Cryopreservation of the Endangered Mahseer (Tor khudree) Spermatozoa: Effect of Dimethyl Sulfoxide, Freezing, Activating Media, and Cryostorage on Post-Thaw Spermatozoa Motility and Fertility

The present study was conducted with the goal of evaluating the viability of spermatozoa of the Deccan mahseer (Tor khudree, Cyprinidae) cryopreserved using different strategies. Immotile spermatozoa pooled from 2–4 males were diluted with modified fish Ringer's solution (pH: 7.48) and protected with dimethyl sulfoxide (Me2SO) at 5%, 10%, and 15% and subjected to different equilibration periods. Diluted samples (1:10) were drawn into 500 µL plastic straws and frozen at a distance of 5 cm from the level of liquid nitrogen (LN2) and preserved for 385 days in LN2. Me2SO at 10% resulted in higher post-thaw spermatozoa motility rate (46.7%) than 5% or 15% (up to 33.3%) after 385 days of cryopreservation. Of the different equilibration periods, 20–40 min generally produced higher motility (%) rates than 0, 10, 50, 60, 70, 80, or 90 min. The highest post-thaw motility of spermatozoa was obtained when they were frozen at 2 cm (−120.3°C) above the level of LN2 and the optimum freezing rate was found to be 14.5 ± 0...