基于计算机辅助精子分析的大口黑鲈精子冷藏保存效果评估

doi:10.3969/j.issn.1004-1524.20240769

浙江农业学报 ›› 2025, Vol. 37 ›› Issue (7): 1417-1423.DOI: 10.3969/j.issn.1004-1524.20240769

基于计算机辅助精子分析的大口黑鲈精子冷藏保存效果评估

李帆¹(), 张晓锋², 李建营³, 陈小明², 刘伟², 周凡², 孟庆辉²^,^*()

1.上海海洋大学水产与生命学院,上海 201306
2.浙江省水产技术推广总站,浙江杭州 311122
3.杭州师范大学生命与环境科学学院,浙江杭州 311121

收稿日期:2024-08-30 出版日期:2025-07-25 发布日期:2025-08-20
作者简介:李帆(2002—),女,浙江岱山人,本科生,研究方向为鱼类生殖生理。E-mail:lifan021228@163.com
通讯作者: ^*孟庆辉, E-mail: qinghui1234@126.com
基金资助:
浙江省农业新品种选育重大科技专项(2021C02069-2);国家特色淡水鱼产业技术体系(CARS-46);浙江省农业重大技术协同推广项目(2022XTTGSC01)

Evaluation of sperm cryopreservation effectiveness for largemouth bass(Micropterus salmoides) based on computer-assisted sperm analysis

LI Fan¹(), ZHANG Xiaofeng², LI Jianying³, CHEN Xiaoming², LIU Wei², ZHOU Fan², MENG Qinghui²^,^*()

1. College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China
2. Zhejiang Fisheries Technical Extension Center, Hangzhou 311122, China
3. College of Life and Environmental Sciences, Hangzhou Normal University, Hangzhou 311121, China

Received:2024-08-30 Online:2025-07-25 Published:2025-08-20

摘要/Abstract

摘要： 为筛选适于大口黑鲈精子的保存液,延长精子保存时间,研究了4种保存液(Cortland生理盐水、Ringer’s生理盐水、斑马鱼E3培养液、PBS溶液)在4 ℃冷藏条件下对大口黑鲈精子活力的影响,使用计算机辅助精子分析系统(CASA)检测原精及精液冷藏期间大口黑鲈的精子活力比例、直线运动速率、曲线运动速率、平均路径运动速率和鞭打频率等指标的变化。结果显示,随着冷藏时间的延长,大口黑鲈的精子活力比例逐步下降。不同保存液冷藏大口黑鲈精子的效果不同,Ringer’s生理盐水、PBS溶液保存条件下的精子活力比例更高,运动参数的表现也更好。用Ringer’s生理盐水保存72 h,精子活力比例仍可达95%以上。这表明,Ringer’s生理盐水适合用作大口黑鲈精子短期冷藏的保存液。

关键词: 大口黑鲈, 计算机辅助精子分析系统, 精子, 冷藏保存

Abstract:

To evaluate and screen out suitable sperm preservation solutions to extend the storage time of largemouth bass (Micropterus salmoides) sperm, the effects of four preservation solutions, i.e. Cortland saline, Ringer’s saline, Zebrafish E3 culture solution, and PBS solution, on sperm motility were compared during the storage at 4 ℃. The changes on key motility parameters, including the percentage of motile (MOT), curvilinear velocity (VCL), straight line velocity (VSL), average path velocity (VAP) and beatingcross frequency (BCF) of the origin sperm and the sperm stored with preservation solution were assessed using a computer-assisted sperm analysis (CASA) system. It was shown that there was a gradual decline in MOT with the extended storage time. The cryopreservation effectiveness of preservation solutions varied. Among the preservation solutions, Ringer’s saline and PBS solution demonstrated superior performance, with higher MOT and other motion parameters. When the sperm was stored with Ringer’s saline, the MOT could maintain over 95% after 72 h. These findings suggest that Ringer’s saline could be used for the short-term refrigeration of largemouth bass sperm.

Key words: largemouth bass (Micropterus salmoides), computer-assisted sperm analysis, sperm, refrigerated preservation

中图分类号:

S961.2

李帆, 张晓锋, 李建营, 陈小明, 刘伟, 周凡, 孟庆辉. 基于计算机辅助精子分析的大口黑鲈精子冷藏保存效果评估[J]. 浙江农业学报, 2025, 37(7): 1417-1423.

LI Fan, ZHANG Xiaofeng, LI Jianying, CHEN Xiaoming, LIU Wei, ZHOU Fan, MENG Qinghui. Evaluation of sperm cryopreservation effectiveness for largemouth bass(Micropterus salmoides) based on computer-assisted sperm analysis[J]. Acta Agriculturae Zhejiangensis, 2025, 37(7): 1417-1423.

图/表 4

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保存液 Preservation solution	NaCl	KCl	MgSO₄·7H₂O	NaHCO₃	NaH₂PO₄·2H₂O	CaCl₂	KH₂PO₄	葡萄糖 Glucose
Cortland	7.250	0.380	0.230	1.000	0.410	0.162	—	1.000
Ringer	6.500	0.140	—	0.200	0.010	0.120	—	—
E3	0.344	0.015	0.098	—	—	0.058	—	—
PBS	8.000	0.200	—	—	1.440	—	0.240	—

保存液 Preservation solution	NaCl	KCl	MgSO₄·7H₂O	NaHCO₃	NaH₂PO₄·2H₂O	CaCl₂	KH₂PO₄	葡萄糖 Glucose
Cortland	7.250	0.380	0.230	1.000	0.410	0.162	—	1.000
Ringer	6.500	0.140	—	0.200	0.010	0.120	—	—
E3	0.344	0.015	0.098	—	—	0.058	—	—
PBS	8.000	0.200	—	—	1.440	—	0.240	—

t/h	MOT/%	STR/%	VCL/(μm·s^-1)	VSL/(μm·s^-1)	VAP/(μm·s^-1)	BCF/Hz	MAD/(°)	DHS/μm
2	99.98±5.24	59.93±2.24	202.06±22.08	46.53±15.64	77.64±18.57	38.82±5.66	20.97±5.76	60.17±15.71
6	96.26±5.61	77.94±6.87	110.29±17.81	45.67±16.32	58.60±16.89	29.30±6.91	19.87±5.23	44.88±11.35
24	80.00±4.77	70.39±8.53	47.00±11.66	15.14±9.47	21.51±8.83	10.76±1.24	11.47±3.43	24.51±8.52
48	63.58±3.26	55.95±3.49	28.54±3.97	5.78±1.87	10.33±2.16	5.16±0.36	7.40±2.17	16.10±3.18
72	54.13±6.31	76.16±9.82	25.26±3.75	9.84±3.24	12.92±2.99	6.46±1.08	6.58±1.11	19.23±2.60

t/h	MOT/%	STR/%	VCL/(μm·s^-1)	VSL/(μm·s^-1)	VAP/(μm·s^-1)	BCF/Hz	MAD/(°)	DHS/μm
2	99.98±5.24	59.93±2.24	202.06±22.08	46.53±15.64	77.64±18.57	38.82±5.66	20.97±5.76	60.17±15.71
6	96.26±5.61	77.94±6.87	110.29±17.81	45.67±16.32	58.60±16.89	29.30±6.91	19.87±5.23	44.88±11.35
24	80.00±4.77	70.39±8.53	47.00±11.66	15.14±9.47	21.51±8.83	10.76±1.24	11.47±3.43	24.51±8.52
48	63.58±3.26	55.95±3.49	28.54±3.97	5.78±1.87	10.33±2.16	5.16±0.36	7.40±2.17	16.10±3.18
72	54.13±6.31	76.16±9.82	25.26±3.75	9.84±3.24	12.92±2.99	6.46±1.08	6.58±1.11	19.23±2.60

保存液	t/h	VCL/	VSL/	VAP/	DHS/μm	MAD/(°)	BCF/Hz	STR/%
Preservation solution		(μm·s^-1)	(μm·s^-1)	(μm·s^-1)
Ringer	2	74.49±14.32	30.43±11.84	39.24±12.33	31.82±18.57	15.90±3.66	19.62±6.16	75.90±5.27
	72	70.78±15.84	26.18±12.38	35.10±13.01	26.46±2.43	15.78±4.08	17.55±6.51	72.08±7.26
PBS	2	78.31±15.61	30.47±15.34	40.04±15.12	29.81±13.07	16.43±2.39	20.02±7.56	73.01±8.89
	72	75.76±13.63	29.52±15.90	38.77±15.40	37.80±12.67	13.77±3.42	19.39±7.70	72.34±9.63
E3	2	27.42±1.79	6.16±0.27	10.41±0.53	40.67±4.67	6.64±0.59	5.20±0.26	59.19±1.37
	72	10.21±0.95^**	1.43±0.27^**	3.18±0.16^**	24.25±4.54^**	2.86±0.47^**	1.59±0.08^**	44.74±7.41^*
Cortland	2	24.81±3.82	5.59±0.53	9.44±1.15	38.01±7.47	5.31±0.51	4.72±0.57	59.42±2.54
	72	11.30±0.36^**	1.29±0.13^**	3.29±0.05^**	19.38±2.95^**	3.47±0.33^**	1.65±0.02^**	39.12±3.53^**

基于计算机辅助精子分析的大口黑鲈精子冷藏保存效果评估

Evaluation of sperm cryopreservation effectiveness for largemouth bass(Micropterus salmoides) based on computer-assisted sperm analysis

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