Bansal Krishna, Jhamb Dinesh, Yadav Usha, Meel Mamta, Gupta Akanksha, Bala Renu, Virmani Meenakshi, Punetha Meeti, Kumar Dharmendra, Kumar Pradeep
Animal Physiology and Reproduction Division, ICAR-Central Institute for Research on Buffaloes, Hisar, Haryana, India.
Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, Rajasthan University of Veterinary and Animal Sciences, Bikaner, Rajasthan, India.
Front Vet Sci. 2025 Jul 15;12:1594298. doi: 10.3389/fvets.2025.1594298. eCollection 2025.
For cattle bulls, only ejaculates with a sperm concentration of 500 million/mL or higher are selected for sperm cryopreservation. There is no established ejaculate's minimum sperm concentration threshold for buffalo semen cryopreservation. Therefore, the first objective of the study was to determine the percentage of ejaculates of buffalo bulls having concentrations lower than 500 million/mL, the percentage of ejaculates suitable for cryopreservation across different concentrations of donated ejaculates, and to estimate the impact of ejaculate concentrations on the production of semen doses. The second objective was to evaluate the post-thaw sperm quality of ejaculates that initially met the criteria for cryopreservation. After analyzing 5,347 ejaculates from 31 buffalo bulls, it was found that 9.96% of the buffalo semen ejaculates had a sperm concentration of less than 500 million/mL. Of these, 48% qualified the minimum criteria, mainly based on sperm motility. The maximum semen doses (8.79%) were produced by the ejaculates with a sperm concentration between 1,101 and 1,201 million/mL. For the second objective of the study, the ejaculates were categorized into four groups: <300, 300-400, >400- < 500, and ≥ 500 million sperm/mL, diluted to a final concentration of 80 million sperm/mL and cryopreserved. The frozen-thawed sperm of different groups were evaluated by a computer-assisted sperm analyser (CASA) and a flow cytometer. Remarkably, it was found that sperm motility, integrity of plasma and acrosome membrane, and mitochondrial membrane potential (MMP) significantly decreased ( < 0.05) when the ejaculates of category <300 were cryopreserved, despite being qualified in subjective motility analysis compared to the other categories. Except for the category <300, in all the other three categories, there was no significant difference ( > 0.05) in the percentage of spermatozoa in MMP. The highest and lowest percentages of spermatozoa with high mitochondrial superoxide were found in categories <300 and ≥500, respectively. In conclusion, buffalo semen ejaculates with a concentration greater than 300 million sperm/mL should be considered for cryopreservation.
对于种公牛，只有精子浓度达到5亿/mL及以上的射精精液才会被选用于精子冷冻保存。水牛精液冷冻保存尚无既定的射精精液最低精子浓度阈值。因此，本研究的首要目标是确定水牛种公牛射精精液中精子浓度低于5亿/mL的比例、不同捐赠射精精液浓度下适合冷冻保存的射精精液比例，并评估射精精液浓度对精液剂量生产的影响。第二个目标是评估最初符合冷冻保存标准的射精精液解冻后的精子质量。在分析了31头水牛种公牛的5347份射精精液后发现，9.96%的水牛射精精液精子浓度低于5亿/mL。其中，48%符合最低标准，主要依据精子活力。精子浓度在11.01亿至12.01亿/mL之间的射精精液产生的精液剂量最多（8.79%）。对于本研究的第二个目标，射精精液被分为四组：<3亿、3亿 - 4亿、>4亿 - <5亿和≥5亿精子/mL，稀释至最终浓度8000万精子/mL并进行冷冻保存。通过计算机辅助精子分析仪（CASA）和流式细胞仪对不同组的冻融精子进行评估。值得注意的是，尽管与其他类别相比，<3亿组的射精精液在主观活力分析中合格，但当对其进行冷冻保存时，精子活力、质膜和顶体膜完整性以及线粒体膜电位（MMP）显著下降（<0.05）。除<3亿组外，其他三个类别中MMP阳性精子百分比无显著差异（>0.05）。线粒体超氧化物含量最高和最低的精子百分比分别出现在<3亿组和≥5亿组中。总之，精子浓度大于3亿/mL的水牛射精精液应考虑用于冷冻保存。
