东乡野生稻内生放线菌分离及菌株S123次级代谢产物分析

Isolation of endophytic actinomycetes from Dongxiang wild rice (Oryza rufipogon) and analysis of secondary metabolite of active strain S123

摘要:

【目的】从东乡野生稻(Oryza rufipogon)中分离和鉴定内生放线菌，对其进行抗菌活性筛选，并分析高抗菌活性菌株S123的次级代谢产物。【方法】采用S培养基对东乡野生稻内生放线菌进行分离、纯化，并构建16S rRNA基因序列系统发育进化树进行菌株鉴定。以琼脂扩散法和菌丝生长速率法进行抗菌活性筛选，同时设计简并引物检测菌株I型聚酮合酶(PKS-I)基因。对具广谱抗菌活性的菌株S123进行分批大量发酵，运用多种色谱方法对发酵产物进行分离、纯化，利用MS和NMR分析鉴定化合物的结构。【结果】从东乡野生稻中共分离到11株内生放线菌，分别属于链霉菌属(8株)和假诺卡氏属(3株)。其中有8株具有抗菌活性，8株呈现I型PKS阳性。从高抑菌活性菌株S123中分离到化合物Nigericin和17-O-demethylgeldanamycin，其中Nigericin对金黄色葡萄球菌、枯草芽孢杆菌及水稻纹枯病菌均有抑制活性。【结论】对东乡野生稻内生放线菌进行了分离、鉴定和抗菌活性筛选，并从中分到两种与I型PKS基因相关活性的化合物Nigericin和17-O-demethylgeldanamycin，为研究东乡野生稻内生放线菌的多样性和次级代谢产物的分离提供依据。

关键词:东乡野生稻，内生放线菌，I型PKS，抗菌活性，次级代谢产物

Abstract:

[Objective] To isolate endophytic actinomycetes from Dongxiang wild rice (Oryza rufipogon), and to study the classification, antimicrobial activity and analysis the secondary metabolites from the high active strain S123. [Methods] S medium was used to isolate endophytic actinomycetes from Dongxiang wild rice. The isolates were classified by 16S rRNA gene sequence analysis. Antimicrobial activity of the isolates was tested using agar diffusion and growth rate method. Meanwile, to screen the potential activity compounds, The degenerated primer targeted on type-I polyketide synthase (PKS-I) was designed. Bioactivity strain S123 was subjected to large scale fermentation for isolating secondary metabolites. The compounds were separated and purified by various chromatographic methods, and the structure was determined by mass spectrum and nuclear magnetic resonance analyses. [Results] We obtained 11 actinomycete isolates from Dongxiang wild rice. The isolates belonged to 2 genera, Streptomyces (8) and Pseudonocardia (3). Among of these isolates 8 strains show their anti-microbial activities and 8 strains were positive for type-I PKS genes. Two compounds were isolated from strain S123 and determined as Nigericin and 17-O-demethylgeldanamycin separately. The compound Nigericin showed inhibition against Staphylococcus aureus, Bacillus subtilis and Rhizoctonia solani. [Conclusion] This is the first report of endophytic actinomycetes from Dongxiang wild rice and identified two activity compounds Nigericin and 17-O-demethylgeldanamycin associated with type-I PKS genes. It provides a reliable basis for studying Dongxiang wild rice endophytic actinomycetes diversity and seeking activity secondary metabolites.

Key words:Dongxiang wild rice (Oryza rufipogon), Endophytic actinomycetes, Type-I PKS genes, Antimicrobial activity, Secondary metabolite