Retrospective analysis of sperm metrics in fresh ejaculates of collared peccaries presenting contrasting semen freezability

Abstract

Based on a retrospective study of sperm parameters in raw ejaculates of collared peccaries classified as good and poor freezers, this study aimed to identify whether these animals present different sperm characteristics that allow predicting their contrasting freezability. For this purpose, ejaculates were obtained from twenty-six animals, in which an aliquot of fresh semen was immediately evaluated, while another aliquot was diluted in Tris-yolk-glycerol diluent and cryopreserved. Fresh and frozen/thawed samples were evaluated for sperm motility kinetic parameters, membrane integrity, mitochondrial activity, membrane functionality, sperm morphology and sperm binding capacity. Based on post-thawing sperm metrics, the animals were classified as good and poor freezers. Following this classification, a retrospective study of fresh ejaculates was performed, in which a reclassification of the samples was performed according to the groups obtained after thawing. For the thawed groups, analysis of variance was performed using SAS software and the Tukey mean test to detect the significant differences between the groups. For fresh samples, the Mann Whitney test was used to verify the difference between the groups regarding the kinetic motility parameters (MT, fast, medium, slow and static cells), mitochondrial activity, pH and sperm concentration. For the other fresh parameters, the Unpaired t-test was used, using the GraphPad Prism® software version 9.3 for Windows (GraphPad Software Inc., San Diego, CA, USA). The highest (P < 0.05) mean values of post-thawing sperm motility were verified for good freezers (42.8 ± 2.8 %), to the detriment of bad freezers (23.8 ± 2.7 %). Good freezers also showed the highest (P < 0.05) mean values of progressive sperm motility (25.8 ± 2.5 %), lateral head amplitude (6.1 ± 0.1 μm) and proportion of fast sperm (31.1 ± 2.6 %), while poor freezers showed the highest proportion (P < 0.05) of static sperm (72.8 ± 3.4 %). Regarding raw semen, all ejaculates had a milky appearance and whitish coloration. There was no statistical difference between the groups regarding the kinetic motility patterns (P > 0.05). Poor freezers provided the lowest percentage (P < 0.05) of sperm with intact membranes (78.8 ± 1.7 %) and mitochondrial activity (79.3 ± 1.8 %), compared to good freezers (87.0 ± 1.9 %; 87.3 ± 1.9 %), with no differences regarding the other parameters. In conclusion, sperm plasma membrane integrity and mitochondrial activity in raw semen appear to be useful for predicting the freezability of semen from collared peccaries classified as good and poor freezers.