No abstract

Over the years, the natural stocks of Tor khudree (Cyprinidae) have depleted due to anthropogenic activities and hence it is considered a threatened species in India. Several in situ and ex situ conservation strategies have been suggested for the revival of T. khudree stocks. The total volume of milt obtained from hormone-injected (gonadotropin-releasing hormone analogue1domperidone) T. khudree were signi¢cantly higher (six to seven times) than that of uninjected ¢sh. The total number of spermatozoa per ¢sh was also higher in injected ¢sh (6.5 Â 10 8^7 .6 Â 10 8 ) than that obtained from uninjected counterparts (1.3 Â 10 8^1 .8 Â 10 8 ). On the contrary, the spermatozoa density and spermatocrit were found to be lower in injected ¢sh than that of the controls. Spermatozoa density and spermatocrit ranged between 4.1 Â 10 8^4 .4 Â 10 8 spermatozoa mL À 1 and 38.1^39.4%, respectively, in injected ¢sh, whereas the ¢gures £uctuated between 6.0 Â 10 8^7 .8 Â 10 8 spermatozoa mL À 1 and 61.5^63.1%, respectively, in uninjected ¢sh. However, there was no signi¢cant di¡erence in the spermatozoa motility rates between experimental and control ¢sh. Di¡erent spermatozoa-activating media revealed no signi¢cant di¡erence in spermatozoa motility between hormoneinjected and uninjected mahseers. But motility duration was the longest with NaCl1urea (190^193 s) and the shortest with tap water (50^55 s) in the experimental and control groups. Short-term preservation of the spermatozoa of T. khudree indicated that spermatozoa stored at 4 1C had higher motility rates than those preserved at room temperature either in the presence or absence of oxygen.

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 (Me 2 SO) at 5%, 10%, and 15% and subjected to different equilibration periods. Diluted samples (1:10) were drawn into 500 L plastic straws and frozen at a distance of 5 cm from the level of liquid nitrogen (LN 2) and preserved for 385 days in LN 2. Me 2 SO 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 LN 2 and the optimum freezing rate was found to be 14.5 ؎ 0.1°C /min. Freezing spermatozoa at 1 cm (؊160.4°C) or 3 cm (؊103.0°C) or 4 cm (؊77.9°C) from the level of LN 2 resulted in a significant decrease in post-thaw motility. Among the different activating media tested in the presence of different extenders and cryoprotectants, NaCl (0.3%) was found to be the best, followed by glucose (1%), and tap water. In the presence of different concentrations of Me 2 SO, NaCl (0.3%), glucose (1%), and NaCl plus urea (0.3% ؉ 0.4%) generally produced comparable post-thaw motility percentage and duration at all the concentrations of the cryoprotectant. When the spermatozoa were diluted in the Ringer's solution having pH 7.5 and 7.9, cryoprotected with Me 2 SO at 10% (equilibration period: 30 min) and frozen at 2 cm from LN 2 , the motility rates were 43% and 40%, respectively, after 810 days of cryostorage. High fertilization rate (98%) was obtained for spermatozoa cryopreserved up to 780 days and was close to that of fresh spermatozoa (98.5%). However, the hatching rate and fry survival were marginally lower in the treatment than that of control. After 1 year of rearing in ponds, the growth and survival of fish obtained from cryopreserved and fresh spermatozoa were comparable. The fish produced from cryopreserved spermatozoa were as normal as normally produced fish. The importance of cryopreservation of mahseer spermatozoa vis-à-vis establishment of a frozen gene bank is discussed.

No abstract