Application of Techniques for Sperm Selection in Fresh and Frozen‐Thawed Stallion Semen

Abstract

The objective of this research was to improve the techniques in processing chilled and frozen-thawed horse semen. In a preliminary experiment (Exp. I), different techniques for sperm selection and preparation [Swim-up, Glass wool (GW) filtration, Glass wool Sephadex (GWS) filtration; Percoll] were tested for their suitability for equine spermatozoa and results were compared with the routine procedure by dilution (Exp. I). In the main experiment (Exp. II), two sperm preparation techniques (GWS, Leucosorb) refering to the results of Exp. I and a previous study of our group (Pferdcheilkunde 1996 12, 773) were selected for processing complete ejaculates either for cooled-storage or cryopreservation. In a third experiment (Exp. III), pregnancy rates from inseminations with semen processed according to the techniques tested in Exp. II were compared with those obtained with semen processed according to routine procedures. In Exp. I (six stallions, six ejaculates/stallion), between 48 and 92% of spermatozoa were lost following the different sperm selection procedures (p < 0.05). Preparation of sperm increased percentage of progressively motile spermatozoa (pms) [Swim-up, GW, GWS vs dilution, Percoll (p < 0.05)] and decreased percentage of sperm head abnormalities [Swim-up, GW, GWS vs dilution, Percoll (p < 0.05)] probably by not improving the quality of individual cells, but by elimination of spermatozoa of inferior quality. In Exp. II (eight stallions, three ejaculates/stallion) Leucosorb and GWS procedures allowed the filtration of large volumes (extended ejaculates) for routine laboratory practice. GWS and Leucosorb filtration resulted in increased motility, membrane integrity and sperm viability after storage of spermatozoa until 48 h at +5 degrees C when compared with control (diluted) and centrifuged semen (p < 0.05). Significantly more spermatozoa were recovered after centrifugation (87.8 +/- 15.4%) compared with GWS (63.5 +/- 18.6%) and Leucosorb filtration (53.6 +/- 22.3%). GWS or Leucosorb procedure resulted in successful cryopreservation of stallion semen without centrifugation for removal of seminal plasma. The per cycle conception rate of inseminated mares using 200 x 106 pms transferred within 8 h after collection of semen was not affected by GWS filtration or Leucosorb separation when compared with centrifugation (n.s.; Exp. III). In conclusion, GWS and Leucosorb filtration results in the improvement of semen quality and should be considered as a method for stallion semen processing. Additional studies are needed for the evaluation of potentially higher fertilizing ability of stallion spermatozoa separated by techniques for sperm selection.

Antioxidant supplementation and purification of semen for improved artificial insemination in livestock species

“…The GWS filtration of stallion spermatozoa improved both sperm quality (129) and mare pregnancy rates after deep-horn insemination with low sperm numbers (130). Leukosorb filters also removed defective spermatozoa from stallion semen with an efficiency comparable to GWS (131). Compared to other microspin chromatography column separation methods, the use of glass wool yielded the highest sperm motility in stallions (132).…”

Stallion Sperm Selection: Past, Present, and Future Trends

“…Processing low-quality stallion semen with one layer of Percoll resulted in an improvement in sperm quality, although there was no improvement for "normal" ejaculates [18]. In contrast, sperm quality in "normal" stallion ejaculates was improved by single-layer centrifugation (SLC) using Androcoll-E, a new commercially available species-specific colloid for equine spermatozoa [19,20].…”

Biomimetics in Action: Practical Applications of Single Layer Centrifugation for equine breeding

“…The results of Sharma et al [3] showed that although it was possible to improve sperm quality in human ejaculates of low initial quality, no improvement in sperm quality was seen when "normal" human ejaculates were processed in this manner. Sieme et al [4] obtained similar results using one layer of Percoll ™ for stallion semen.…”