标准化CASA分析准确评价海胆卵泡旁精子活力

IF 3.9 1区农林科学 Q1 FISHERIES

Aquaculture Pub Date : 2025-09-16 DOI:10.1016/j.aquaculture.2025.743197

Sabrina Gacem , Estefanía Paredes , Sara Campos , Francisco Sevilla , Carles Soler , Jesús Yániz , Anthony Valverde , Miguel A. Silvestre

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引用次数: 0

摘要

尽管计算机辅助精子分析（CASA）系统被广泛应用于评估许多脊椎动物物种的精子活力，但其在海胆精子中的应用仍然相对较少，关于这种无脊椎动物模型的数据有限。本研究旨在规范CASA系统用于评估海胆精子活力的帧率、计数室类型、扩展器和稀释率。采集精液，用人工海水稀释或添加0.3%血清牛白蛋白（ASW- bsa）。将稀释后的精液装入4种腔室：drop displacement、Spermtrack®（SK）和R2D10 (R2)；毛细管，Kubus®（KU）；和SpermLide®（SL）。研究了从低到高的三种稀释度。使用商业CASA系统（AI Station v1.2；精子分析技术有限公司，Buñol，西班牙）评估精子活力，该系统采用人工智能进行精子识别。在ASW中稀释后，精子在液滴置换室中不动；然而，当精液在ASW-BSA中稀释时，总运动性（TMOT）和进动性（PMOT）在不同腔型之间没有显著差异，与填充腔型无关。当精液在ASW中稀释时，KU室的速度参数明显高于SL室。然而，在ASW-BSA介质中，这种差异消失了，在SL室中直线速度（VSL）甚至显著更高。随着稀释度的增加（即细胞浓度降低），精子速度增加，线性度降低。准确测量曲线速度（VCL）的最佳帧率为510 fps，而较低的帧率（< 360 fps）足以评估VSL和平均路径速度（VAP）。综上所述，使用CASA评估海胆精子活力的最佳条件是帧速率为510 fps， ASW-BSA稀释至浓度低于75 × 106个精子/mL，并使用毛细管填充室。

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Standardizing CASA analysis to accurately assess sea urchin Paracentrotus lividus sperm motility

Although computer-assisted sperm analysis (CASA) system is widely applied for evaluating sperm motility in numerous vertebrate species, its application to sea urchin sperm remains comparatively underexplored, with limited data available on this invertebrate model. This study aimed to standardize the frame rate, type of counting chambers, extender, and dilution rate to use CASA system to evaluate sea urchin sperm motility. Semen samples were collected and diluted in artificial sea water alone (ASW) or supplemented with 0.3 % serum bovine albumin (ASW-BSA). The diluted semen was filled in 4 chamber types: drop displacement, Spermtrack® (SK) and R2D10 (R2); and capillary, Kubus® (KU); and SpermLide® (SL). Three dilutions ranging from low to high were studied. Sperm motility was evaluated using a commercial CASA system (AI Station v1.2; Sperm Analysis Technologies S.L., Buñol, Spain) that employs artificial intelligence for sperm recognition. Spermatozoa were immotile in drop displacement chambers when diluted in ASW; however total motility (TMOT) and progressivity (PMOT) did not differ significantly across chamber types when semen was diluted in ASW-BSA, independently of filling chamber type. KU chamber showed significantly higher velocity parameters compared to SL chamber when semen was diluted in ASW. However, these differences disappeared when in ASW-BSA medium, and straight line velocity (VSL) was even significantly higher in SL chamber. As dilution increased (i.e., lower cell concentration), sperm velocity increased, and linearity decreased. The optimal frame rate for accurate measurement of curvilinear velocity (VCL) was 510 fps, whereas lower frame rates (< 360 fps) were sufficient for evaluating VSL and average path velocity (VAP). In conclusion, to optimally assess sea urchin sperm motility using CASA requires a frame rate of 510 fps, dilution in ASW-BSA to concentrations below 75 × 10⁶ sperm/mL and using capillary-filling chambers.

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来源期刊

Aquaculture 农林科学-海洋与淡水生物学

CiteScore

8.60

自引率

17.80%

发文量

1246

审稿时长

56 days

期刊介绍： Aquaculture is an international journal for the exploration, improvement and management of all freshwater and marine food resources. It publishes novel and innovative research of world-wide interest on farming of aquatic organisms, which includes finfish, mollusks, crustaceans and aquatic plants for human consumption. Research on ornamentals is not a focus of the Journal. Aquaculture only publishes papers with a clear relevance to improving aquaculture practices or a potential application.