30 Does size matter? Effect of sperm morphometric characteristics on the motility and swimming ability in stallion sperm samples

Historically, horses exhibit sperm quality below that of other domestic species. Stallion sperm demonstrates significant variability in motility and kinematic patterns among individuals. In this study, we determined the morphometric characteristics (area, length, width and perimeter) of the sperm head on the motility and kinematic parameters of spermatozoa in fresh ejaculates from 18 Pura Raza Español stallions. Semen samples were collected during the 2022 breeding season using an artificial vagina. After collection, samples were centrifuged (600 × g, 10 min), equilibrated, for 5 min, in INRA extender, and diluted to a concentration of 25 million spz/mL. Samples were evaluated for motility and kinematic parameters using the MOT module of the SCA™ CASA system (Microptic). At least 500 sperm of each ejaculate were evaluated immediately after equilibration and again after 60 min of incubation at 37.5°C in 1-mL Nunclon™ dishes. At both analysis times, morphometric smears were prepared, air-dried, stained (using Diff Quick™), and analysed (24 h after staining) using the MORPH module of SCA, with at least 100 sperm heads per sample. Within stallion, data at each assessment time were averaged and analysed using a linear Pearson correlation test. Our findings indicated that area, perimeter, and length of sperm were positively correlated (r: ~0.26; P < 0.05) with total and progressive motility at the 5-min evaluation after ejaculation. At the 60-min evaluation, the correlation values increased (r: ~0.35; P < 0.01) for perimeter and length. Conversely, sperm width showed no significant correlation with motility. Similar results (r: ~0.2, P < 0.05) were observed between velocities (curvilinear velocity, straight-line velocity, and average path velocity) and perimeter and length, with a slightly lower correlation with area. Moreover, perimeter and length were positively correlated with sperm beating cross frequency (r: ~0.20; P < 0.05) at 5 min, but even more so after 60 min of incubation (r: ~0.34; P < 0.01). Our results suggest that stallion sperm with larger and longer heads exhibit increased motility and kinematic patterns, both at 5 min and 60 min postcollection. Further studies are required to determine the biological significance of this relationship, as well as its potential involvement in sperm freezing ability in the species.

We thank National Centre of Equine Breeding (CENRE, SPAIN) for providing the stallions and samples.