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Summary: Female but not male mosquitoes are vectors for multiple deadly human diseases including malaria, dengue, yellow fever, and West Nile virus. However, the underlying neural substrates of sexually dimorphic behaviors remain largely unknown in mosquitoes. In this study, we found striking sexual dimorphism in brain regions in two major disease vectors, Aedes aegypti and Culex quinquefasciatus, through voxel-wise comparison of the whole brain. Female-enlarged regions include those associated with chemosensation and vision, while male-enlarged regions are linked to hearing and memory. However, some brain regions associated with vision and memory are sexually dimorphic in Ae. aegypti but not Cx. quinquefasciatus. As the first global voxel-based comparative neuroanatomical analysis of mosquito brains between sexes, this study not only sheds light on the neural substrates underlying sex-specific behaviors, but also identifies regions of interest for future research to disrupt female-specific behaviors critical to disease transmission.