BACKGROUND: Cryopreservation is a common practice to preserve fish sperm for prolonged periods. OBJECTIVE: To examine the effect of different freezing protocols on sperm characteristics, fertilization, and hatching rate of turbot. MATERIALS AND METHODS: Milt was obtained from ten 8-year-old turbot (54.3 ± 1.7 cm in length and 3,106 ±283 g in weight) at the peak of spawning season. Six batches of milts with >90% motility was pooled and diluted to 1:3 by adding dimethyl sulfoxide (DMSO, 10%) as cryoprotectant. Then straws filled with semen were subjected to three freezing protocols (cooling rates). Sperm characteristics were assessed using sperm class analyzer before and after cryopreservation. Cryopreserved and fresh sperm were used for artificial fertilization to assess fertilization and hatching rates. RESULTS: Cryopreservation protocol has significantly deleterious effects on total motility, progressive motility, curvilinear velocity, straight line velocity, average path velocity, linearity index, straightness index, oscillation index, and amplitude of lateral head displacement of sperm. However, the beat frequency of cryopreserved sperm was found to be similar to control sperm. The fertilization rate of sperm subjected to three freezing protocols were similar, varying between 65.3 % and 75.6 %, and the hatching rates varied from 51.2% to 70.7%. CONCLUSION: The results show the potential application of cryopreservation in fish hatcheries.

Keywords: cryopreservation; sperm motility; sperm quality; turbot.