Puig-Timonet Adrià, Castillo-Martín Miriam, Pereira Barbara A, Pinart Elisabeth, Bonet Sergi, Yeste Marc
Department of Biology, Institute of Food and Agricultural Technology, University of Girona, Spain.
Department of Biology, Institute of Food and Agricultural Technology, University of Girona, Spain.
Theriogenology. 2018 Apr 15;111:9-18. doi: 10.1016/j.theriogenology.2018.01.014. Epub 2018 Feb 3.
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 10 and 10 CFU mL. 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 × 10 ± 6.2 × 10 CFU mL and PBD2: 9.1 × 10 ± 2.4 × 10 CFU mL) and 5 μM (day 10, PBD1: 1.2 × 10 ± 5.1 × 10 CFU mL; PBD2: 8.7 × 10 ± 2.9 × 10 CFU mL), the control with antibiotic was found to be more effective (day 10, 8.3 × 10 ± 1.6 × 10 CFU mL). 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.
本研究评估了两种不同的抗菌肽（AMP）：猪β-防御素-1（PBD1）和-2（PBD2）在三种浓度（1.5μM、3μM和5μM）下能否作为液态保存公猪精液中抗生素的合适替代品。对保存在17°C下9 - 10天的液态公猪精液进行了两项独立实验。在第一个实验中，我们评估了添加每种AMP的三种浓度对保存10天的公猪精液细菌生长和精子质量的影响。在第二个实验中，在人工接种10⁵和10⁶CFU/mL大肠杆菌后，测定了这些AMP在9天内控制细菌生长的能力。在两个实验中，通过流式细胞术评估精子活力，用计算机辅助精子分析（CASA）测定精子活力，并通过在Luria Bertani琼脂上进行细菌培养来评估对微生物生长的抑制作用。发现PBD1和PBD2在5μM时显著（P < 0.05）降低精子活力（第10天总活动精子百分比，对照组：31.6%±1.2%，PBD1：6.5%±0.3%，PBD2：5.6%±0.4%）。尽管在3μM（第10天，PBD1：1.4×10³±6.2×10³CFU/mL，PBD2：9.1×10³±2.4×10³CFU/mL）和5μM（第10天，PBD1：1.2×10³±5.1×10³CFU/mL；PBD2：8.7×10³±2.9×10³CFU/mL）时观察到对细菌生长的最高抑制作用，但发现抗生素对照组更有效（第10天，8.3×10²±1.6×10²CFU/mL）。总之，PBD1和PBD2可以以3μM的浓度添加到不含抗生素的公猪精液稀释液中，但不能完全控制所有细菌生长。
