Is the sperm-binding test to the hen's egg perivitelline membrane reliable for the functional assessment of equine and donkey semen?

Abstract

The evaluation of male fertility is traditionally conducted in vivo, but this method is costly and influenced by various factors, such as females and management. The sperm-binding test to the hen's egg perivitelline membrane (HEPM) is an affordable in vitro alternative with promising results in various species, although its application in equids is still not well studied. Thus, this study aimed to: i) assess the feasibility of the sperm-binding test to HEPM at two insemination doses (IDs) for equine and donkey semen; ii) evaluate the influence of seminal plasma and semen cryopreservation on the in vitro sperm-binding capacity to HEPM from both species. In Experiment I, the sperm-binding capacity to HEPM of equid semen with sperm viabilities (SVs) of 0%, 25%, 50%, 75%, and 100% was compared at insemination doses of 10 million (ID10) and 50 million (ID50) total sperm per apparatus containing the membrane. In Experiment II, the sperm-binding capacity to HEPM of fresh and frozen-thawed semen, with or without seminal plasma, was compared. All mean comparisons were evaluated using the PROC GLIMMIX package, considering a statistical difference when P<0.05. The sperm-binding capacity to HEPM was influenced by the insemination dose and the species, being higher for donkeys and for the ID50 dose. When the species was disregarded, the test allowed differentiation of binding potential between infertile seminal samples (0% SV) and samples that exhibited fertile potential (≥25% SV). However, it did not allow differentiation of binding potential between samples with lower and higher SVs (25%, 50%, and 75%). Additionally, cryopreservation and seminal plasma did not influence the sperm-binding capacity to HEPM. In conclusion, this study demonstrated that the sperm-binding test to HEPM can be used to assess the in vitro potential of equid sperm binding capacity. Further studies regarding insemination dose, incubation time of the inseminated HEPM, and the number of repetitions per seminal sample are required to achieve greater accuracy of the test.