Influence of age and weight on seminal parameters of golden-headed lion tamarin (Leontopithecus chrysomelas) in ex situ conditions and potential use of seminal coagulum for molecular procedures

The golden-headed lion tamarin (Leontopithecus chrysomelas) is an endangered primate endemic to the Atlantic Forest. Conservation efforts for the species involve applying reproductive biotechniques to preserve genetic resources and ensure the management of populations in both ex situ and in situ conditions. This study aims to initiate investigations into seminal and molecular factors influencing the reproductive potential of sexually mature males. Semen was collected using the penile vibrostimulation technique, and seminal parameters were assessed in two groups: the 'Old' group (average age 11.6 years; n=6) and the 'Young' group (average age 4.8 years; n=6). ANOVA results indicated age-related influences on plasma membrane integrity (p=0.049), acrosomal integrity (p=0.009), and DAB IV (p=0.026) for both groups. Linear regression revealed significant correlations between seminal parameters and age (plasma membrane integrity (p=0.021), acrosomal integrity (p=0.05), and DAB III (p=0.024)), alongside animal weight (plasma membrane integrity (p=0.010), acrosomal integrity (p=0.009), DAB III (p=0.33), and DAB IV (p=0.066)). In an effort to advance reproductive techniques and sperm selection, a protocol utilizing a discontinuous Percoll gradient was employed. Despite its effectiveness in isolating gametes, there were no significant gains in the reevaluated parameters post-selection, necessitating adjustments in the methodology. While semen cryopreservation is common in wild species, challenges arise due to seminal coagulum in many neotropical primate ejaculates, hindering gamete use in reproductive procedures. Given the precious nature of and the considerable effort involved in collecting semen from these animals, it would be desirable to maximize the sample's utility. The liquid fraction could be applied in reproductive biotechniques, while the spermatozoa contained in the clot could be utilized as a non-invasive approach for molecular evaluation of these gametes. This study established a protocol for RNA extraction from sperm retained in the seminal coagulum, highlighting its genetic richness often discarded post-processing. In summary, our study emphasizes the importance of early cryopreservation of semen to safeguard the reproductive potential of L. chrysomelas. Additionally, we propose further exploration of RNA quantity in gametes as a non-invasive tool for inferring male fertility, given the pivotal role of sperm RNA transcripts in regulating the activatio