Enhancing sperm cryo-preservability, ultrastructure, and testicular histology in rams through oral royal jelly administration

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This investigation explored the effects of royal jelly (RJ) administration in rams on reproductive parameters, with a specific focus on semen quality, post-thaw characteristics, sperm function, and testicular health. Rahmani Rams (n=15) were divided into 3 groups: the control group received 1 mL of distilled water orally (RJ0), and two treatment groups received 250 mg (RJ250) or 500 mg (RJ500) of RJ three days per week, every other day per head for 6 months. Oral administration of RJ resulted in significant improvements in sperm mass motility, viability, and plasma membrane integrity (p < 0.001). Conversely, ejaculate volume, sperm concentration, and the percentage of morphological abnormalities showed no significant differences between groups (p > 0.05). The RJ administration resulted in a significant reduction in oxidative markers such as MDA, H 2 O 2 , and NO, and an increase in TAC and CAT compared to the RJ0 group (p <0.01) in raw and post-thawed semen. In cryopreserved semen, the progressive motility, viability, and membrane function were significantly higher in all RJ-treated rams (p <0.01). The majority of sperm parameters, including progressive motility, distance average path, distance curve linear, distance straight line, average path velocity, curve linear velocity, and straight line velocity, were superior in the RJ groups compared to the untreated ram group (p <0.05). The orally administered RJ affected both sperm membrane and acrosome status significantly (p <0.01). Specifically, RJ supplementation significantly increased the proportion of viable spermatozoa with intact acrosomes, while concurrently reducing the percentages of both viable acrosome-damaged and non-viable acrosome-intact cells across all treatment groups (p <0.05). Sperm viability was significantly higher in the RJ250 and RJ500 groups compared to the RJ0 treatment (p < 0.001). Furthermore, RJ administration significantly decreased the incidence of early apoptotic, late apoptotic, and necrotic spermatozoa relative to the RJ0 group (p < 0.05). Regarding the microbial profile, both coliform and total bacterial counts in post-thawed semen were significantly lower in RJ-treated rams than in the RJ0 group (p < 0.05). Oral administration of RJ significantly preserved testicular histoarchitecture, characterized by the maintenance of seminiferous tubule integrity, an increased population of spermatogonia, and the distinct presence of intraluminal spermatozoa. Furthermore, RJ treatment maintained the critical ultrastructural components of the spermatozoa, including the structural integrity of the plasma membrane, the mitochondrial sheath, and the nucleus. Collectively, these findings suggest that oral RJ supplementation enhances reproductive performance and post-thaw semen quality in rams, likely mediated by its potent antioxidant properties and its protective role during the cryopreservation process.

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