Comparison of different methods of semen cryopreservation in Melopsittacus undulatus

Melopsittacus undulatus is largely used as a potential model for assisted reproduction in other endangered parrot species. Semen was collected from nineteen healthy adult males, by massage technique, during the breeding season. After preliminary evaluations through CASA analysis, eight birds were selected as semen donors and five ejaculates of each bird were utilized to evaluate the effect of 10% ethylene glycol (EG) addition, in two times, during 40min of equilibration time at 4°C, and to compare two programmable freezing curves (a rapid and a slow temperature descent rate) and freezing on nitrogen vapors. Diluted semen (modified TALP, pH 8.4) was divided into two aliquots, the first for freezing individual samples, the second to be mixed in a semen pool from the eight birds. Potential inseminating doses of 10μl, containing 1×10⁶ spermatozoa, were frozen. The effect of EG addition on semen motility and kinetic parameters was analyzed and the three freezing methods were compared. EG addition caused a significant decline of semen motility in individual samples, not in semen pools. The three freezing curves resulted in significant differences in thawed-semen parameters, with nitrogen vapors showing the worst results, while the higher total and progressive motility values were obtained with the 'fast' protocol. Thawed-semen pools motility and kinetic parameters appeared lower than those of the individual ejaculates. The differences found between single ejaculates and semen pools emphasize the importance of performing artificial insemination tests in order to assess the in vivo performance of single ejaculates in the parrot species.