Abstract：
Fourier harmonic analysis (FHA) is a robust method for identification of minute changes in sperm nuclear shape that are indicative of reduced fertility. The current study was designed to develop a fertility prediction model for Nili-Ravi buffalo bulls through FHA of sperm. In experiment I, FHA technique was standardized, average sperm nuclear perimeter was measured and sperm nuclear shape plot of buffalo bull was constructed. Sperm of buffalo bulls (n = 10) were stained with YOYO-1 and Hoechst-33342 to differentiate live and dead, and digital images were captured using phase contrast and fluorescent microscopy. The images were analyzed by ImageJ software and 100 sperm/bull were evaluated. The results are described as mean ± SEM values of mean harmonic amplitude (mharm), skewness harmonic amplitude (skharm), kurtosis harmonic amplitude (kurharm) and variance harmonic amplitude (varharm) at Fourier frequencies 0-5 along with the cartesian and polar coordinate plots of buffalo bull sperm. In experiment II, a fertility prediction model was developed based on FHA of buffalo bull sperm. Semen samples of low (n = 6), medium (n = 3) and high (n = 8) fertility bulls were investigated for FHA of sperm and harmonic amplitudes (HA) were generated. Firstly, to determine if live and dead sperm population have unique nuclear shape distribution; the mean, skewness, kurtosis and variance HA 0-5 of 1700 live and 1294 dead spermatozoa of 17 bulls were evaluated. T-test signified a difference in the mharm0 (2.363 ± 0.01 vs. 2.439 ± 0.02), skharm0 (-0.0002 ± 0.07 vs. -0.266 ± 0.09), kurharm0 (-0.156 ± 0.07 vs. 0.260 ± 0.18), kurharm2 (0.142 ± 0.11 vs. 1.031 ± 0.32) and varharm4 (0.109 ± 0.00 vs. 0.082 ± 0.00) of live vs. dead sperm population (p < 0.05). Therefore, 100 live sperm/bull were further evaluated for mean, skewness, kurtosis and variance HA 0-5 values among high (n = 6) and low-fertility (n = 6) groups. Results of T-test showed higher values of mharm2 (0.739 ± 0.01 vs. 0.686 ± 0.00), mharm4 (0.105 ± 0.001 vs. 0.007 ± 0.001), and skharm0 (0.214 ± 0.109 vs. -0.244 ± 0.097) in high vs. low-fertility group (p < 0.05). In next step, five significantly different combinations of discriminant measures between high and low-fertility groups were obtained by discriminant analysis. In conclusion, mharm4, skharm0 and varharm2 correctly identified 91.7 % of bulls into their respective fertility groups, and upon cross validation the value of the canonical correlation was 0.928.
摘要：
傅里叶谐波分析（FHA）是一种可靠的方法，用于识别精子核形状的微小变化，这些变化表明生育力降低。本研究旨在通过精子的FHA开发Nili-Ravi水牛公牛的生育力预测模型。在实验I中,FHA技术是标准化的，测量了水牛平均精子核周长，并构建了水牛精子核形状图。水牛公牛的精子（n=10）用YOYO-1和Hoechst-33342染色，以区分活的和死的，使用相衬和荧光显微镜捕获数字图像。通过ImageJ软件分析图像并评估100个精子/公牛。结果描述为平均谐波振幅(mharm)的平均值±SEM值，偏度谐波振幅(skharm)，傅立叶频率0-5的峰度谐波振幅（kurharm）和方差谐波振幅（varharm）以及水牛公牛精子的笛卡尔和极坐标图。在实验二,建立了基于水牛精子FHA的生育力预测模型。低精液样本(n=6)，研究了中等（n=3）和高（n=8）生育力公牛的精子FHA，并产生了谐波振幅（HA）。首先,为了确定活的和死的精子群体是否具有独特的细胞核形状分布；平均值，偏斜度,评估了17头公牛的1700只活精子和1294只死精子的峰度和方差HA0-5。T检验表示mharm0的差异（2.363±0.01与2.439±0.02),skharm0(-0.0002±0.07vs.-0.266±0.09),kurharm0(-0.156±0.07vs.0.260±0.18)，kurharm2(0.142±0.11vs.1.031±0.32)和varharm4(0.109±0.00vs.0.082±0.00)的活vs.精子死亡人数(p<0.05)。因此,进一步评估了100只活精子/公牛的平均值，偏斜度,高（n=6）和低生育率（n=6）组之间的峰度和方差HA0-5值。T检验结果显示mharm2的值较高（0.739±0.01vs.0.686±0.00),mharm4(0.105±0.001vs.0.007±0.001)，和skharm0(0.214±0.109vs.-0.244±0.097）inhighvs.低生育率组(p<0.05)。下一步,通过判别分析获得了高生育率和低生育率组之间5种显著不同的判别组合.总之,mharm4，skharm0和varharm2正确地将91.7％的公牛识别为各自的生育组，交叉验证后，典型相关值为0.928。
