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渗透压对嗜线虫致病杆菌YL001生长及抑菌活性的影响
李忝珍1, 张淑静1, 刘 启,等1
西北农林科技大学 无公害农药研究服务中心
摘要:
【目的】研究渗透压对嗜线虫致病杆菌(Xenorhabdus nematophila)YL001菌体生长、抑菌活性的影响,为进一步提高YL001抑菌活性及开发利用奠定基础。【方法】以NaCl为渗透压调节剂,分别测定其质量浓度为0,5,10,20,30和40 g/L条件下,嗜线虫致病杆菌YL001不同时间段细胞生长量、pH、还原糖变化;采用琼脂扩散法、生长速率法及活体组织法测定了YL001发酵液的抑菌活性;同时采用HPLC及LC-MS分析了甲醇提取物中酰胺类化合物(Nematophin)与水溶性苯并芘类化合物(Xenocoumacins,包括XCN1和XCN2)含量差异。【结果】在NaCl质量浓度为0,5 g/L时,YL001细胞生长最适且无显著差异,吸光值分别达0.63,0.62,随着NaCl质量浓度的进一步增大细胞生长量显著下降;在NaCl质量浓度为0,5,10 g/L时,YL001发酵液对苏云金芽孢杆菌、蜡状芽孢杆菌、猕猴桃溃疡病菌、金黄色葡萄球菌的抑菌作用较好且无显著差异,随着NaCl质量浓度的进一步增大抑菌作用显著降低;在NaCl质量浓度为0,5 g/L时,YL001发酵液对水稻稻瘟病菌、番茄灰霉病菌、苹果树腐烂病菌、苹果轮纹病菌、苹果炭疽病菌、水稻纹枯病菌、油菜菌核病菌的抑制率均高于80%且均无显著差异,明显优于其他较高NaCl质量浓度的处理;在NaCl质量浓度为0,5,10 g/L时,YL001发酵液对番茄灰霉病的治疗作用较好且无显著差异,在NaCl质量浓度为0,5 g/L时,对番茄灰霉病的保护作用较好且无显著差异,治疗作用和保护作用均随着NaCl质量浓度的进一步增大而显著下降;甲醇提取物中抑菌活性物质Nematophin含量、XCN1的相对含量随NaCl质量浓度的增大显著下降,如NaCl质量浓度为0 g/L时,甲醇提取物中抑菌活性物质Nematophin含量、XCN1的相对含量较NaCl质量浓度为40 g/L时分别提高7.82和27.55倍;XCN2的相对含量在NaCl质量浓度为5 g/L时最高,显著高于其他处理。【结论】低渗透压有利于X.nematophila YL001菌体生长、抑菌活性提高及抑菌活性代谢产物Nematophin、Xenocoumacins的产生。
关键词:  嗜线虫致病杆菌  渗透压  生长规律  抑菌活性  Nematophin  Xenocoumacins
DOI:
分类号:
基金项目:公益性行业(农业)科研专项(200903052);国家自然科学基金项目(31171910);陕西省自然科学基金重点项目(2014JZ004);中央高校基本科研创新重点项目(Z109021304)
Effect of osmotic stress on in vitro growth and antimicrobial active constituents of Xenorhabdus nematophila YL001
LI Tianzhen,ZHANG Shujing,LI Tianzhen1,ZHANG Shujing1,LIU Qi,et al
Abstract:
【Objective】This study investigated the effects of osmotic stress on growth,antimicrobial activity and active secondary metabolites of Xenorhabdus nematophila YL001 to provide foundation for improving antimicrobial activity and utilization of YL001.【Method】The growth,pH and glucose residue of X.nematophila YL001 were measured at different periods under different salt concentrations (0,5,10,20,30 and 40 g/L).Antimicrobial activities of fermented liquid of X.nematophila YL001 were measured using growth rate,agar diffusion method,and biopsy method.The differences of metabolites Nematophin and Xenocoumacins (including XCN1 and XCN2) in methanol extract were detected by HPLC and LC-MS,respectively.【Result】At the NaCl concentrations of 0 and 5 g/L,YL001 cells had the best growth without significant difference,and the absorbance values reached 0.63 and 0.62,respectively.With the further increase of NaCl,the growth decreased significantly.The inhibitory effects of YL001 against Bacillus thuringiensis,Bacillus cereus,Pseudomonas syringae and Staphylococcus aureus at NaCl concentrations of 0,5 and 10 g/L were best without significant difference,and the antibacterial activity gradually decreased with the increase of NaCl concentration.The inhibitory rates against Magnaporthe grisea,Botryti cinerea,Valsa mali Miyabe et Yamada,Physalospora piricola,Clomerela cinyulate,Thanatephorus cumeris,and Sclerotinia sclerotiorum were higher than 80% without significant difference under the concentrations of 0 and 5 g/L,and the effects were better than that of higher NaCl concentrations.Protective effect against Botryti cinerea was best at NaCl concentrations of 0,5 and 10 g/L without significant difference,while the therapeutic effect was best at the concentrations of 0 and 5 g/L without significant difference.Both effects decreased significantly with the further increase of NaCl concentration.The content of Nematophin and the relative content of XCN1 in methanol extract decreased drastically with the increase of NaCl concentration.Their concentrations at 0 g/L NaCl were 7.82 and 27.55 times of that at 40 g/L NaCl.The highest relative content of XCN2 was obtained at 5 g/L NaCl,and was significantly higher than other treatments.【Conclusion】Low osmotic pressure is more conducive to cell growth and improves antimicrobial activity and accumulation of Nematophin and Xenocoumacins of X.nematophila YL001.
Key words:  Xenorhabdus neamatophila  osmotic pressure  in vitro growth  antimicrobial activity  Nematophin  Xenocoumacins