Images of original pupae of Zeugodacus cucurbitae (Coquillett) were normalized, grayed, and segmented to identify male and female pupae of this species via machine vision. The image of each pupa was divided into 25 small areas. The differences in surface texture features in each small area within 11 days were compared. The texture characteristics of both male and female pupae were screened by combining the eclosion of both sexes of Z. cucurbitae (Coquillett). Results indicated that the pectinate setae on the abdominal backplane could be used as a basis for the identification of the male and female pupa of Z. cucurbitae (Coquillett). Moreover, machine vision correctly identified these characteristics with an accuracy of 96.0%. This study lays a foundation for the identification of male and female pupae using machine vision and also for the comprehensive control of Z. cucurbitae (Coquillett).