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

Abstract

Single Layer Centrifugation (SLC) of spermatozoa through a species-specific colloid has been shown to be effective in selecting the best spermatozoa from stallion semen. The method is easier to use and less time-consuming than density gradient centrifugation (DGC), and has been scaled-up to allow whole ejaculates to be processed in a practical manner. The major applications for SLC-sperm selection are as follows: to improve sperm quality in artificial insemination (AI) doses, particularly for "problem" stallions; to increase the "shelf-life" of normal sperm samples, either by processing the fresh semen before preparing AI doses, or by processing the stored semen dose to extract the best spermatozoa; to circumvent the problem of spermatozoa that do not tolerate cooling to 4-6°C; to improve cryosurvival by removing dead and dying spermatozoa prior to cryopreservation, or selecting the live spermatozoa post-thawing; to select morphologically normal spermatozoa with intact chromatin from sub-fertile stallion semen for ICSI, thus increasing the number of blastocysts; to remove pathogens (viruses, bacteria); to accelerate the process of flow cytometric sex selection by removing the dead and dying sperm before passage through the laser beam; to conserve rare breeds. These applications are discussed and practical examples provided.

Quality of bull spermatozoa after preparation by single-layer centrifugation

“…Pregnancy rates are also higher when SLC-selected samples prepared from "normal" ejaculates are used for AI compared with the corresponding untreated controls (fresh and cooled sperm doses). Furthermore, SLC-selected stallion and boar spermatozoa survive freezing better and survive longer after thawing than unselected controls (Morrell, 2011;Martinez-Alborcia et al, 2013). Thus, our previous studies in other species indicate that SLC improves both sperm quality and sperm survival, and that pregnancy rates are higher after AI.…”

Stallion Sperm Selection: Past, Present, and Future Trends

“…Because potential applications of SLC in equine breeding have been reviewed recently [34], only a brief update will be provided here.…”

“…When SLC-selected spermatozoa from "problem" ejaculates were inseminated, pregnancy rates were increased to approximately 65%, depending on the nature of the problem [36]. Pregnancies were achieved in four of four mares when spermatozoa with low tolerance to cooling were processed by SLC, cooled, and inseminated after 24 hours [34]. 2.…”

Colloids: Applications in Sperm Preparation for Assisted Reproduction

“…Removing all the seminal plasma from the spermatozoa and selecting the robust spermatozoa by SLC allows them to survive being cooled, and their fertilizing capability is retained [27]. Alternatively, SLC-selected sperm samples can be maintained at around 15° for at least 48 h and still retain their motility [30] and fertilizing capacity (J. Newcombe, personal communication)…”

“…Similarly to equine breeding, the pig breeding industry relies heavily on the insemination of fresh, cooled semen. Although cryopreservation protocols exist for boar semen, there is a perception that pregnancy rates are lower and litter sizes smaller if cryopreserved semen is used for AI instead of fresh semen [30].…”