一种桑树细菌性病原内生拮抗细菌的筛选、鉴定及其生防活性

Screening, Identification of an Antagonistic Endophyte to Pathogens of Mulberry Bacterial Diseases from Mulberry Leaves and Its Biocontrol Activity

摘要: 本文从健康桑树叶片中筛选对桑青枯病和桑疫病病原菌有拮抗作用的内生细菌，并对其生防机制初步探索。实验利用常规方法从严格表面消毒的桑树叶片中分离纯化内生菌，采用抑菌圈法筛选拮抗菌；同时有机溶剂分段萃取拮抗菌产物，抑菌圈法分析活性物质存在组段，并定性分析活性物质的种类。结果显示，从三个品种健康桑树叶片中分离获得7株内生细菌，其中3株具有拮抗桑树病原菌的作用，分离自品种9703的编号为381的菌株作用效果最强。结合形态学和生理生化特征以及16SrRNA序列分析鉴定其为多粘类芽孢杆菌(Paenibacillus polymyxa)。菌株381产生的活性抑菌物质主要存在于发酵液中，可用乙酸乙酯有效萃取。定性鉴定表明活性抑菌物质可能是生物碱、黄酮、二萜或三萜。同时平板定性分析显示菌株381可产生有机酸、纤维素酶和蛋白酶。研究结果对桑树细菌性病害无公害防治提供理论基础和参考依据。

Abstract: The endophytic bacteria which antagonized the pathogens of mulberry bacterial wilt and mulberry bacterial blight were screened from healthy mulberry leaves, and the biocontrol mechanism was also explored. The endophytic bacteria were isolated and purified from the mulberry leaves which were strictly surface-sterilized by conventional methods. The antagonistic bacteria were screened by the inhibition zone method. At the same time, the organic solvents were used to extract the products of the strain, and the active substance group was analyzed by the inhibition zone method, and the types of active substances were qualitatively analyzed. Results showed that 7 endophytic bacteria were isolated from the three healthy mulberry trees, in which 3 strains antagonized mulberry pathogens, and the strain numbered 381 isolated from 9703 had the strongest effect. Based on morphological, physiological and biochemical characteristics and 16S rRNA sequence analysis, the strain 381 was identified as Paenibacillus polymyxa. The active bacteriostatic substances produced by strain 381 existed mainly in the fermentation broth and could be effectively extracted with ethyl acetate. Qualitative tests indicated that the active bacteriostatic substances may be alkaloids, flavonoids, diterpenes or triterpenes. At the same time, qualitative analysis on the plates showed that the 381 strain could produce organic acids, cellulases and proteases. The results in this paper provided theoretical basis and reference basis for the pollution-free prevention and control of mulberry bacterial diseases.