Aquaculture Research Station, Louisiana Agricultural Experiment Station, Louisiana State University Agricultural Center, Baton Rouge, LA 70803, USA.
Cryobiology. 2011 Feb;62(1):74-82. doi: 10.1016/j.cryobiol.2010.12.006. Epub 2010 Dec 19.
Hybrid catfish created by crossing of female channel catfish (Ictalurus punctatus) and male blue catfish (Ictalurus furcatus) are being used increasingly in foodfish aquaculture because of their fast growth and efficient food conversion. However, the availability of blue catfish males is limited, and their peak spawning is at a different time than that of the channel catfish. As such, cryopreservation of sperm of blue catfish could improve production of hybrid catfish, and has been studied in the laboratory and tested for feasibility in a commercial dairy bull cryopreservation facility. However, an approach for commercially relevant production of cryopreserved blue catfish sperm is still needed. The goal of this study was to develop practical approaches for commercial-scale sperm cryopreservation of blue catfish by use of an automated high-throughput system (MAPI, CryoBioSystem Co.). The objectives were to: (1) refine cooling rate and cryoprotectant concentration, and evaluate their interactions; (2) evaluate the effect of sperm concentration on cryopreservation; (3) refine cryoprotectant concentration based on the highest effective sperm concentration; (4) compare the effect of thawing samples at 20 or 40°C; (5) evaluate the fertility of thawed sperm at a research scale by fertilizing with channel catfish eggs; (6) test the post-thaw motility and fertility of sperm from individual males in a commercial setting, and (7) test for correlation of cryopreservation results with biological indices used for male evaluation. The optimal cooling rate was 5°C/min (Micro Digitcool, IMV) for high-throughput cryopreservation using CBS high-biosecurity 0.5-ml straws with 10% methanol, and a concentration of 1×10(9)sperm/ml. There was no difference in post-thaw motility when samples were thawed at 20°C for 40s or 40°C for 20s. After fertilization, the percentage of neurulation (Stage V embryos) was 80±21%, and percentage of embryonic mobility (Stage VI embryo) was 51±22%. There was a significant difference among the neurulation values produced by thawed blue catfish sperm, fresh blue catfish sperm (P=0.010) and channel catfish sperm (P=0.023), but not for Stage VI embryos (P≥0.585). Cryopreserved sperm from ten males did not show significant variation in post-thaw motility or fertility at the neurulation stage. This study demonstrates that the protocol established for high-throughput cryopreservation of blue catfish sperm can provide commercially relevant quantities and quality of sperm with stable fertility for hybrid catfish production and provides a model for establishment of commercial-scale approaches for other aquatic species.
通过将雌性斑点叉尾鮰（Ictalurus punctatus）和雄性蓝蟹（Ictalurus furcatus）杂交而培育的杂交鲶鱼，由于其生长迅速和高效的食物转化，在食用鱼养殖中越来越受欢迎。然而，蓝蟹雄性的供应量有限，其产卵高峰期与斑点叉尾鮰不同。因此，蓝蟹精子的冷冻保存可以提高杂交鲶鱼的产量，并已在实验室进行了研究，并在商业奶牛冷冻保存设施中进行了可行性测试。然而，仍然需要一种商业相关的生产冷冻保存的蓝蟹精子的方法。本研究的目的是通过使用自动化高通量系统（MAPI，CryoBioSystem Co.）开发商业规模蓝蟹精子冷冻保存的实用方法。目标是：（1）改进冷却速度和冷冻保护剂浓度，并评估它们的相互作用；（2）评估精子浓度对冷冻保存的影响；（3）根据最高有效精子浓度优化冷冻保护剂浓度；（4）比较在 20°C 或 40°C 下解冻样品的效果；（5）在研究规模下通过用斑点叉尾鮰卵受精来评估解冻精子的受精能力；（6）在商业环境中测试个体雄性精子的解冻后活力和受精能力；（7）测试冷冻保存结果与用于雄性评估的生物学指标的相关性。使用 CBS 高生物安全性 0.5-ml 吸管和 10%甲醇以 5°C/min 的最佳冷却速率（Micro Digitcool，IMV）进行高通量冷冻保存，精子浓度为 1×10(9)sperm/ml。当样品在 20°C 解冻 40s 或 40°C 解冻 20s 时，解冻后活力没有差异。受精后，神经形成（V 期胚胎）的百分比为 80±21%，胚胎运动（VI 期胚胎）的百分比为 51±22%。解冻蓝蟹精子的神经形成值之间存在显著差异，新鲜蓝蟹精子（P=0.010）和斑点叉尾鮰精子（P=0.023），但 VI 期胚胎没有差异（P≥0.585）。来自十个雄性的冷冻保存精子在解冻后活力或神经形成阶段的受精能力没有显示出显著差异。本研究表明，为蓝蟹精子高通量冷冻保存建立的方案可为杂交鲶鱼的生产提供具有稳定生育能力的商业相关数量和质量的精子，并为其他水生物种建立商业规模方法提供模型。
