How does the microbial load affect the quality of equine cool-stored semen?

Area of Science:

• Reproductive Biology
• Microbiology
• Sperm Cryopreservation

Background:

• Bacterial contamination in stallion ejaculates negatively impacts sperm quality during storage.
• Existing methods to reduce microbial load in semen have varying efficacy.
• Understanding the relationship between bacterial load, reactive oxygen species (ROS), and sperm function is crucial for improving preservation techniques.

Purpose of the Study:

• To evaluate bacterial elimination techniques in stallion semen.
• To investigate the correlation between total microbial load (TML) and ROS production.
• To determine the impact of TML on the functionality of cool-stored sperm.

Main Methods:

• Ejaculates from 11 stallions were processed using three methods: extended semen, conventional centrifugation, and Single Layer Centrifugation (SLC) with Androcoll-E.
• Bacteriological analysis was performed to determine Total Microbial Load (TML) at various storage intervals (0-72h at 5°C).
• Sperm viability, lipid peroxidation, motility (CASA), and ROS production (flow cytometry) were assessed.

Main Results:

• TML correlated significantly with sperm viability, H2O2 production, total ROS, and motility parameters (total, progressive, VCL, VSL, VAP).
• SLC removed 99.34% of the microbial load, significantly reducing H2O2 production.
• Bacterial load increased H2O2 production in cool-stored semen, decreasing sperm tolerance to refrigeration.

Conclusions:

• Single Layer Centrifugation (SLC) is the most effective technique for removing bacteria from stallion semen.
• Reducing bacterial load, particularly H2O2 production, via SLC improves sperm quality and functionality during cold storage.
• SLC facilitates the selection of more functional sperm by minimizing the detrimental effects of bacterial contamination.