EFFECT OF CRYOPRESERVATION PROTOCOLS ON THE SPERM KINEMATICS AND VITAL SEMEN ATTRIBUTES IN FROZEN-THAWED MARWARI STALLION SPERM

Studies involving the effects of various procedures and techniques of cryopreservation and their impact on conception rates in horse breeding programs have been previously reported in the literature. The current study aims to compare the efficiency of the automated system (controlled-rate freezer) and the conventional vapour freezing system (manual system) for freezing the stallion semen after cooling at 4 °C and to evaluate the efficiency by studying their post-thaw sperm kinematics and sperm characteristics. Sixty-four ejaculates from eight adult fertile stallions were used in this study. Ejaculates from each stallion were collected twice a week for four weeks. Each ejaculate was extended with Lactose-Glucose-DMF extender and the samples were then packed into 0.5 ml straws, which were divided into two parts: one for cryopreservation in a controlled-rate automated freezer and the other for a conventional vapour freezing system. Semen samples were evaluated for sperm motility, liveability, acrosome integrity, plasma-membrane integrity and mitochondrial membrane potential at fresh, equilibrated and frozen-thawed stages of cryopreservation. Though there was non-significant difference in post-thaw total sperm motility and plasma membrane integrity, however, viability, acrosome integrity and mitochondrial membrane integrity of the stallion spermatozoa were significantly (P<0.05) higher in the automated controlled freezing method in comparison to the conventional vapour freezing system. The kinematic parameters were also recorded be significantly (P<0.05) higher in the automated in comparison to the manual system of freezing. In conclusion, the conventional vapour and automated freezing methods allow significant differences in results for sperm kinematic and vital sperm parameters. The conventional vapour freezing system allows cost reduction with the cryopreservation procedure and is easy to handle and transport, making it suitable for use in breeding projects.