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Animals : an open access journal from MDPI2023; 13(3); doi: 10.3390/ani13030382

Optimization of the Equine-Sperm Freeze Test in Purebred Spanish Horses by Incorporating Colloidal Centrifugation.

Abstract: The Purebred Spanish Horse, according to our clinical experience, is characterized by having a high number of stallions that do not meet the international commercial recommendations for equine-sperm cryopreservation. This means that artificial insemination with frozen semen from these stallions is less widespread than in other breeds. In this study, we investigated if the incorporation of single-layer colloidal centrifugation prior to cryopreservation in clinical conditions could increase the number of ejaculates of Purebred Spanish stallions suitable for this processing, observing the influence of centrifugation and freezing extender protocol on post-thawed sperm motility. Using colloidal centrifugation, the percentage of ejaculates available to be frozen was increased from 35% (6/17) to 71% (12/17), doubling the number of samples that could have been subjected to cryopreservation. We only found significant differences in linearity (LIN) and lateral head displacement (ALH) after 5 min of incubation at 37 °C between colloidal and simple centrifugation processing techniques. No significant differences were found between the two different colloidal protocols in any of the variables considered. Colloidal centrifugation allowed us to obtain, from worse fresh-quality ejaculates, thawed sperm doses with similar quality to that of good-quality ejaculates. BotuCrio® produced, in general, higher motility parameters and its characteristics than the other extenders analyzed, with significant differences found in comparison to Inra-Freeze® and Lac-Edta in both total (MOT) and progressive motility (PMOT) when using colloidal centrifugation and only in PMOT when applying simple centrifugation. Colloidal centrifugation optimized the efficiency of cryopreservation, as it allowed us to increase the number of ejaculates of Purebred Spanish Horses suitable to be frozen. Including these semen processing techniques in the freeze test could help to optimize equine-sperm cryopreservation protocols, especially when dealing with individuals or breeds for which initially low sperm quality prevents or limits their inclusion in sperm cryopreservation programs.
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Publication Date: 2023-01-22 PubMed ID: 36766271PubMed Central: PMC9913238DOI: 10.3390/ani13030382Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This study has found that using colloidal centrifugation significantly improved the number of semen samples from Purebred Spanish Horses suitable for freezing and artificial insemination, addressing an issue with the breed’s low sperm quality affecting its suitability for cryopreservation.

The Problem with Purebred Spanish Horse’s Sperm Cryopreservation

  • Many Purebred Spanish Horses’ sperm samples do not meet international standards for cryopreservation (freezing). This makes frozen artificial insemination less common for this breed.
  • Without intervention, only about 35% (6 out of 17) of the samples, in the study were suitable for freezing.

Methodology of the Study

  • The researchers’ approach for increasing the number of suitable samples involved incorporating single-layer colloidal centrifugation before the freezing.
  • They investigated the effect this method has on the sperm’s motility after thawing.
  • They used two different protocols of centrifugation to see if there were significant differences in outcomes.
  • They studied the impact of different freezing extenders on the sperm’s quality and motility.

Findings and Implications

  • Applying single-layer colloidal centrifugation prior to freezing increased the percentage of suitable samples for cryopreservation to 71% (12 out of 17).
  • Significant differences were only found in linearity and lateral head displacement between colloidal and simple centrifugation methods after incubation at 37°C (degrees Celsius) for 5 minutes.
  • There were no significant variations in the outcomes using the two different colloidal centrifugation protocols.
  • The same quality of sperm after thawing could be obtained from poorer quality fresh ejaculates as from high-quality ones, thus effectively uplifting the quality of poor samples.
  • Among the different types of freezing extenders used, BotuCrio was found to enhance sperm motility parameters better than others.

The study has important implications for improving fertility in Purebred Spanish horses. The researchers concluded that the single-layer colloidal centrifugation optimized the efficiency of cryopreservation because it increased the number of suitable samples. This could help in optimizing equine sperm cryopreservation protocols particularly for individuals or breeds with initially low sperm quality.

Cite This Article

APA
Gutiérrez-Cepeda L, Crespo F, Blazquez JC, Serres C. (2023). Optimization of the Equine-Sperm Freeze Test in Purebred Spanish Horses by Incorporating Colloidal Centrifugation. Animals (Basel), 13(3). https://doi.org/10.3390/ani13030382

Publication

Animals : an open access journal from MDPI
ISSN: 2076-2615
NlmUniqueID: 101635614
Country: Switzerland
Language: English
Volume: 13
Issue: 3

Researcher Affiliations

Gutiérrez-Cepeda, Luna
  • Animal Medicine and Surgery Department, Veterinary Faculty, UCM, Avda. Puerta de Hierro s/n, 28040 Madrid, Spain.
Crespo, Francisco
  • Centro Militar de Cría Caballar (CCFAA), C/Arsenio Gutiérrez Palacios s/n, 05005 Ávila, Spain.
Blazquez, Juan Carlos
  • Centro Militar de Cría Caballar (CCFAA), C/Arsenio Gutiérrez Palacios s/n, 05005 Ávila, Spain.
Serres, Consuelo
  • Animal Medicine and Surgery Department, Veterinary Faculty, UCM, Avda. Puerta de Hierro s/n, 28040 Madrid, Spain.

Grant Funding

  • FPU fellowship AP2008-02034 / FPU fellowship AP2008-02034

Conflict of Interest Statement

The authors declare no conflict of interest.

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Citations

This article has been cited 3 times.
  1. Al-Kass Z, Morrell JM, Ntallaris T. Effect of Centrifugation of Stallion Semen Through a Low Density Colloid Prior to Freezing on Sperm Cryosurvival. Animals (Basel) 2025 Jun 25;15(13).
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  2. Zabala SM, Serres C, Montero N, Crespo F, Lorenzo PL, Pérez-Aguilera V, Oliet A, Hijón V, Moreno S, González-Zorn B, Gutiérrez-Cepeda L. Innovative Approaches to Avoid Antibiotic Use in Equine Semen Cryopreservation: Advancing Sustainable Reproductive Technologies. Animals (Basel) 2025 May 9;15(10).
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  3. Boni R, Ruggiero R, Di Palma T, Ferrara MA, Preziosi G, Cecchini Gualandi S. Stallion Sperm Freezing with Different Extenders: Role of Antioxidant Activity and Nitric Oxide Production. Animals (Basel) 2024 Aug 25;14(17).
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