Abstract
Egg yolk is widely used as a cryoprotectant in dog semen extenders, but there is a risk of contamination with animal pathogens. In addition, egg yolk may vary in composition, making it difficult to standardize the extender. Lecithin is an animal protein‐free alternative to egg yolk for semen cryopreservation. Recently, it was shown that 1% of soybean lecithin type II‐S was better than 2% for freezing canine semen. The aim of the study was to compare two different types of soybean lecithin, with egg yolk as a control. Ejaculates from eight dogs were divided into three equal parts and diluted with a Tris‐based extender, containing either 20% egg yolk, 1% soybean lecithin Type II‐S or 1% soybean lecithin Type IV‐S. The samples were then frozen. Sperm motility was evaluated by computer‐assisted sperm analysis (CASA), acrosome integrity (FITC‐PNA/PI) and sperm membrane integrity (SYBR‐14/PI) post‐thaw, as well as after 2 and 4 hr incubation at 37°C. Post‐thaw sperm chromatin structure assay and plasma membrane integrity were evaluated by flow cytometry. Total motility, sperm plasma membrane integrity and acrosome integrity were significantly better in the egg yolk extender than in the two soybean lecithin‐based extenders. Individual motility post‐thaw differed more than in the fresh samples, illustrating individual differences in tolerance to the cryostress. The DNA Fragmentation Index (% DFI) was significantly lower in the Tris egg yolk (TEY) extender compared to any of the soybean‐based extenders. The number of high green stained spermatozoa were significantly higher in Type IV‐S compared to the control TEY extender. In conclusion, egg yolk was superior to the two lecithin‐based extenders to cryopreserve canine semen.
Highlights
The ability to cryopreserve semen has made it possible to exchange genetic material over long distances, and enable long time storage.Egg yolk is widely used as a cryoprotectant in dog semen extenders but there are some concerns and risks with the use of egg yolk including a risk of bacterial contamination and a potential risk of causing disease (Bousseau et al, 1998)
Both of these studies report that post-thaw sperm quality was similar to freezing in a Tris egg yolk (TEY) extender
Sperm plasma membrane integrity was significantly better in the TEY than in either of the two soybean lecithin-based extenders at any given time (p < 0.001, Figure 3, Table 1)
Summary
The ability to cryopreserve semen has made it possible to exchange genetic material over long distances, and enable long time storage. Cryopreservation of dog semen has been performed with 0.04% or 1.5% soybean lecithin (Beccaglia et al, 2009; Hidalgo et al, 2014), respectively Both of these studies report that post-thaw sperm quality was similar to freezing in a Tris egg yolk (TEY) extender. In a study by Axnér and Lagerson (2016), it was concluded that an extender with 1% soybean lecithin extender resulted in better post-thaw sperm motility than 2% soybean lecithin for freezing dog semen. The aim of this study was to compare two tris-based extenders containing 1% soybean lecithin from two different sources: type II-S (Sigma P5638) which was used by Axnér and Lagerson (2016), and type IV-S (Sigma P3644) that had previously yielded similar results as an egg yolk extender for the cryopreservation of goat (Salmani et al, 2014) and rabbit spermatozoa (Nishijima et al, 2015). A TEY extender was used as a control for the cryopreservation of dog semen
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