Vitrification of epididymal sperm from Iberian ibex (Capra pyrenaica)

Vitrification, a process in which living cells undergo glasslike solidification, is a relatively new cryopreservation method that can successfully preserve the embryos, oocytes, and even the sperm of certain species. For sperm, at least in the species studied so far, kinetic vitrification would appear to be a better alternative. The simplicity of this sperm cryopreservation technique can be useful in field laboratories for wild species because requires less equipment, is much faster, simpler and cost-effective than conventional freezing. The aim of this work was to evaluate comparatively the effectivity of kinetic vitrification and conventional freezing of epididymal sperm from Iberian Ibex (Capra pyrenaica). Testes were obtained from mature ibexes that were legally hunted in the Tejeda and Almijara Game Reserve, in southern Spain (36oN latitude, Province of Malaga, Spain) during the rutting season (November/December 2013/2014). Epididymal spermatozoa were collected by the retrograde flushing method, using 1 mL of Tris-citric acid-glucose medium (TCG) at ambient temperature (11-13oC in the field laboratory). Sperm from left epididymis were frozen with TCG-6% egg yolk and 5% glycerol, and sperm from right epididymis were vitrified with TCG-6% egg yolk with 100 mM sucrose. There weren’t differences between treatments (frozenthawed vs vitrified-warmed sperm) for the percentage of motile sperm, percentage of sperm with membrane integrity determined by the hypo-osmotic swelling test, and percentage of sperm with morphological abnormalities (%). However there were significant differences for quality (score 0-4) of motility (2.4±0.2 and 1.4±0.2), percentage of progressive motility (22.7±4.3 and 7.0±1.6%), percentage of intact acrosome (73.8±4.0 and 55.9±2.5%), percentage of viable sperm according to the nigrosin-eosin staining (45.5±4.1 and 29.2±4.1%), percentage of dead sperm with damaged acrosome (5.5±1.0 and 17.3±2.3%) and percentage of live sperm with intact acrosome (45.1±5.5 and 26.5±4.6) respectively. Although better results were found using the conventional freezing method, given the simplicity of sperm vitrification its use under certain field conditions can be recommended for this type of species. Improvement of the technique might, however, provide better post-vitrification outcomes; new vitrifying solutions and additives should be assessed in future work.