In vivo semen characterization and seasonal variation in Procavia capensis.

Abstract: Seasonal reproduction is common among wild mammals, but male fertility traits are often understudied. The rock hyrax (Procavia capensis) is a seasonal breeder with a narrow reproductive window, yet its semen characteristics and seasonal variation remain poorly understood. Our objectives were to develop and validate an electroejaculation protocol for semen collection, and to enable, for the first time, in vivo assessments of sperm morphology, ultrastructure, morphometry, and seasonal variation in semen quality in both captive and wild populations. A total of 70 semen collection attempts were conducted: 17 in captive males at approximately monthly intervals over 1 year and 53 in wild males just before peak mating season and again 2-4 weeks later, across 3 consecutive years (2021-2023). Electroejaculation was well tolerated and effective, particularly around the mating season, eliciting consistent responses and yielding sperm-containing ejaculates in 88.7% of wild procedures. Sperm morphometry revealed a mean total length of ∼56 µm, with ultrastructural features resembling other eutherian mammals. Seasonal analysis demonstrated significantly higher sperm concentration, motility, and normal morphology during peak mating season compared to later samples. Post-peak samples showed increased structural abnormalities, including midpiece and principal piece defects, and signs of disrupted spermatogenesis and epididymal maturation. Cytology and high-resolution scanning electron microscopy confirmed these findings, revealing compromised sperm integrity and elevated round cell counts outside the breeding peak. This study establishes the first in vivo semen collection protocol and comprehensive semen evaluation in the rock hyrax, revealing seasonal variation in male fertility and enabling repeatable, non-lethal reproductive monitoring. Lay summary: Male fertility traits are poorly characterized in most wild mammals. The rock hyrax (also known as rock rabbit) is a seasonal breeder with a short mating period each year, but little was known about its male reproductive physiology because semen studies relied on post-mortem material. We established a non-lethal protocol for semen collection in rock hyraxes and applied it both in captivity and under natural desert conditions. This enabled detailed analyses of sperm's shape (morphology), architecture (ultrastructure), and dimensions (morphometry), and seasonal semen changes. We found that sperm concentration, motility, and normal morphology were highest during peak breeding season but declined sharply only a few weeks later, accompanied by structural abnormalities and signs of disrupted spermatogenesis. Our study reveals sharp seasonal shifts in male fertility and demonstrates a safe, repeatable method for studying reproduction in wild mammals, with potential applications in conservation and comparative studies among seasonal species.