Effect of Cryopreservation on Arabian Stallion Semen After Adding INRA96 and DMF Based Extenders

Cryopreservation is very important technique in AI centers of stallions, it preserves sperms for long period and spread the superior genetic merits between different animal’s breeds. However, using of cryopreserved sperms lead to decreasing the fertility between animals, due to lethal damage of sperms during preservation process; so, this study aimed to use the two different freezing media with decreasing post thawing sperm damage. A total of 54 ejaculates were collected from nine pure fertile Egyptian Stallions (6 ejaculates per stallion), individually housed at a Veterinary Clinic in Giza Government. Semen sample was collected by Missouri AV on a regular basis (two collections ⁄ week) during the 2021 breeding season in presence of teaser mare. The collected ejaculates were sent to the laboratory immediately for evaluation by CASA (total concentration, progressive motility, static motility, and sperm abnormalities). Ejaculates were filtrated for removal gel fraction; filtrated ejaculates were diluted by EDTA-glucose media for centrifugation and the resulting sperm pellets were split into 2 equal aliquots and then extended in freezing media. INRA96 Milk-based Extender with glycerol and Egg yolk- based Extender with DMF (dimethylformamide) were used in this study as freezing extenders. Diluted semen was packaged into 0.5 ml straw then cooling at 4 ◦C and freezing by vapor of liquid nitrogen and after that preserved by freezing in liquid nitrogen container at -196⁰ C. After keeping the frozen straws in liquid nitrogen for one week, at least 2 straws were taken for thawing and evaluating post thawing-freezing motility. Finally thawed-frozen semen was inseminated inside fertile mares for calculation the conception rate after one month. Post thawing motility were evaluated in extended semen by two different extenders. The obtained results showed a change in the motility by decreasing in INRA-diluted semen compared to DMF-diluted semen. Conception rate was recorded after insemination and showed a high significant in DMF-diluted semen than INRA diluted semen. We concluded that the frozen semen with DMF based diluent did not decrease the motility of sperms after thawing and achieved high conception rate when compared with INRA based glycerol diluent.