云南金铁锁根可培养放线菌多样性及其抗菌活性研究

Diversity and antimicrobial activities of culturable endophytic actinomyces in the roots of Psammosilene tunicoides in Yunnan province

摘要:

【目的】探索云南金铁锁根部可培养放线菌资源及筛选高抗病原菌活性菌株。【方法】以云南4个地方金铁锁的根为研究对象，采用不同温度预处理及14种分离培养基对其进行放线菌分离；选用74株内生放线菌菌株，以5种病原菌为指示菌，使用倒置法测定供试放线菌的抗菌活性。【结果】从金铁锁根部分离获得121株内生放线菌，基于16S rRNA基因序列比对和系统发育分析，将其归属于10个目、15个科和24个属，其中链霉菌(Streptomyces)为绝对优势菌群，其后依次为诺卡氏菌属(Nocardia)、小单孢菌属(Micromonospora)和分枝菌酸小杆菌属(Mycolicibacterium)。分离效果最好的培养基为D4(酵母粉0.3 g，干酪素0.3 g，葡萄糖0.3 g，骨粉0.3 g)；最优的预处理温度为80℃；74株内生放线菌中的37株对至少一种指示病原菌具有抑制活性，且活性强的主要类群为链霉菌属。【结论】金铁锁根部蕴藏着丰富的放线菌种质资源，且蕴含着具有产次级代谢产物能力的菌株，这将为后续的医药及农业生产提供了丰富的菌种资源，同时为其栽植、保护、开发及应用提供了一定的基础数据和理论依据。

Abstract:

[Objective] To investigate the diversity of culturable endophytic actinomyces in the roots of Psammosilene tunicoides in Yunnan province and screen out the strains with high antimicrobial activities.[Methods]The roots of P.tunicoides from four different places in Yunnan province were pretreated at different temperatures, from which the culturable endophytic actinomyces were isolated with 14 media. The antimicrobial activities of 74 isolates against 5 pathogens were evaluated with inverted plates. [Results]Totally 121 strains of endophytic actinomyces were isolated from the roots of P.tunicoides. The results of 16S rRNA gene sequence alignments and phylogenetic analysis showed that the 121 strains could be categorized to 24 genera, 15 families of 10 orders. Streptomyces was the absolutely dominant genus, followed by Nocardia, Micromonospora, and Mycolicibacterium. D4 (0.3 g yeast powder, 0.3 g casein, 0.3 g glycose, 0.3 g bone meal) was the best medium for isolation among all the tested media and 80 ℃ was the optimal pretreatment temperature. Among 74 strains of endophytic actinomyces, 37 strains showed inhibitory activities against at least one indicator pathogen, and the main genus with strong antimicrobial activity was Streptomyces. [Conclusion] The roots of P.tunicoides harbor rich actinomyces and the functional strains with the ability to produce secondary metabolites, which can provide abundant strain resources for pharmaceutical and agricultural production in the future as well as basic data and a theoretical basis for the cultivation, protection, development, and application of P.tunicoides.