利迪链霉菌A01活性代谢产物对甘蓝枯萎病菌的抑制作用及其机理

The Antifungal Activity and Action Mechanism of Metabolite Produced by Streptomyces lydicus Strain A01 Against Fusarium oxysporum f.sp.conglutinans

【目的】明确生防利迪链霉菌A01菌株的活性代谢产物纳他霉素对甘蓝枯萎病菌的抑制作用及其机理,为其下一步的产品开发和应用提供科学依据。【方法】分别采用带毒平皿法和凹玻片法测定活性产物对病菌菌丝生长和孢子萌发的抑制作用,采用电导率法和美蓝染色法分别测定其对菌丝体细胞膜通透性和细胞存活性的影响,借助电镜观察其对菌丝超微形态和结构的影响。【结果】纳他霉素对甘蓝枯萎病菌菌丝生长和孢子萌发的最低抑菌浓度分别小于或等于31.92和41.04μg.mL-1,抑制中浓度分别为8.69和3.06μg.mL-1;30μg.mL-1以上浓度的纳他霉素处理90 min后可使菌丝悬浮液电导率明显升高,其作用强度与纳他霉素的浓度和作用时间呈正相关;处理后的菌丝生长异常或畸形,并出现细胞器解体、细胞液泡化等现象;10μg.mL-1以上纳他霉素处理120 min后菌丝大部分可被美蓝染成蓝色。【结论】纳他霉素对甘蓝枯萎病菌菌丝生长和孢子萌发均有明显的抑制作用,能够增加病菌细胞膜的通透性,破坏菌丝细胞的形态和结构,导致其丧失存活能力。

[Objective] The objective of this study is to understand the antifungal activity and action mechanism ofnatamycin produced by Streptomyces lydicus strain A01 against Fusarium oxysporum f. sp. conglutinans, and to provide a scientific basis for product development and application of strain A01. [Method] Mycelial radial growth assay and microscope concave slide method were used to test suppression of mycelia growth and inhibition of spore germination. Electrical conductivity method and methylene blue stain were used to test permeability of pathogen plasma membrane and cell viability of this pathogen. SEM and TEM observations were used to observe morphology and ultrastructure of mycelia. [ Result ] The MICs of natamycin against the mycelia growth and the conidial germination of F. oxysporum f. sp. conglutinans were less than or equal to 31.92 and 41.04 μg·mL-1, respectively, and the IC50 and EC50 were 8.69 and 3.06μg·mL-1, respectively. After being treated by natamycin at concentration above 30 μg·mL-1 for more than 90 min, the electrical conductivity of the mycelial suspension was significantly increased. The action intensity was correlated positively with natamycin concentration and the action time. The changes in morphology and cell ultrastructure of the treated mycelia were observed under SEM and TEM, such as abnormal growth or malformation, disintegration of organelles, cell vacuolization and so on. The mycelia treated with natamycin over 10μg·mL-1 for 120 min could be stained with methylene blue. [ Conclusion ] The active metabolite of strain A01, natamycin, has obvious inhibitory activities against F. oxysporum f. sp. conglutinans. The bioactivity was based on that natamycin could increase the permeability of plasma membrane, destroy the mycelial morphology and cell ultrastructure, thus resulted in the disappearance of cell viability of the pathogen mycelium.