacteriome integrity and Wigglesworthia density will be characterized. This project aims to unravel fundamental molecular mechanisms that coordinate host-microbiota biology. The disruption of pivotal Wigglesworthia-mediated metabolic processes within tsetse offers novel targets for biocontrol.