Single-Layer Colloid Centrifugation as a Method to Process Urine-Contaminated Stallion Semen After Freezing-Thawing

• Published in: Journal of equine veterinary science Vol. 87; p. 102910
• Main Authors: Podico, Giorgia, Ellerbrock, Robyn E., Curcio, Bruna R., Cheong, Soon Hon, Lima, Fabio S., Canisso, Igor F.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-04-2020
• Subjects: Animals; Centrifugation; Centrifugation - veterinary; Colloids; Cryopreservation; Freezing; Horse; Horses; Male; Semen; Sperm Motility; Urospermia; Cryopreservation; Centrifugation; Horse; Urospermia
• ISSN: 0737-0806; 1542-7412
• DOI: 10.1016/j.jevs.2020.102910

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.
•Urine contamination affects post-thawed semen parameters in a dose-dependent manner.•Single-layer colloid centrifugation enhances morphology, motilities, and viability in frozen-thawed semen with low urine contamination.•Recovery rate after single-layer colloid centrifugation does not differ in frozen-thawed semen with a low to a high level of urine contamination.