Single-Layer Colloid Centrifugation as a Method to Process Urine-Contaminated Stallion Semen After Freezing-Thawing.
Abstract: Urospermia is a major ejaculatory dysfunction affecting stallions. It has been thought that urine-contaminated semen should not be cryopreserved; however, on select cases, urine contamination of semen cannot be avoided. A recent study suggested that urospermic semen can be cryopreserved after cushion centrifugation and extension. Thus, this study aimed to assess the use of single-layer colloid centrifugation (SLC) to process frozen-thawed urine-contaminated stallion semen. Raw ejaculates (n = 55) from eight stallions were split into three groups: no urine, low (20%), or high (50%) urine contamination. Semen was extended 1:1, cushion-centrifuged, and resuspended at 200 million sperm/mL in BotuCrio. Resuspended semen was loaded in 0.5 mL straws and cryopreserved in liquid nitrogen. Samples were thawed (37°C for 30 seconds) and processed by SLC (400 g/30 minutes). Percentages of total motility (TM) and progressive motility (PM) were assessed with computer-assisted semen analyzer. Sperm viability (%VIAB) and yield were assessed with a NucleoCounter before and after gradient centrifugation. Data were analyzed with two-way ANOVA and Tukey's test. The motility parameters TM before SLC (control: 35 ± 2; low: 33 ± 0.7; high: 22 ± 1.8) after SLC (control: 51 ± 3.6; low: 42 ± 2.2; high: 25 ± 2.8) and PM before SLC (control: 24 ± 1.8; low: 21 ± 1.14; high: 12 ± 1.5) and after SLC (control: 40.3 ± 3.2; low: 31 ± 3.9; high: 14 ± 2) significantly decreased with increasing urine contamination. Urine contamination marginally reduced (P < .05) sperm viability after cryopreservation before SLC (control: 45 ± 0.7; low: 27 ± 0.2; high: 27 ± 0.3) and after SLC (control: 54 ± 0.5; low: 49 ± 0.7; high: 38 ± 0.6). Recovery rates of sperm after centrifugation were not significantly different between groups. In conclusion, urine contamination affects sperm motility parameters in a dose-dependent manner. Post-thaw SLC selected sperm with higher motility and viability in control and low groups but only selected sperm with higher viability in the high group.
Copyright © 2020 Elsevier Inc. All rights reserved.
Publication Date: 2020-01-07 PubMed ID: 32172909DOI: 10.1016/j.jevs.2020.102910Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The research article investigates the viability of using a single-layer colloid centrifugation method to process frozen and then thawed horse semen that has been contaminated with urine. The findings reveal that although urine contamination significantly reduces semen motility, the centrifugation method could successfully salvage semen with improved motility and viability in samples with low contamination.
Objective and Methodology
- The objective of this study is to examine the effectiveness of a single-layer colloid centrifugation (SLC) process to process frozen-thawed stallion semen contaminated with urine.
- Urospermia, a condition in which urine mixes with semen during ejaculation, is a significant problem affecting stallions.
- Previously, it was believed that contaminated semen could not be cryopreserved. However, this study seeks to challenge this belief.
- Semen samples (n = 55) from eight stallions were divided into three groups based on the level of urine contamination: no urine, low (20%) and high (50%).
- The semen was then extended, cushion-centrifuged and subsequently suspended in BotuCrio, which is a cryoprotective medium, at a concentration of 200 million sperm/mL.
- The prepared samples were then cryopreserved in liquid nitrogen, thawed, and finally processed through single-layer colloid centrifugation.
Analysis and Findings
- Researchers assessed total motility (TM), progressive motility (PM), sperm viability (%VIAB) and yield before and after the centrifugation process using a computer-assisted semen analysis tool and NucleoCounter.
- Data was analysed using two-way ANOVA and Tukey’s test to measure statistical significance.
- Findings showed that total and progressive motility decreased as the level of urine contamination increased.
- A slight reduction in sperm viability post-cryopreservation was observed with increased urine contamination.
- Recovery rates of sperm after centrifugation were not significantly different between the groups.
Conclusion
- In conclusion, the study found that urine contamination affects sperm motility in a dose-dependent manner. The more urine present, the lower the motility.
- The use of post-thaw SLC could select sperm with higher motility and viability in control and low groups, but it only selected sperm with a higher viability in the high group.
- Overall, the results suggest that using single-layer colloid centrifugation could be an effective method to process stallion semen contaminated with urine, especially in low contamination cases.
Cite This Article
APA
Podico G, Ellerbrock RE, Curcio BR, Cheong SH, Lima FS, Canisso IF.
(2020).
Single-Layer Colloid Centrifugation as a Method to Process Urine-Contaminated Stallion Semen After Freezing-Thawing.
J Equine Vet Sci, 87, 102910.
https://doi.org/10.1016/j.jevs.2020.102910 Publication
Researcher Affiliations
- Department of Veterinary Clinical Medicine, College Veterinary Medicine, University of Illinois Urbana-Champaign, Urbana, IL.
- Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA.
- Department of Veterinary Clinical Medicine, College Veterinary Medicine, University of Illinois Urbana-Champaign, Urbana, IL.
- Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY.
- Department of Veterinary Clinical Medicine, College Veterinary Medicine, University of Illinois Urbana-Champaign, Urbana, IL.
- Department of Veterinary Clinical Medicine, College Veterinary Medicine, University of Illinois Urbana-Champaign, Urbana, IL. Electronic address: canisso@illinois.edu.
MeSH Terms
- Animals
- Centrifugation / veterinary
- Colloids
- Freezing
- Horses
- Male
- Semen
- Sperm Motility
Citations
This article has been cited 3 times.- Podico G, Spencer KM, Magalhaes HB, Canisso IF. Semen Quality of the First and Second Ejaculates Collected from Breeding Inactive Stallions after Cooling and Freezing. Vet Sci 2023 Feb 21;10(3).
- Gutiérrez-Cepeda L, Crespo F, Blazquez JC, Serres C. Optimization of the Equine-Sperm Freeze Test in Purebred Spanish Horses by Incorporating Colloidal Centrifugation. Animals (Basel) 2023 Jan 22;13(3).
- Alves NC, Diniz SA, Viegas RN, Arigoni AL, Freitas MM, Lana ÂQ, Lagares MA. Action of swim-up and caffeine on equine frozen sperm. Anim Reprod 2022;19(4):e20220056.
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