Llavanera Marc, Ribas-Maynou Jordi, Delgado-Bermúdez Ariadna, Recuero Sandra, Muiño Rodrigo, Hidalgo Carlos O, Tamargo Carolina, Bonet Sergi, Mateo-Otero Yentel, Yeste Marc
Biotechnology of Animal and Human Reproduction (TechnoSperm), Faculty of Sciences, University of Girona, C/ Maria Aurèlia Campany, 69, ES-17003, Girona, Spain.
Department of Biology, Unit of Cell Biology, Faculty of Sciences, University of Girona, ES-17003, Girona, Spain.
J Anim Sci Biotechnol. 2021 Nov 9;12(1):115. doi: 10.1186/s40104-021-00634-7.
Genetic selection in cattle has been directed to increase milk production. This, coupled to the fact that the vast majority of bovine artificial inseminations (AI) are performed using cryopreserved sperm, have led to a reduction of fertility rates over the years. Thus, seeking sensitive and specific sperm biomarkers able to predict fertility rates is of vital importance to improve cattle reproductive efficiency. In humans, sperm chromatin condensation evaluated through chromomycin A3 (CMA3) has recently been purported to be a powerful biomarker for sperm functional status and male infertility. The objectives of the present study were: a) to set up a flow cytometry method for simultaneously evaluating chromatin condensation and sperm viability, and b) to test whether this parameter could be used as a predictor of in vivo fertility in bulls. The study included pools of three independent cryopreserved ejaculates per bull from 25 Holstein males. Reproductive outcomes of each sire were determined by non-return rates, which were used to classify bulls into two groups (highly fertile and subfertile).
Chromatin condensation status of bovine sperm was evaluated through the combination of CMA3 and Yo-Pro-1 staining and flow cytometry. Sperm quality parameters (morphology, viability, total and progressive motility) were also assessed. Pearson correlation coefficients and ROC curves were calculated to assess their capacity to predict in vivo fertility. Sperm morphology, viability and total motility presented an area under the ROC curve (AUC) of 0.54, 0.64 and 0.68, respectively (P > 0.05), and thus were not able to discriminate between fertile and subfertile individuals. Alternatively, while the percentage of progressively motile sperm showed a significant predictive value, with an AUC of 0.73 (P = 0.05), CMA3/Yo-Pro-1 staining even depicted superior results for the prediction of in vivo fertility in bulls. Specifically, the percentage of viable sperm with poor chromatin condensation showed better accuracy and precision to predict in vivo fertility, with an AUC of 0.78 (P = 0.02).
Chromatin condensation evaluated through CMA3/Yo-Pro-1 and flow cytometry is defined here as a more powerful tool than conventional sperm parameters to predict bull in vivo fertility, with a potential ability to maximising the efficiency of dairy breeding industry.
奶牛的基因选择旨在提高产奶量。这与绝大多数牛人工授精（AI）使用冷冻保存的精子这一事实相结合，导致多年来生育率下降。因此，寻找能够预测生育率的敏感且特异的精子生物标志物对于提高奶牛繁殖效率至关重要。在人类中，最近有人声称通过放线菌素 A3（CMA3）评估的精子染色质浓缩是精子功能状态和男性不育的有力生物标志物。本研究的目的是：a）建立一种同时评估染色质浓缩和精子活力的流式细胞术方法，b）测试该参数是否可作为公牛体内生育能力的预测指标。该研究包括来自25头荷斯坦公牛的每头公牛的三个独立冷冻射精样本池。每头种公牛的繁殖结果通过不返情率确定，该指标用于将公牛分为两组（高生育力和低生育力）。
通过CMA3和Yo-Pro-1染色与流式细胞术相结合评估牛精子的染色质浓缩状态。还评估了精子质量参数（形态、活力、总活力和前向运动活力）。计算了Pearson相关系数和ROC曲线以评估它们预测体内生育能力的能力。精子形态、活力和总活力的ROC曲线下面积（AUC）分别为0.54、0.64和0.68（P>0.05），因此无法区分生育力高和生育力低的个体。另外，虽然前向运动精子的百分比显示出显著的预测价值，AUC为0.73（P = 0.05），但CMA3/Yo-Pro-1染色在预测公牛体内生育能力方面甚至表现出更好的结果。具体而言，染色质浓缩差的活精子百分比在预测体内生育能力方面显示出更好的准确性和精确性，AUC为0.78（P = 0.02）。
通过CMA3/Yo-Pro-1和流式细胞术评估的染色质浓缩在此被定义为比传统精子参数更强大的预测公牛体内生育能力的工具，具有最大化奶牛养殖业效率的潜在能力。
