الفهرس | Only 14 pages are availabe for public view |
Abstract Cryopreservation is a non-physiological and complex method that involves a high level of adaptation of biological cells to the thermal and osmotic shocks, occur during the dilution, cooling–freezing as well as the thawing procedures. During cryopreservation, freeze-thaw damages are unavoidable that result in reduced semen quality. The present study was conducted on two native swamp buffalo-bulls and five breeds of bulls (two Holstein, one Simmental, one Brown-Swiss one Angus and one Belgian blue), maintained on the experimental farm of Animal Reproduction Research Institute (ARRI). Their age ranged between 2 and 4.5 years and of body weight 450-700 Kg. Throughout the experimental period (February – March 2021). each animal received a daily ration composed of about 15 kg green berseem, 4-5 kg concentrates mixture in pellet form, and 4-5 kg rice hay. Water was allowed ad libitum. The animals were kept under similar conditions of nutrition and management. The semen was collected by AV early in the morning, once weekly from each bull over a period of six successive weeks. Two successive ejaculates with about 10 min intervals were obtained from each animal and evaluated for individual motility (%), sperm concentration, plasma membrane integrity by HOST, acrosomal status by a dual staining procedure, mitochondrial activity by MTT test and sperm cell DNA integrity by Comet assay. After evaluation , two ejaculates (from each bull) pooled and diluted with a Tris-based egg yolk extender to obtain a concentration of 40×106 sperm/ml. Samples were cooled slowly to 5oC over a period of 1.5 hours and Summary and Conclusions 80 loaded in 0.25 ml straws (IMV, France). The straws placed 4 cm above liquid nitrogen in the vapor phase in a foam box for 15 minutes before being plunged and stored in liquid nitrogen tank until thawing (one week after freezing). Thawing occurred at 37o C in a water bath for 30 seconds. All straws were evaluated for viability index, plasma membrane integrity, acrosomal intactness, mitochondrial activity and DNA integrities. The present study revealed a significant (P < 0.05) decrease in most parameters of cryopreserved semen of buffalo bulls and bull than fresh semen. The evaluation of cryopreserved semen resulted in loss of sperm individual motility (26.67% and 29.86%), membrane integrity (29.17% and 28.25%), acrosomal integrities (17.66% and 27.09%) as well as mitochondrial activity (40.00% and 41.94%) for buffalo bulls and bulls respectively. Moreover, loss of sperm cell with intact DNA were 4.11% &1.82 and DNA in head 1.96 and 1.35% , respectively. However, there is a significant (P < 0.05) increase in DNA of tail (1.95 and 1.35%), tail length(1.51pixels and 2.13) and Olive tail moment ( 0.09 and 0.17) respectively. from this study we can conclude that: Based on the findings of the present study, it was inferred that both buffalo bulls and bulls spermatozoa could be frozen in a satisfactory way. Cryopreservation deserves some changes in motility, functional integrities of sperm membrane, acrosomes, DNA and mitochondrial activity of bubaline as well as bovine spermatozoa. HOS test could be a valuable and practical tool to assess the functional capacity of fresh and frozen-thawed bubaline and bovine sperm Summary and Conclusions 81 cells and could be added in the routine analysis of semen samples for artificial insemination. DNA integrity of both species (as evaluated by Comet assay) was better preserved with the current freezing technique. Moreover, our results indicated that semen from bulls were more vulnerable to the side effects of freezing as compared to those of buffalo bulls in terms of individual motility and acrosomal integrity |