Cryoprotection of humanin-like peptides in seminal plasma for ejaculated spermatozoa of crossbred bulls.

BACKGROUND: Cryopreservation process negatively affects spermatozoa functions. Humanin, a small polypeptide encoded in the mitochondrial genome, is well known for its role in cell survival. OBJECTIVE: To quantify the endogenous levels of humanin in seminal plasma of crossbred Frieswal bulls and to study its role in cryoprotection. The presence of humanin in bull spermatozoa was also investigated. MATERIALS AND METHODS: A total of 40 semen samples were separated into two groups based on the initial progressive motility (IPM): Good (IPM >70%) and Poor (IPM <50%) groups; and/or based on the post-thaw motility (PTM): Freezable (PTM>50%) and Non-freezable (PTM < 50%) groups. Humanin concentration in seminal plasma (SP-HN) was quantified using ELISA. RESULTS: SP-HN concentration ranged from undetectable to 67.6 pg/mL with a median level of 35.2 pg/mL. SP-HN level was significantly higher in the good quality semen group than in the poor quality semen group (p<0.001), and also significantly higher in the freezable group than in the non-freezable group (p<0.001). SP-HN level was positively correlated with initial progressive motility, post-thaw semen motility, viability, acrosome intactness and plasma membrane integrity, but negatively correlated the level of reactive oxygen species and malondialdehyde content. Immunochemical localization showed the presence of humanin in the proximal region of the middle piece of spermatozoa. CONCLUSION: Endogenous humanin level had significant correlation with semen quality and might protect sperm cells against freeze-induced oxidative stress. doi.org/10.54680/fr22510110712.

Effects of unconjugated gold, silver and titanium dioxide nanoparticles on bovine spermatozoa at various stages of cryopreservation.

BACKGROUND: The increasing use of nanoparticles (NP) for gender-selected spermatozoa, sperm-enriched semen and novel extenders raises the concern of undesirable effects on fertility and sperm function. OBJECTIVE: To investigate the effects of gold (Au-), silver (Ag-), and titanium dioxide (TiO2-) NPs on the motility and sperm functions in bovine spermatozoa at various stages of cryopreservation. MATERIALS AND METHODS: Frieswal (Sahiwal × Holstein Friesian) bull semen ejaculates (N = 24) were challenged with unconjugated and ligand-free Au-, Ag-, and TiO2-NPs. RESULTS: At post-dilution (fresh) stage, there was no significant difference observed in progressive motility and viability amongst the control and any nanoparticle-treated groups, though plasma membrane integrity was significantly reduced in nanoparticle-treated groups (p < 0.05). The acrosome intactness was also significantly reduced in the groups of Ag-NP and TiO2 -NP (p < 0.05), while there was no effect observed in the Au-NP group. At post-equilibration stage, a significant reduction in motility, viability, and plasma membrane integrity was observed in all three nanoparticle-treated groups (p < 0.05). There was no difference in intact acrosome between the control and Au-NPs groups; which was significantly higher than the Ag-NP and TiO2 -NP groups (p < 0.05). At post-thaw stage, all NP groups resulted in a significant reduction of motility, viability, acrosome intactness and plasma membrane integrity (p < 0.05). Besides, TiO2-NPs appear to be significant more toxic (p < 0.05) among three NP groups, and Au-NPs appear to be lesser toxic. CONCLUSION: Bovine spermatozoa are adversely affected by Au-, Ag- and TiO2 -NPs that may impair sperm motility and other functions. doi.org/10.54680/fr22310110512.

Antibiogram of microorganisms isolated from fresh and frozen semen of crossbred frieswal bulls.

BACKGROUND: The bacterial contaminants in the semen are a major concern for most of the semen production laboratories because they adversely affect the semen quality. During sperm cryopreservation, the inclusion of antimicrobials in extenders may help to minimize bacterial growth. However, due to bacterial resistance to commonly used antimicrobials, they cannot fully assure microbiological safety to the frozen semen. OBJECTIVE: To estimate the microbial load and antibiogram of microorganisms isolated from the fresh and frozen bull semen. MATERIALS AND METHODS: The bacterial load was estimated in fresh and frozen semen samples of crossbred Frieswal bulls by the pour plate method. Microorganisms were identified as Gram positive and Gram negative by Gram staining. The representative bacterial colonies were streaked onto different specific media which were further confirmed by biochemical tests. Bacterial isolates were subjected to in vitro antibiotic sensitivity test. RESULTS: The average microbial load of fresh and frozen semen samples was found to be 8397.4 ± 524.3 cfu per mL and 680.9 ± 105.4 cfuper mL, respectively. Microorganisms belonging to Staphylococcus aureus, Staphylococcus epidermidis, Proteus, Klebsiella, Bacillus cereus, Bacillus subtilis, Actinomyces, E. coli, Rhodococcus, Neisseria and Micrococcus were identified in the semen samples. The antibiotic sensitivity testing of the bacterial isolates revealed that benzyl penicillin was found to be the least effective against the isolated organisms while gentamicin and spectinomycin were found to be most effective among the antibiotics used. Lincomycin, tylosin and streptomycin showed moderate efficacy against the bacterial isolates. CONCLUSION: Gentamicin, tylosin, lincomycin, and spectinomycin (GTLS) antibiotic combination is more effective against bacterial isolates and may be added to semen extender to better control bacterial load and semen quality. doi.org/10.54680/fr22610110512.