Fertility after insemination with frozen-thawed sperm using N-methylacetamide extender on the Combatiente Español avian breed

The aim of this study was to evaluate the in vitro and in vivo quality of frozen-thawed sperm obtained from the Combatiente Español avian breed, with sperm having previously been diluted in N-methylacetamide (NMA). Experimental groups were established: fresh control semen (C); semen diluted without cryoprotectant (T1); semen diluted with extender containing NMA (T2); frozen-thawed sperm (with NMA) containing 500 × 10⁶ spermatozoa (T3); frozen-thawed sperm (with NMA) containing 250 × 10⁶ spermatozoa (T4). In the different groups, sperm motility and viability were assessed using a computer-assisted semen analyzer and flow cytometer, respectively. To evaluate the fertilizing capacity of the sperm, the percentage of fertile eggs was determined. The fertility rate after insemination with frozen-thawed semen was poor, and the concentration of the inseminating dose did not affect fertility rate (9.4 ± 2.7% and 7.0 ± 2.3%, respectively). The results indicate insemination using diluted semen without CPA leads to a reduced fertility, and the addition of 9% NMA to the extender has a greater negative effect on this in vivo variable. Furthermore, inclusion of NMA in the freezing-thawing processes reduced capacity of sperm for fertilization. Sperm viability was reduced during the freezing process, and the dilution in NMA extender affected both sperm viability and motility. The results indicate rooster fertility is negatively affected by sperm dilution, NMA addition and the frozen-thawed effects. Frozen-thawed sperm from Combatiente Español roosters maintained fertilizing capacity for no more than 6 days after insemination, whereas for fresh sperm this capacity was maintained for 14 days.