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Evaluation of porcine beta defensins-1 and -2 as antimicrobial peptides for liquid-stored boar semen: Effects on bacterial growth and sperm quality

Puig-Timonet, Adrià, Castillo-Martín, Miriam, Pereira, Barbara A., Pinart, Elisabeth, Bonet, Sergi, Yeste, Marc
Theriogenology 2018 v.111 pp. 9-18
Escherichia coli, agar, antibiotics, antimicrobial peptides, boars, computers, flow cytometry, microbial culture, microbial growth, semen, sperm motility, sperm quality, viability
The present study evaluated whether two different antimicrobial peptides (AMP): porcine beta defensins-1 (PBD1) and −2 (PBD2) at three concentrations (1.5 μM, 3 μM and 5 μM) could be a suitable alternative to antibiotics in liquid-stored boar semen. Two separate experiments were conducted with liquid-stored boar semen preserved at 17 °C for 9–10 days. In the first one, we evaluated the impact of adding three concentrations of each AMP on the bacterial growth and sperm quality of boar semen stored for 10 days. In the second experiment, the ability of these AMPs to control bacterial growth was determined over a 9-day period, following artificial inoculation with Escherichia coli at 107 and 108 CFU mL−1. In both experiments, sperm viability was assessed through flow cytometry, sperm motility was determined with Computer Assisted Sperm Analysis (CASA) and the inhibitory effect on microbial growth was evaluated by bacteria culture on Luria Bertani agar. PBD1 and PBD2 were found to significantly (P < 0.05) decrease sperm motility at 5 μM (% total motile spermatozoa at day 10, Control: 31.6% ± 1.2% vs. PBD1: 6.5% ± 0.3% and PBD2: 5.6% ± 0.4%). Although the highest inhibitory effect on bacterial growth was observed at 3 μM (day 10, PBD1: 1.4 × 106 ± 6.2 × 105 CFU mL−1 and PBD2: 9.1 × 105 ± 2.4 × 105 CFU mL−1) and 5 μM (day 10, PBD1: 1.2 × 105 ± 5.1 × 104 CFU mL−1; PBD2: 8.7 × 104 ± 2.9 × 104 CFU mL−1), the control with antibiotic was found to be more effective (day 10, 8.3 × 103 ± 1.6 × 103 CFU mL−1). In conclusion, PBD1 and PBD2 may be added to antibiotic-free extenders for boar semen at a concentration of 3 μM, but do not completely control all bacterial growth.