Chicken Semen Cryopreservation and Its Impact on Reproductive Efficiency

Abstract

Aims: The present study aimed to evaluate fertility and hatchability rates of cryopreserved semen from the Horasi chicken ecotype, thereby completing the development of a functional semen cryopreservation protocol for this ecotype. Study Design: Factorial experiment, in a completely randomized design. Place and Duration of Study: Department of Laboratory (semen laboratory), National Artificial Insemination Centre (NAIC), between April and October 2025. Methodology: Semen was collected from Horasi cockerels using abdominal massage technique and examined for motility, viability, and morphology. Assessed semen was pooled and diluted (1:2 ratio) with specific extenders: Beltsville Poultry Semen Extender (BPSE) and CARI each supplemented with 4% of a specific cryoprotectant (dimethyl sulfoxide (DMSO) or dimethyl formamide (DMF)), and packed in 0.5 mL, and 0.25mL French straws before cryopreservation. Thawing was performed under two conditions (5oC for 100 sec, and 37oC for 30 sec), while artificial insemination was carried out using two different semen volumes (0.5mL, and 0.25mL), and frequencies (once or twice weekly). Results: All tested parameters significantly (p<0.05) affected fertility and hatchability outcomes. The optimal combination of BPSE with 4% DMSO, thawed at 5oC for 100 sec, twice weekly inseminated with 0.5mL of semen, resulted in better fertility and hatchability rates (81.27 ± 12.23 and 76.27 ± 8.99, respectively) comparable to fresh semen (84.28 ± 6.52 and 72.66 ± 14.79, respectively). In contrast, the poorest performance (fertility: 10.73 ± 8.04, hatchability: 2.64 ± 4.90) was observed with CARI containing 4% DMF, thawed at 37oC for 30 sec, and inseminated once weekly with 0.25mL of semen. Conclusion: Cryopreservation of chicken semen using BPSE with 4% DMSO followed by thawing at 5oC for 100 sec and two times insemination per week with 0.5mL package resulted into acceptable level of fertility and hatchability in Horasi chicken ecotype. The developed protocol can be used in gene bank conservation and breeding programs to support sustainable use of the Horasi chicken ecotype.