Effects of Kinds of Cryoprotectants on the Characteristics of Frozen Fowl Semen

Abstract

The purpose of this study was to evaluate the sperm viability, normal acrosome and mitochondrial activity in the frozen-thawed fowl semen by different cryoprotectants. The experiment was carried out on 10 sexually adult roosters of Ogye. The semen was collected twice a week and pooled semen was diluted 1:1 EK extender containing no cryoprotectant at 5℃. After equilibration for 30 minutes, diluted chicken semen was diluted 1:1 extender containing either 7% dimethylacetamide (DMA), 7% dimethylformamide (DMF) or 7.5% methylacetamide (MA) at final concentration and was put in 0.5 mL plastic straws and frozen for 30 minutes by exposure to liquid nitrogen vapor 4 cm above the surface of liquid nitrogen, followed by plunging into liquid nitrogen. Frozen semen was thawed in water bath at 5℃ for 2 minutes. For cytometric analysis, the frozen-thawed semen was diluted with EK extender to a final concentration of 90 million spermatozoa per mL. Sperm membrane integrity was evaluated as SYBR-14 and propidium iodide (PI). Acrosome integrity was assessed with fluorescein isothiocyanate-labeled PSA and PI. The percentage of mitochondrial function was estimated by using Rhodamine123 (R123) and PI. In conclusion, freezing rooster semen by using 7% DMF as cryoprotectant was significantly highest in rates of survival and mitochondrial function while its rate of damage of acrosome was significantly lowest. As a result, DMF is the cryoprotectant that has the lowest influences on sperm membranes and acrosome integrity. Therefore it could be used for freezing method of animal genetic conservation method for poultry diversity.

Cryopreservation of Rooster Semen Using N-Methylacetamide as Cryoprotective Agent

“…In the present study, sperm viability was similar among the three different concentrations of MA used. The present study findings were in agreement with that of the other reports where there was significant reduction in live sperm after cryopreservation [15][16][17]. A study reported that semen cryopreservation resulted in 34% and 30% sperm viability for 9% and 12% MA concentrations in Korean Oge chicken [15].…”

“…In our study with use of 9%MA there was no fertile eggs. The result is in contrary to other reports where MA was used as cryoprotectant [7,10,[15][16][17]25]. However, the high fertility (>70%) reported in some studies [7,10] could not be replicated in other studies where different breeds of chicken were utilized [15][16][17].…”

Pellet Method of Semen Cryopreservation: Effect of Cryoprotectants, Semen Diluents and Chicken Lines

“…Chalah et al [1] applied a step freezing programme for chicken semen cryopreservation with DMF as a cryoprotectant in plastic vials and obtained 79% fertility. Korean native chicken semen cryopreserved with 7% DMF gave higher live sperm, mitochondrial activity and lower acrosome damage [3]. The present study differs from the earlier reports where semen was vitrified using DMF as cryoprotectant.…”

“…Alternative cryoprotectants belonging to amide group such as dimethylacetamide, dimethylformamide and methylacetamide were used for cryopreserving poultry semen and the semen can be used for insemination without removal of the cryoprotectant. Dimethylformamide (DMF) was used as cryoprotectant for chicken [1][2][3] and guinea fowl semen cryopreservation [4,5]. In these reports, the semen diluted with DMF was cryopreserved in plastic straws or in plastic vials using stepwise freezing protocols.…”

Effect of cryoprotectants and thawing temperatures on chicken sperm quality

“…Apart from the type of cryoprotectant (CPA), also thawing procedure is crucial for avian post‐thaw semen quality (Iaffaldano, Di Iorio, Cerolini, et al., ). Lower semen quality values were observed using a thawing rate of 37°C for 30 s (Santiago‐Moreno et al., ; Choi et al., ; Madeddu et al., ) when compared to 5°C for 2 min (Hanzawa et al., ; Mphaphathi et al., ; Sasaki et al., ).…”