Panyaboriban, Saritvich, Pukazhenthi, Budhan S., Brown, Megan E., Crowe, Chris, Lynch, Warren E., Singh, Ram P., Techakumphu, Mongkol, and Songsasen, Nucharin. 2016. "Influence of cooling and thawing conditions and cryoprotectant concentration on frozen-thawed survival of white-naped crane (Antigone vipio) spermatozoa." Cryobiology 73 (2):209-215. https://doi.org/10.1016/j.cryobiol.2016.07.009
To assist in genetic resource management and recovery efforts of the white-naped crane (Antigone vipio), we conducted two experiments to evaluate the effect of cooling condition, thawing rate, and cryoprotectant concentration on sperm survival post-thaw. Semen was collected from four mature males during breeding season (March and April) and evaluated for volume, sperm concentration, motility, and membrane integrity. In Experiment 1, ejaculates (nÂ =Â 8) were diluted with Beltsville Poultry Semen Extender (BPSE) containing 10% dimethylsulfoxide (Me2SO) and frozen using either one (average cooling rateÂ =Â 2.5Â Â°C/min) or two step (average cooling rateÂ =Â 7 and 9Â Â°C/min, respectively) cooling method. The frozen samples were thawed using one of two thawing rates: 37Â Â°C 30Â s vs. 4Â Â°C 1Â min. In Experiment 2, samples were diluted with crane semen extender containing either 6% or 10% Me2SO, frozen using two-step method and then thawed at 37Â Â°C for 30Â s. Both cooling condition (two-stepÂ >Â one-step) and thawing rate (37Â Â°C 30Â sÂ >Â 4Â Â°C 1Â min) impacted sperm motility, progression and kinetic characteristics (PÂ Â 0.05) affect plasma membrane or acrosomal integrity. Concentration of Me2SO did not impact frozen-thaw survival. We conclude that white-naped crane sperm cryopreserved using a combination of two-step cooling and thawing at 37Â Â°C 30Â s was superior to other cooling and thawing combinations regarding to sustaining sperm motility with good motility kinetics. Findings represent the first steps towards the development of effective cryopreservation protocols and establishment of a genome resource bank for this threatened species.