الفهرس 
ACKNOWLEDGEMEN
Semen storage methodsOnly 14 pages are availabe for public view
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Abstract
were included in this study. All experiments were Three experiments conducted at the Research Poultry Farm, Department of Poultry Production, Faculty of Agriculture, Assiut University, Egypt. This study aimed 1) to evaluate semen quality of the local strains, 2) to find the best approach for freezing sperms and 3) to study the ultra-structure of sperm cell after freezing/ thawing process.
Experiment I
Analysis of sperm motility, velocity and morphometry of three Egyptian indigenous chicken strain.
This study aimed to evaluate sperm swimming velocity and other semen characteristics in relation to some morphometric measures (sperm and flagellum length). Also, to measure the concentrations of some ions in seminal plasma in three Egyptian local chicken strains (Dandarawi, Sharkasi ,and Fayoumi). Thirty chicks from each strain were reared on the floor until reaching sexual maturity. Males were reallocated into individual battery cages and given 2 weeks to acclimatize before starting the experiment. Ten adult roosters from each strain were selected and trained for semen collection. Semen was collected by the abdominal massage.
Studied Traits:
1- Ejaculate volume (mL), sperm concentration, motility (%), sperm curve linear velocity (VCL), average path velocity (VAP), straight line velocity (VSL) and straightness (STR) were measured using computer-assisted sperm analysis (CASA) software.
2- The lengths of the entire sperm, head plus mid-piece and flagellum (µm). Also, the head and the flagellum relative lengths and the head: flagellum ratios were calculated.
3- The concentrations of calcium, magnesium, potassium and sodium in seminal plasma were measured.
The obtained results could be summarized as follows:
1- No significant differences were observed among strains in the percentages of motile spermatozoa and in percentages of sperm showing progressive motility. Dandarawi roosters had significantly (p=0.007) higher sperm concentration per mL, and higher percentages of rapid spermatozoa (p=0.001) and smaller percentages of slow sperms (p=0.001) and showed significantly higher values of VCL (p<0.0001), VSL (p<0.0001) and VAP (p<0.0001). Sharkasi roosters returned significantly higher ejaculate volume (p=0.011) and higher percentages of slow swimming spermatozoa (p=0.001).
2- Dandarawi and Sharkasi exhibited longer sperm and flagellum. While there were no significant differences in the length of head plus midpiece among different strains (p<0.05).
3- Chemical composition of seminal plasma revealed higher potassium concentrations in Sharkasi compared to those of Fayomi (p<0.05); while the concentrations in Dandarawi were intermediate.
Experiment II
Effect of using two levels of two different cryoprotectants with or without sucrose on the fertilizing capacity of Dandarawi rooster’s frozen sperm
This experiment was designed to identify the optimal levels of Glycerol (GLY) and Dimethylacetamide (DMA) used as cryoprotectants as well as, to evaluate the effect of adding sucrose as an osmoregulator on the motility and fertilizing ability of the post-thawed Dandarawi rooster spermatozoa.
Twenty adult males and fifty females were used. These birds were housed in individual cages under the photoperiod of 16 h light: 8 h dark and fed a commercial diet. Pooled semen samples were diluted immediately by Lake and Ravie extender 1: 1 (v: v). Diluted semen was divided into five aliquots. The first aliquot was stored at 5°C and served as a control. The remaining aliquots were mixed with two levels of the cryoprotectant either with or without sucrose. Fast freezing (pellets) technique was used with dimethylacetamide, while gradual freezing in straws method was used for glycerol. After thawing, motility, fertility, scientific and commercial hatchability (%) were measured.
In trial 1, treatments were control, 8% GLY, 11% GLY, 8% GLY+ 5 % sucrose and 11% GLY+ 5 % sucrose. No significant differences in post-thawed sperm motility, fertility and hatchability percentages can be attributed to glycerol levels. The presence of sucrose (5%) led to a significant increase (P<0.05) in scientific and commercial hatchability when mixed with both levels of GLY (8 and 11 %). Trial 2) the treatments were control, 5% DMA, 7% DMA, 5% DMA+ 5 % sucrose and 7% DMA+ 5 % sucrose. There were no significant differences between all frozen semen treatments for motility (%). Hens inseminated with frozen semen using 7% DMA had the highest fertility rate (p< 0.01) followed by 5 and 7 % DMA with sucrose; while 5 % DMA treatment gave the lowest fertility rate. The presence of sucrose (5%) led to a significant increase (p<0.01) in scientific and commercial hatchability when mixed with 5 % DMA. While the positive effect of sucrose didn’t appear with the higher level of DMA (7 %).
Experiment III
Ultrastructure of Fresh and Frozen-Thawed Spermatozoa of Dandarawi Chicken.
This experiment was conducted to assess the level of damage occurring to the sperm parts as a result of using different freezing/thawing methods (pellets with dimethylacetamide and slow freezing with glycerol) with or without adding sucrose. The study was achieved using transmission electron microscopy (TEM). Twenty Dandarawi cockerels were selected and used as semen donors. Birds were exposed to the same environmental conditions that were provided in experiment II. Semen was collected from all cockerels, pooled and diluted 1: 1 (v: v) with Lake and Ravie extender. Diluted semen was divided into five aliquots and prepared as the following; an aliquot was fresh semen and served as a control. Aliquots 2 and 3 were frozen in pellets using 5 %dimethylacetamide (DMA) as a cryoprotectant with or without 5% sucrose, respectively. Aliquots 4 and 5 were frozen in straws using 8 % glycerol (GLY) with or without 5% sucrose, respectively.
The results showed that, approximately 66.7 % of spermatozoa frozen with 8 % GLY alone had abnormal and damaged acrosomes, more than double of the ratio observed in spermatozoa frozen with 5 % DMA (30 %). There were no damages in the acrosome region of unfrozen and frozen spermatozoa in the presence of sucrose. Approximately, 15 % of spermatozoa frozen with GLY alone showed excessive granulated nuclear material. These abnormalities were less seen in other frozen treatments and were not observed in unfrozen spermatozoa. Almost all unfrozen spermatozoa appeared to have normal midpiece, while abnormalities were evident and ranged from 18 to 52 % in frozen/thawed spermatozoa using DMA and GLY with or without sucrose. The ratio of the intact tail was greatly decreased (13%) by freezing/thawing procedures using GLY or DMA without sucrose when compared to the unfrozen treatment (92%). However, the addition of sucrose helped in maintaining the integrity of the tail and increased the ratio to reach 30 and 33% for the DMA and GLY treatments, respectively.
In general, varying degrees of damages were observed in the acrosome, nucleus, midpiece and tail for all treatments. The damage was more intensive for the treatments without sucrose. Spermatozoa frozen with GLY showed more cryodamage than DMA for acrosome, nucleus and the midpiece regions. Furthermore, addition of sucrose resulted in protecting the acrosome region against cryo-injures, but this protective effect was less seen in the nucleus and midpiece. Sucrose also lessened the severity of damages occurring to the tail region of the spermatozoon.