Abstract
Chicken semen conservation is an important tool for programs of genetic diversity management and of endangered breeds' conservation. However, the method still needs to be improved in order to be applied in a wide variety of environments and breeds. Our objective was to compare the effects of 2 external cryoprotectants saccharides (sucrose and raffinose) on the sperm freezability of a Thai local breed, Pradu Hang Dum, in which semen was frozen with a simple freezing method using nitrogen vapors and dimethyl formamide (DMF). Thirty-six males were selected on their motility vigor score for the experiments. In a first experiment, a large range of sucrose and raffinose doses were tested. Semen quality was evaluated after incubation at 5°C or after cryopreservation in straws in the saline Blumberger Hahnen Sperma Verdünner diluent + DMF (6% v/v) with or without sucrose/raffinose. The best targeted doses of sucrose and raffinose were then kept for experiment 2 that was focused on cryopreserved semen. In this experiment, semen quality was measured on frozen-thawed sperm: different objective motility data evaluated by computer-assisted sperm analysis (CASA), membrane integrity, acrosome integrity, mitochondria function evaluated using flow cytometry, lipid peroxide production assessed by the thiobarbituric acid test. Fertility obtained with frozen-thawed semen supplemented or not with sucrose or raffinose was also evaluated after artificial insemination of laying hens. The presence of sucrose at the osmotically inactive dose 1 mmol significantly increased the vigor motility, membrane integrity, acrosome integrity, and mitochondrial functions of frozen-thawed sperm (P < 0.05), and showed the highest levels of fertility after sperm cryopreservation (91% vs. control 86%, P < 0.001). Raffinose showed negative effects on in vitro semen quality from 1 to 100 mmol. Fertility was also negatively (P < 0.001) affected by raffinose (fertility rate 66 to 70%). We thus showed in the present study the high success of a simple chicken sperm cryopreservation method with an external cryoprotectant easily available and cheap, the sucrose, used at an osmotically inactive low concentration.