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Ganan, N.; Gonzales, R.; Garde, J.; Martinez, F.; Vargas, A.; Gomendio, M.; Roldan, E.R.S.
Assessment of semen quality, sperm cryopreservation and heterologous IVF in the critically endangered Iberian lynx (_Lynx pardinus_)
2009  Reproduction, Fertility and Development (21): 848-859

Semen traits and factors affecting sperm cryopreservation were assessed in the Iberian lynx (_Lynx pardinus_), a species regarded as the most endangered felid in the world. For cryopreservation, semen was washed, resuspended in a Tes-Tris-based diluent (TEST) or a Tris-based diluent (Biladyl), both with 20% egg yolk and 4% glycerol, loaded into straws, cooled to 5C using an automated programmable system and frozen on nitrogen vapour. Heterologous IVF of _in vitro_-matured domestic cat oocytes was used to test the fertilising ability of cryopreserved spermatozoa. Electroejaculates from five males were obtained. Characterisation of the electroejaculates revealed mean ( s.e.m.) values of 3.3 0.6 106 total spermatozoa, 73.6 4.6% motile spermatozoa, 23.7 4.0% morphologically normal spermatozoa and 40.7 2.3% spermatozoa with intact acrosomes. After thawing a higher percentage of motile spermatozoa was seen in TEST than in Biladyl (34.0 6.2% _v._ 7.5 4.8%, respectively; _P_ < 0.05); however, there were no differences in the percentage of intact acrosomes between the two diluents. Iberian lynx spermatozoa fertilised domestic cat oocytes _in vitro_, with higher fertilisation rates observed for spermatozoa cryopreserved in TEST than in Biladyl, although the difference did not reach statistical significance (20.5 4.5% _v._ 11.5 6.8%, respectively). There were positive significant relations between the fertilisation rates and both the percentage of normal spermatozoa and the percentage of spermatozoa with an intact acrosome before cryopreservation (_P_ = 0.04). This first report of the collection and cryopreservation of Iberian lynx semen and analysis of fertilising ability is an important step in the development of assisted reproductive techniques for this critically endangered felid species. Semen traits and factors affecting sperm cryopreservation were assessed in the Iberian lynx (_Lynx pardinus_), a species regarded as the most endangered felid in the world. For cryopreservation, semen was washed, resuspended in a Tes-Tris-based diluent (TEST) or a Tris-based diluent (Biladyl), both with 20% egg yolk and 4% glycerol, loaded into straws, cooled to 5C using an automated programmable system and frozen on nitrogen vapour. Heterologous IVF of _in vitro_-matured domestic cat oocytes was used to test the fertilising ability of cryopreserved spermatozoa. Electroejaculates from five males were obtained. Characterisation of the electroejaculates revealed mean ( s.e.m.) values of 3.3 0.6 106 total spermatozoa, 73.6 4.6% motile spermatozoa, 23.7 4.0% morphologically normal spermatozoa and 40.7 2.3% spermatozoa with intact acrosomes. After thawing a higher percentage of motile spermatozoa was seen in TEST than in Biladyl (34.0 6.2% _v._ 7.5 4.8%, respectively; _P_ < 0.05); however, there were no differences in the percentage of intact acrosomes between the two diluents. Iberian lynx spermatozoa fertilised domestic cat oocytes _in vitro_, with higher fertilisation rates observed for spermatozoa cryopreserved in TEST than in Biladyl, although the difference did not reach statistical significance (20.5 4.5% _v._ 11.5 6.8%, respectively). There were positive significant relations between the fertilisation rates and both the percentage of normal spermatozoa and the percentage of spermatozoa with an intact acrosome before cryopreservation (_P_ = 0.04). This first report of the collection and cryopreservation of Iberian lynx semen and analysis of fertilising ability is an important step in the development of assisted reproductive techniques for this critically endangered felid species.

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