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Anatomy of bat genital organs has been conventionally studied by gross and microscopic observations to date. Here, we employ both histological observation and diceCT (diffusible iodine-based contrast-enhanced computed tomography) to study the detailed three-dimensional morphological structure of the male genital organs in bats, using the greater horseshoe bat, Rhinolophus ferrumequinum. This is the first study to three-dimensionally describe the whole reproductive organs of bats in detail. Our highly resolved three-dimensional reconstruction reveals that the male organs of R. ferrumequinum consist of paired testes, epididymides, deferent ducts, and five accessory genital glands. The boundary between the ampullary and vesicular glands has been difficult to identify in previous observations, but our diceCT imaging allowed us to clearly differentiate the two. We found that the ampullary gland is located at the terminal part of the deferent ducts, and the vesicular gland lies distal to the ampullary glands. This species possesses a single and carrot-shaped urethral gland, which is not found in most chiropteran families. The presence of the urethral gland in this species and its secretions suggest that after copulation this species is capable of forming a vaginal plug, which can seal the female’s vaginal orifice to block the entrance of spermatozoa from other males. The presence of the urethral gland and elongated epididymal tail and the fact that some individuals can terminate their hibernation and reactivate imply forced copulation on hibernating females can occur in R. ferrumequinum.