Impact of three field-practical thawing methods on bovine semen motility, viability, and membrane and acrosome integrity at 0 and 2 hours post-thaw.

The effectiveness of artificial insemination with frozen semen depends largely on semen quality, which can be influenced by the thawing method. This study evaluated the effect of three field-practical semen thawing methods on bovine sperm motility, viability, and plasma membrane and acrosome integrity at 0 and 2h post-thaw. The treatments were: T1 (water at 37 °C), T2 (axilla at 36.1°C), and T3 (room temperature at 23.4 °C). A total of 42 frozen semen straws was analyzed. Motility was assessed using a CASA system, viability was evaluated with Hoechst/propidium iodide staining, and plasma membrane and acrosome integrity were examined using the hypoosmotic swelling (HOS) test and Coomassie blue (CB) staining, respectively. Statistical analysis included ANOVA and t-tests. At 0h, thawing using a water bath (T1) or the axilla (T2) resulted in higher (P < 0.05) total motility, progressive motility, rapid motility, and number of rapidly motile sperm compared to thawing at room temperature (T3). Thawing using a water bath also resulted in higher (P<0.05) sperm viability and percentage of HOS+/CB+ cells at 0h than thawing with T2 or T3. At 2h, post-thaw, T1 resulted in superior (P<0.05) total motility, progressive motility, rapid motility, and viability compared with T2 and T3 thawing methods. Holding samples from 0 to 2h reduced sperm quality parameters in all three thawing methods. In conclusion, thawing semen in warm water at 37 °C yielded better sperm quality parameters at both 0 and 2h post-thaw than thawing in the axilla or at room temperature.