ε-聚赖氨酸对关中黑猪精液常温保存效果的研究

发布时间：2018-05-20 03:35

本文选题：关中黑猪精液 + ε-聚赖氨酸　；参考：《西北农林科技大学》2017年硕士论文

【摘要】：近年来由于抗生素不合理使用,致使部分细菌对常规抗生素产生耐药性。若继续在稀释液中使用常规抗生素,可能致使耐药细菌通过精液感染母猪。为减少抗生素的使用,降低细菌耐药性的产生和提供更高生物安全的关中黑猪精液用于深部输精,本研究在稀释液中添加一种具有广谱抑菌能力的天然的阳离子抗菌肽——ε-聚赖氨酸用于精液常温保存。首先,本试验采用高通量测序法检测了不同时间段精液中细菌的种类,并通过16SrDNA法鉴定了公猪精液中分离的部分细菌的菌属;其次,选择精液中常见的细菌作为体外抑菌试验菌种,测定ε-聚赖氨酸对其最小抑菌浓度;最后,探究了不同浓度ε-聚赖氨酸和0.16 g/Lε-聚赖氨酸替代部分庆大霉素对精液常温保存效果的影响。本研究获得的主要结果如下:1.常温保存第1天精液中的细菌主要为革兰氏阳性菌,如表皮葡萄球菌、地衣芽孢杆菌和浅绿气球菌,常温保存第5天主要为革兰氏阴性菌,如鲁菲不动杆菌、木糖氧化无色杆菌和嗜麦芽寡养单胞菌。2.ε-聚赖氨酸对鲁菲不动杆菌,表皮葡萄球菌、浅绿气球菌和大肠杆菌均有抑制效果;表皮葡萄球菌和大肠杆菌的最小抑菌浓度为0.02 g/L,鲁菲不动杆菌和浅绿气球菌最小抑菌浓度为0.04 g/L,而ε-聚赖氨酸对木糖氧化无色杆菌和嗜麦芽寡养单胞菌的抑制效果不佳。仅添加50%和25%标准添加量的庆大霉素(0.25 g/L)均对木糖氧化无色杆菌和嗜麦芽寡养单胞菌的抑制效果不佳;而0.16 g/Lε-聚赖氨酸可分别替代50%和75%标准添加量的庆大霉素,取得与标准添加量庆大霉素相近的抑菌效果(P0.05)。3.在稀释精液中添加0.04~0.64 g/L的ε-聚赖氨酸不影响其渗透压(P0.05),当添加1.28 g/Lε-聚赖氨酸时显著提高其渗透压(P0.05),0.04~1.28 g/Lε-聚赖氨酸均能有效抑制精液中细菌的生长和提高精液的常温保存效果;其中0.16 g/Lε-聚赖氨酸常温保存效果最佳(P0.05)。4.进一步研究发现稀释液中,ε-聚赖氨酸与庆大霉素具有协同抑菌的作用;0.16 g/Lε-聚赖氨酸可分别替代50%和75%标准添加量的庆大霉素,并取得与标准添加量庆大霉素相近的抑菌效果和精液常温保存效果(P0.05)。综上所述,关中黑猪精液中的革兰氏阴性菌为优势菌群,0.04 g/Lε-聚赖氨酸有效抑制表皮葡萄球菌,大肠杆菌、鲁菲不动杆菌、浅绿气球菌的生长;仅添加ε-聚赖氨酸对木糖氧化无色杆菌和嗜麦芽寡养单胞菌抑制效果不佳,进一步研究发现0.16 g/Lε-聚赖氨酸可替代部分庆大霉素用于抑制这两种细菌生长;0.04 g/L~1.28 g/Lε-聚赖氨酸均能提高精液的保存效果,其中其最适添加量为0.16 g/L;进一步研究发现ε-聚赖氨酸与庆大霉素在稀释液中具有协同抑菌的作用。
[Abstract]:In recent years, some bacteria have become resistant to conventional antibiotics due to irrational use of antibiotics. Continued use of conventional antibiotics in diluents may cause drug-resistant bacteria to infect sows through semen. In order to reduce the use of antibiotics, reduce the production of bacterial resistance and provide higher biosafety of Guanzhong black pig semen for deep insemination, In this study, a natural cationic antimicrobial peptide-蔚-Polysine was added to the diluent to preserve semen at room temperature. Firstly, high throughput sequencing method was used to detect the species of bacteria in semen of different periods, and the genus of some bacteria isolated from boar semen was identified by 16SrDNA method. The common bacteria in semen were selected as bacteriostasis test strains in vitro, and the minimal inhibitory concentration of 蔚 -polylysine on them was determined. The effects of different concentrations of 蔚 -polylysine and 0.16 g / L 蔚 -poly-lysine on the preservation of semen at room temperature were investigated. The main results of this study are as follows: 1. On the first day of cryopreservation, the bacteria in semen were mainly Gram-positive bacteria, such as Staphylococcus epidermidis, Bacillus licheniformis and Balloon palliformis, and Gram-negative bacteria, such as Acinetobacter Ruffiensis, were mainly found in semen on the 5th day of preservation at room temperature. Acinetobacter lufii, Staphylococcus epidermidis, Balloonus graminis and Escherichia coli were all inhibited by Xyloxide and Maltobacillus maltophilia. The minimum inhibitory concentration of Staphylococcus epidermidis and Escherichia coli was 0.02 g / L, the minimum inhibitory concentration of Acinetobacter Ruffiensis and Anoacter palliforme was 0.04 g / L, while 蔚 -Polylysine was not good in inhibiting xylose oxidized and maltophilia. Only 50% and 25% standard addition of gentamicin 0.25 g / L) could not inhibit Xymoxanthobacterium and Maltophotrophomonas maltophilia, while 0.16 g / L 蔚 -Polysine could replace 50% and 75% standard addition of gentamicin, respectively. The antimicrobial effect of gentamicin was similar to that of the standard dosage of gentamicin. Adding 0.04 ~ 0.64 g / L 蔚 -polylysine to diluted semen did not affect the osmotic pressure (P _ (0.05). When 1.28 g / L 蔚 -polylysine was added, the osmotic pressure was significantly increased (P _ (0.05) P _ (0.04) and 1.28 g / L ~ (-) -polylysine could effectively inhibit the growth of bacteria in semen and improve the cryopreservation effect of semen at room temperature. Among them, 0.16 g / L 蔚 -Poly (lysine) was best preserved at room temperature (P0.05N. 4). Further study showed that 蔚 -polylysine and gentamicin had synergistic bacteriostasis effect in dilution solution, 0.16 g / L 蔚 -poly-lysine could replace 50% and 75% standard addition of gentamicin, respectively. The bacteriostatic effect of gentamicin was similar to that of standard dosage and the effect of semen preservation at room temperature was obtained (P 0.05). In conclusion, Gram-negative bacteria in the semen of Guanzhong Black Pig were the dominant bacteria group (0.04 g / L 蔚 -Polysine) which could effectively inhibit the growth of Staphylococcus epidermidis, Escherichia coli, Acinetobacter lufi and Balloonus palliforme. The inhibitory effect of 蔚 -polylysine only on Xyloxide and Maltophostomonas maltophilia was not good. It was found that 0.16 g / L 蔚 -polylysine could be used to inhibit the growth of these two bacteria by 0.04 g/L~1.28 / L 蔚 -Polysine instead of gentamicin. The optimum addition amount was 0.16 g / L, and it was found that 蔚 -polylysine and gentamicin had synergistic bacteriostatic effect in the diluent.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S828

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