紫花地丁根际菌的筛选鉴定及活性物质初步研究

Isolation and identification of actinomycetes from rhizosphere of viola yedoensis and preliminary research on its active substance

目的从药用植物紫花地丁根部土壤中筛选出具有抗菌活性的根际菌并对其活性物质进行初步分析。方法采用稀释涂布法分离根际菌,以金黄色葡萄球菌、大肠埃希菌、表皮葡萄球菌、铜绿假单胞菌、费氏柠檬酸杆菌为指示菌经固定琼脂块法筛选活性菌株;通过16 s rDNA基因序列数据分析及系统发育进化树确定活性菌株的分类;采用纸片扩散法测定活性菌株发酵液抗菌活性,采用萃取法分析菌株发酵液抗菌活性物质极性。结果共分离出21株根际菌,其中2株活性菌株对指示菌有较强抑菌作用,分别命名为Y1-7、Y1-13a;菌株Y1-7为Streptomyces sp.属,菌株Y1-13a为Streptomyces californicus属。活性菌株发酵液对金黄色葡萄球菌、大肠埃希菌、表皮葡萄球菌均具有较强抑菌作用,抑菌圈直径10.5～17.0 mm,且Y1-13a对铜绿假单胞菌及费氏柠檬酸杆菌具有较强抑菌作用。Y1-7发酵液抑菌活性物质为脂溶性物质,Y1-13a抑菌活性物质为水溶性物质。结论菌株Y1-7、Y1-13a次级代谢产物均有一定的抑菌作用,具有进一步研究及开发利用的价值。

Objective To screen rhizosphere bacteria with antimicrobial activity from root soil of medicinal plant, and to analyze their active substances. Methods Rhizosphere bacteria were isolated by dilution coating method. Staphylococcus aureus, Escherichia coli, Staphylococcus epidermidis, Pseudomonas aeruginosa and Citrate fernii were used as indicator bacteria to screen the active strains. The 16 s rDNA gene sequence data and phylogenetic tree were constructed to determine its taxonomic position. The antibacterial activity of the fermentation broth of the active strain was determined by disk diffusion method and the polarity of the antimicrobial active substance was analyzed by the extraction method. Results A total of 21 rhizosphere strains were isolated, and 2 active strains, named Y1-7, Y1-13 a and Y1-7 as Streptomyces sp., were screened out. The strain Y1-13 a belongs to Streptomyces californicus genus, and the fermentation broth of active strain has strong bacteriostatic effect on Staphylococcus aureus, Escherichia coli and Staphylococcus epidermidis. The diameter of inhibition circle was 10.5 ～ 17.0 mm, and Y1-13 a had a strong bacteriostatic effect on Pseudomonas aeruginosa and Citrate fermentum fermentative broth. The bacteriostatic active substance of Y1-7 fermentation broth was lipophilic substance Y1-13 a antibacterial active substance was watersoluble substance, and Y1-13 a had strong bacteriostatic effect on Pseudomonas aeruginosa and Citrate fermentum. Conclusion The secondary metabolites of strain Y1-7 and Y1-13 a have certain bacteriostatic effect, which has the value of further research and utilization.