beta-Nerve growth factor (beta-NGF) is a seminal plasma element, responsible for inducing ovulation in camelids. The main organ of beta-NGF production remains nondescript. The aims of this study were to (a) characterize gene expression and protein localization of beta-NGF and its main receptor tyrosine kinase receptor A (TrKA) in the llama male reproductive tract, and (b) determine whether the seminal beta-NGF interacts with ejaculated sperm by localizing beta-NGF and TrKA in epididymal, ejaculated, and acrosome-reacted (AR) sperms and, additionally, by identifying beta-NGF presence in sperm-adsorbed proteins (SAP). Both beta-NGF and TrkA transcripts are widely expressed along the male reproductive tract, with a higher expression level of beta-NGF at prostate (p < 0.05). beta-NGF immunolabeling was only positive for prostate, whereas TrKA label was present in epithelial and muscular cells of testis, prostate, bulbourethral glands, and epididymis. Using an immunofluorescent technique, beta-NGF was colocalized with TrKA in the middle piece of ejaculated and AR sperm. However, only TrKA was observed in epididymal sperm indicating that beta-NGF could have a seminal origin. This was also confirmed by the identification of four beta-NGF isoforms in SAP. This study extends the knowledge about the participation of beta-NGF/TrkA in llama reproduction, providing evidence that may have roles in the regulation of sperm physiology.