独岛枝芽胞杆菌MCCC 1A00493产生的4-乙烯基苯酚鉴定和作用南方根结线虫特性研究

Identification and characteristics of 4-vinylphenol from Virgibacillus dokdonensis MCCC 1A00493 against Meloidogyne incognita

摘要:

【目的】根结线虫危害严重，难防难控，前期研究发现一株深海来源的独岛枝芽胞杆菌（Virgibacillus dokdonensis）MCCC 1A00493对南方根结线虫具有良好的体外拮抗效果，本研究分离鉴定菌株发酵液中杀线虫活性物质，对其作用线虫的多种模式进行研究，为菌株有效控制植物病原线虫的应用奠定理论基础。【方法】采用硅胶柱层析和半制备高效液相色谱对独岛枝芽胞杆菌发酵上清液中的杀线虫活性物质进行分离纯化，采用液相色谱质谱联用、核磁共振对纯化物进行结构鉴定；并对其趋避、诱杀、熏杀和卵孵化抑制活性进行检测。【结果】结构鉴定确定独岛枝芽胞杆菌MCCC 1A00493发酵液中分离的杀线虫活性物质为4-乙烯基苯酚。4-乙烯基苯酚对线虫具有触杀、熏杀、卵孵化抑制和高浓度驱避低浓度引诱活性，15 μg/mL 4-乙烯基苯酚72 h触杀线虫校正死亡率为71.23%±9.06%；20 mg/mL 4-乙烯基苯酚24 h熏杀线虫死亡率达100%；100 μg/mL 4-乙烯基苯酚作用线虫卵10 d后，卵孵化抑制率达66.2%；在琼脂平板上，10 mg/mL的4-乙烯基苯酚对线虫有驱避作用，1 mg/mL的4-乙烯基苯酚对线虫有引诱作用；在Pluronic凝胶中，25 mg/mL的4-乙烯基苯酚驱避线虫，5 mg/mL和10 mg/mL的4-乙烯基苯酚引诱并杀死线虫。【结论】首次报道4-乙烯基苯酚对植物病原线虫具有多种作用模式，尤其是高浓度下对根结线虫具有趋避作用而低浓度下具有诱杀作用的新特性。因此，4-乙烯基苯酚可以作为一种潜在的生物杀线虫剂。

关键词:南方根结线虫;独岛枝芽胞杆菌;4-乙烯基苯酚;杀线虫活性;作用模式

Abstract:

[Objective] Root-knot nematode is destructive and difficult to be controlled. Virgibacillus dokdonensis MCCC 1A00493 is antagonistic to Meloidogyne incognita in vitro. In this study, we isolated and identified the nematicidal compound in the fermentation broth of MCCC 1A00493 and explored the modes of action, aiming at laying a theoretical foundation for the effective control of plant pathogenic nematodes.[Methods] The nematicidal compound in the supernatant of MCCC 1A00493 fermentation broth was isolated and purified by silica gel column chromatography and semi-preparative high-performance liquid chromatography, and the structure of the purified product was identified by liquid chromatography-mass spectrometry and nuclear magnetic resonance. The repellent activity, trapping activity, fumigation activity, and egg hatching-inhibiting activity of the compound were tested.[Results] The compound was identified to be 4-vinylphenol, which had the contact activity, fumigation activity, egg hatching-inhibiting activity, repellent activity at high concentration, and trapping activity at low concentration against the nematodes. At 15 μg/mL, 4-vinylphenol demonstrated contact activity, as manifested by the high corrected death rate (71.23%±9.06%) of M. incognita after 72 h. At 20 mg/mL, it showed fumigation activity and the death rate was 100% after 24 h. Inhibition rate of M. incognita egg hatching reached 66.2% after 10 days of exposure to 100 μg/mL 4-vinylphenol. Nematodes were repelled by 10 mg/mL, but attracted by 1 mg/mL 4-vinylphenol on agar plates. In Pluronic gel, nematodes were repelled by 25 mg/mL, and attracted and killed by 5 and 10 mg/mL 4-vinylphenol.[Conclusion] The compound 4-vinylphenol, identified from V. dokdonensis, possessed multiple modes of action against M. incognita, especially the repellent effect at high concentration and trapping effect at low concentration. Thus, it can be used as a potential bionematicide.

Key words:Meloidogyne incognita;Virgibacillus dokdonensis;4-vinylphenol;nematicidal activity;mode of action