FREE SHIPPING over $40
OVER 10000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Theriogenology2018; 114; 212-220; doi: 10.1016/j.theriogenology.2018.03.028

How does the microbial load affect the quality of equine cool-stored semen?

Abstract: Contaminating bacteria present in stallion ejaculates may compromise sperm quality during storage. Different procedures have been used to reduce the load of microorganisms in semen and avoid bacterial growth during storage. The aims of this study were: 1) to evaluate different techniques to eliminate bacteria in semen 2) to study the relationship between total microflora load (TML) and ROS production; and 3) to determine if TML affects the functionality of cool-stored sperm. Ejaculates from 11 stallions were split and processed in 3 ways: A. extended semen; B. conventional centrifuged semen, and C. Single layer centrifugation through Androcoll-E (SLC). All samples were preserved in INRA 96 at 5 °C for 72 h. Aliquots from native semen and from different treatments were taken for bacteriological analysis at T0, T24, T48 and T72h of storage and Total microbial load (TML: CFU (colony-forming units/ml) was calculated. The ROS production (dichlorodihydrofluorescein diacetate for H2O2, dihydroethidium for superoxide anion and CellROX deep red for total ROS), viability (YO-PRO-1-Ethidium) and lipid peroxidation (BODIPY-C11) were assessed by flow cytometry, and motility by CASA. The bacteria isolated were Corynebacterium spp, Arcanobacterium spp, Bacillus spp, Dermobacter, Staphylococcus spp, Streptococcus spp, Penicilium spp. TML of semen showed correlations with live sperm (r: -0.771), dead sperm (r: 0.580), H2O2 production (r: 0.740), and total ROS production (CellROX (+)) (r: -0.607), Total motility (r: 0.587), Progressive motility (r: -0.566), VCL (r: -0.664), VSL (r: -0,569), VAP (r: -0.534) (p ≤ 0.05). SLC removed 99.34% of the microbial load, which was assicated with a significanlty reduced H2O2 production (p ≤ 0.05). However, only samples treated with Androcoll-E had a higher total ROS production (CellROX +) (p ≤ 0.05). These results suggest that CellROX stain probably identifies superoxide production rather than H2O2 and this higher superoxide production may reflect an intense sperm functionality. The bacterial load increased the production of H2O2 in cool-stored semen which was associated with lower tolerance to refrigeration. SLC was the sperm processing technique that was most efficient at removing bacteria, reducing H2O2 production and selecting the most functional sperm.
Copyright © 2018 Elsevier Inc. All rights reserved.
Get Full Text
Publication Date: 2018-03-26 PubMed ID: 29653389DOI: 10.1016/j.theriogenology.2018.03.028Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article

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 research article investigates how the presence of bacteria in horse semen affects its quality during storage, and the effectiveness of various techniques to reduce bacterial contamination. The study concludes that a specific process named Single Layer Centrifugation (SLC) is most effective to manage bacteria, maintain sperm quality and enhance overall sperm functionality.

Objective and Methodology

  • This research sought to understand the impact of microorganisms on the functionality and quality of cooled-stored horse semen, with the specific goals of evaluating techniques to reduce bacterial presence, examining the connection between bacteriological load and reactive oxygen species (ROS) production, and ascertaining how the bacteria load impacts the semen’s functionality.
  • The semen samples were obtained from 11 stallions, divided, and processed in three different ways – via standard extension, standard centrifugation, and Single Layer Centrifugation through a specialized medium called ‘Androcoll-E’.
  • The bacteriological load in all samples was measured at different time intervals (immediately, and after 24, 48, and 72 hours of storage). Other parameters assessed included ROS production, sperm viability and motility, and lipid peroxidation.

Findings and Conclusions

  • A variety of bacteria were identified in the semen samples, including strains of Corynebacterium, Arcanobacterium, Bacillus, Dermobacter, Staphylococcus, Streptococcus, and Penicilium.
  • The bacteria load was found to affect nearly all measured parameters, including sperm survival, ROS production, and motility, suggesting a fairly robust impact on the semen’s overall quality and functionality in storage.
  • The SLC process was shown to be the most effective technique, removing over 99% of the bacterial load. Moreover, this process resulted in less production of hydroxyl radicals (HO) and an increased total ROS production, indicating higher sperm functionality.
  • The higher ROS production (specifically, superoxide production) demonstrated by samples treated with the Androcoll-E medium suggests that this procedure may be more effective in reducing contamination and maintaining semen viability in cool storage. This could potentially improve the results of artificial insemination protocols.

Cite This Article

APA
Varela E, Rey J, Plaza E, Muñoz de Propios P, Ortiz-Rodríguez JM, Álvarez M, Anel-López L, Anel L, De Paz P, Gil MC, Morrell JM, Ortega-Ferrusola C. (2018). How does the microbial load affect the quality of equine cool-stored semen? Theriogenology, 114, 212-220. https://doi.org/10.1016/j.theriogenology.2018.03.028

Publication

Theriogenology
ISSN: 1879-3231
NlmUniqueID: 0421510
Country: United States
Language: English
Volume: 114
Pages: 212-220

Researcher Affiliations

Varela, E
  • Unit of Infection Diseases, University of Extremadura, Caceres, Spain.
Rey, J
  • Unit of Infection Diseases, University of Extremadura, Caceres, Spain.
Plaza, E
  • Laboratory of Equine Reproduction and Equine Spermatology, University of Extremadura, Caceres, Spain.
Muñoz de Propios, P
  • Animal Reproduction and Obstetrics, University of León, León, Spain.
Ortiz-Rodríguez, J M
  • Laboratory of Equine Reproduction and Equine Spermatology, University of Extremadura, Caceres, Spain.
Álvarez, M
  • Animal Reproduction and Obstetrics, University of León, León, Spain.
Anel-López, L
  • Animal Reproduction and Obstetrics, University of León, León, Spain.
Anel, L
  • Animal Reproduction and Obstetrics, University of León, León, Spain.
De Paz, P
  • Animal Reproduction and Obstetrics, University of León, León, Spain.
Gil, M C
  • Laboratory of Equine Reproduction and Equine Spermatology, University of Extremadura, Caceres, Spain.
Morrell, J M
  • Faculty of Veterinary Medicine and Animal Sciences, SLU, Uppsala, Sweden.
Ortega-Ferrusola, C
  • Animal Reproduction and Obstetrics, University of León, León, Spain. Electronic address: cortf@unileon.es.

MeSH Terms

  • Animals
  • Bacteria / classification
  • Bacteria / isolation & purification
  • Cold Temperature
  • Male
  • Semen / microbiology
  • Semen Preservation / veterinary

Citations

This article has been cited 15 times.
  1. Boni R, Ruggiero R, De Luca F, Preziosi G, Ferrara MA, Ostuni A, Guerriero S, Gallo A, Murano C, Cecchini Gualandi S. Enhancing Stallion Semen Cryopreservation: Selected Antioxidant Extracts and Sperm Freezability. Antioxidants (Basel) 2025 Nov 16;14(11).
    doi: 10.3390/antiox14111363pubmed: 41300520google scholar: lookup
  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).
    doi: 10.3390/ani15101368pubmed: 40427246google scholar: lookup
  3. González N, Peñalosa A, de Blas I, Gil L. Sustainable Alternatives to the Reduction of Plastic Straws Used with Chilled Equine Semen. Animals (Basel) 2024 Nov 25;14(23).
    doi: 10.3390/ani14233388pubmed: 39682354google scholar: lookup
  4. Malaluang P, Niazi A, Guo Y, Nagel C, Guimaraes T, Rocha A, Aurich C, Morrell JM. Bacterial diversity in semen from stallions in three European countries evaluated by 16S sequencing. Vet Res Commun 2024 Jun;48(3):1409-1421.
    doi: 10.1007/s11259-024-10321-3pubmed: 38305959google scholar: lookup
  5. Zabala SM, Serres C, Montero N, Crespo F, Lorenzo PL, Pérez-Aguilera V, Galán C, Domínguez-Gimbernat M, Oliet A, Moreno S, González-Zorn B, Gutiérrez-Cepeda L. Strategies to Reduce the Use of Antibiotics in Fresh and Chilled Equine Semen. Animals (Basel) 2024 Jan 5;14(2).
    doi: 10.3390/ani14020179pubmed: 38254348google scholar: lookup
  6. Egyptien S, Deleuze S, Ledeck J, Ponthier J. Sperm Quality Assessment in Stallions: How to Choose Relevant Assays to Answer Clinical Questions. Animals (Basel) 2023 Oct 6;13(19).
    doi: 10.3390/ani13193123pubmed: 37835729google scholar: lookup
  7. Domain G, Ali Hassan H, Wydooghe E, Bogado Pascottini O, Johannisson A, Morrell JM, Niżański W, Van Soom A. Influence of Single Layer Centrifugation with Canicoll on Semen Freezability in Dogs. Animals (Basel) 2022 Mar 11;12(6).
    doi: 10.3390/ani12060714pubmed: 35327111google scholar: lookup
  8. Quiñones-Pérez C, Martínez A, Ortiz I, Crespo F, Vega-Pla JL. The Semen Microbiome and Semen Parameters in Healthy Stallions. Animals (Basel) 2022 Feb 22;12(5).
    doi: 10.3390/ani12050534pubmed: 35268102google scholar: lookup
  9. Struyfs C, Breukers J, Spasic D, Lammertyn J, Cammue BPA, Thevissen K. Multiplex Analysis to Unravel the Mode of Antifungal Activity of the Plant Defensin HsAFP1 in Single Yeast Cells. Int J Mol Sci 2022 Jan 28;23(3).
    doi: 10.3390/ijms23031515pubmed: 35163438google scholar: lookup
  10. Cojkic A, Niazi A, Guo Y, Hallap T, Padrik P, Morrell JM. Identification of Bull Semen Microbiome by 16S Sequencing and Possible Relationships with Fertility. Microorganisms 2021 Nov 25;9(12).
    doi: 10.3390/microorganisms9122431pubmed: 34946031google scholar: lookup
  11. Quiñones-Pérez C, Hidalgo M, Ortiz I, Crespo F, Vega-Pla JL. Characterization of the seminal bacterial microbiome of healthy, fertile stallions using next-generation sequencing. Anim Reprod 2021;18(2):e20200052.
    doi: 10.1590/1984-3143-AR2020-0052pubmed: 34394753google scholar: lookup
  12. Eini F, Kutenaei MA, Zareei F, Dastjerdi ZS, Shirzeyli MH, Salehi E. Effect of bacterial infection on sperm quality and DNA fragmentation in subfertile men with Leukocytospermia. BMC Mol Cell Biol 2021 Aug 13;22(1):42.
    doi: 10.1186/s12860-021-00380-8pubmed: 34388964google scholar: lookup
  13. Papas M, Catalán J, Recuero S, Morrell JM, Yeste M, Miró J. Single Layer Centrifugation Improves the Quality of Fresh Donkey Semen and Modifies the Sperm Ability to Interact with Polymorphonuclear Neutrophils. Animals (Basel) 2020 Nov 16;10(11).
    doi: 10.3390/ani10112128pubmed: 33207812google scholar: lookup
  14. Kelesidis T, Tran E, Arastoo S, Lakhani K, Heymans R, Gornbein J, Middlekauff HR. Elevated Cellular Oxidative Stress in Circulating Immune Cells in Otherwise Healthy Young People Who Use Electronic Cigarettes in a Cross-Sectional Single-Center Study: Implications for Future Cardiovascular Risk. J Am Heart Assoc 2020 Sep 15;9(18):e016983.
    doi: 10.1161/JAHA.120.016983pubmed: 32896211google scholar: lookup
  15. Quiñones-Pérez C, Martínez A, Crespo F, Vega-Pla JL. Comparative Semen Microbiota Composition of a Stallion in a Taylorella equigenitalis Carrier and Non-Carrier State. Animals (Basel) 2020 May 17;10(5).
    doi: 10.3390/ani10050868pubmed: 32429567google scholar: lookup
Subscribe to our newsletter
Join 99,911 horse owners like you who receive our email updates on horse health and nutrition.