Bacterial communities in bovine ejaculates and their impact on the semen quality

Although bacterial contamination of ejaculates may cause difficulties in cattle reproduction, standard protocols for a routine microbiological analysis of bovine semen are still missing. Understanding of the mechanisms of bacterial damage to spermatozoa may contribute to the prevention and management of bacteriospermia in the future. Therefore, this study was designed to investigate bacterial profiles of fresh bovine ejaculates (n = 30), while at the same time we focused on assessing the relationships between bacteriospermia and selected sperm quality parameters as well as an array of oxidative stress and inflammatory markers. The samples were divided into three quality groups according to the sperm motility: Excellent (EX) - over 90% > Good (GO) - between 89% and 80% > Moderate (MO) - under 80%. The results showed a significant increase in reactive oxygen species (ROS) generation in the GO group when compared to the EX group. In the MO group, a deterioration of almost all quality parameters was observed when compared to the EX group. In particular, sperm motility, mitochondrial membrane potential, ROS production and IL-6 concentration exhibited a significant decline. Pearson correlation analysis revealed positive associations among bacterial load and the presence of leukocytes in semen (r = 0.965), malondialdehyde concentration (r = 0.816) and DNA fragmentation (r = 0.784). MALDI-TOF MS Biotyper analysis showed a prevalence of the Staphylococcus genus. The quantification of bacterial colonies revealed a significantly increased (P < 0.01) bacterial load in the MO group when compared with the EX as well as the GO group. Overall, our results suggest that sperm quality may be affected by both, bacterial composition, and bacterial load. It appears that an increased presence of bacterial species triggers the immune response, causes oxidative stress, and thereby contributes to sperm structural alterations while diminishing their fertilization ability.Abbreviations: EX: Excellent; GO: Good; MO: Moderate; MOT: Motility; ROS: Reactive Oxygen Species; MMP: Mitochondrial Membrane Potential; IL-1: Interleukin 1; IL-6: Interleukin 6; IL-8: Interleukin 8; IL-12: Interleukin 12; CRP: C-reactive protein; DNA: Deoxyribonucleic acid; MALDI-TOF MS: Matrix-assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry; LPO: Lipid peroxidation; CFU: Colony-forming units MDA: Malondialdehyde; CASA: Computer-assisted Sperm Analysis; WS: Working solution; RIPA: Radio-immunoprecipitation assay; TBARS: Thiobarbituric Acid Reactive Substances; BHB: D-β-hydroxybutyrate.