Generally, sexual hormone action or sexual behaviors tend to be suppressed during hibernation in the most hibernating mammals. Recently, we found out that the Japanese little horseshoe bat showed the distension of gonads in males and vaginal opening in females during wintering by examining the annual reproductive cycle. Furthermore, we confirmed the frequent flight of bats within and outside the hibernacula during periodic arousal. In this study, we aim to clarify why reproductive organ and flight activity in the Japanese little horseshoe bat were activated during wintering. Field study was conducted from March 2013 to March 2016 in Osawa cave, Niigata prefecture. Bats within colony were captured every month and then the reproductive statuses of both sexes were examined. The mate behavior of bats during periodic arousal was also observed. In some males, active spermatozoa were retained in cauda epididymis during wintering despite atrophied testis. Males with distended cauda epididymis aroused periodically and copulated with torpid females. Winter copulation may play an important role in increasing fertilization success in addition to autumn mating for the bat species with delayed fertilization.