Thesis (Ph.D.) - Cairo University - Faculty of Veterinary Medicine - Department of Theriogenology

Different aspects of cryopreservation of dog semen have been investigated for the first time in Egypt. Studies were conducted on ejaculated semen (38 dogs) and epididymal sperm (20 dogs) during the period from July 2013 till October 2015. Ten experiments were done to determine the effect of non-canine commercial (BullXcell and INRA-96), Tris and milk based extenders, different cryoprotectants viz., glycerol, dimethyl sulfoxide (DMSO), dimethyl formamide, (DMF) ethylene glycol (EG) and propylene glycol (PG), egg yolk from different avian species (chicken, duck, goose and ostrich), replacing of egg yolk in semen extender by low density lipoprotein (LDL), centrifugation, packaging size, freezing rate and thawing rate on sperm cryosurvival, possibility of cryopreservation of epididymal sperm and extent of damage imposed on sperm by freezing and thawing. Results were evaluated by pre-freeze sperm motility, post-thaw sperm motility and viability, acrosomal integrity, hypo-osmotic swelling test and in vitro-fertilization. It was found that, Tris-based extenders were superior to milk-based ones and that BullXcell, could successfully be used for dog semen cryopreservation. In Tris extenders, fructose was superior to glucose. Moreover, addition of trehalose or lecithin to the extender improved the post-thaw semen quality. DMSO had a cryoprotective role comparable to glycerol; both were better than other cryoprotectants. Duck egg yolk improved freeze-thaw semen quality compared to egg yolk from chicken, ostrich or goose. LDL could successfully substitute egg yolk in dog semen extenders. Centrifugation and packaging of semen in 0.25ml or 0.50 ml straws had no significant effect on post-thaw quality. Freezing of semen in LN vapor, 5 cm above the surface of LN for 10 min was superior to freezing at a faster rate. Thawing of frozen semen at 70 OC for 6 sec gave better results than thawing at slower or faster rates. Epididymal sperm could successfully be frozen using the same protocol as that for ejaculated semen. The freezability of semen from individual dogs is important factor in the success of any cryopreservation program. Freeze-thaw associated ultrastructural changes in the head and mid-piece regions of spermatozoa were similar to those found in other animal species. Further studies are required to assess their effect on fertility

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