Spermatogenesis is important for male fertility, but has not been well-studied in Opsariichthys bidens, an economically important freshwater fish in China. In this study, there was investigation of the cytological features of spermatogenesis in O. bidens using light microscopy, transmission electron microscopy, and immunofluorescence detection of microtubules. O. bidens has tubular testis. Spermatogenesis in O. bidens is of the cystic type, in which the spermatogenic cells develop into spermatozoa in cysts. There was asynchronous development of primary spermatocytes within a single cyst. Spermiogenesis was classified as Type I, which develops into a Type I aquasperm with an oval nucleus, a small and simple midpiece, a flagellum and no acrosome. There was a nuage in spermatogonia, spermatocytes, and spermatids in different developmental stages of spermatids which may have important functions in fish spermatogenesis. Furthermore, microtubule dynamics may be involved in spermatid reshaping, material transport, and polar distribution of organelles during spermiogenesis.