Effect of warming regimen on canine epididymal spermatozoa following vitrification using a coconut water extender supplemented with egg yolk

BACKGROUND: Preservation of canine genetic material through epididymal sperm recovery and vitrification has gained increasing importance in breeding programs and germplasm conservation. The warming procedure represents a critical step affecting post-vitrification sperm survival and functionality. This study evaluated the effect of two warming temperatures on the quality of vitrified canine epididymal spermatozoa. METHODS: Epididymal spermatozoa were collected from twelve clinically healthy dogs undergoing routine orchiectomy. Samples were vitrified using a coconut water-based extender supplemented with egg yolk and subsequently warmed at either 37 °C or 42 °C for 2 min. Sperm quality was evaluated before vitrification and after warming by assessment of motility, viability, morphology, and kinematic parameters using computer-assisted sperm analysis (CASA). RESULTS: Vitrification significantly reduced sperm quality compared with fresh samples. However, warming at 37 °C resulted in significantly higher total motility, viability, and percentage of morphologically normal spermatozoa compared with warming at 42 °C. Several kinematic parameters were also better preserved following warming at 37 °C. CONCLUSIONS: The warming protocol significantly influences post-vitrification quality of canine epididymal spermatozoa. Warming at 37 °C for 2 min provided superior preservation of sperm function compared with warming at 42 °C and may represent a practical approach for canine germplasm banking and assisted reproduction.