泉州湾红树林根际土壤放线菌的分离鉴定及链霉菌W444次级代谢产物的鉴定

Isolation and identification of actinomycetes from rhizosphere soil samples of mangrove forests in Quanzhou Bay and secondary metabolites of Streptomyces sp. W444

摘要:

目的 从泉州湾2种红树林的根际土壤中分离并鉴定放线菌，进行抗真菌活性初筛以获取目标菌株，并对链霉菌属(Streptomyces) W444的次级代谢产物进行分离与鉴定。方法 从泉州湾洛阳江采集2种不同红树林植物的根际土壤样品，采用稀释涂布法对土样中的放线菌进行分离，并构建16S rRNA基因系统发育树。采用琼脂打孔扩散法进行抗真菌活性筛选，对获取的目标菌株Streptomyces sp. W444进行放大规模发酵，并对其次级代谢产物进行了分离与鉴定。根据生物合成基因簇定位和分析，推导星孢菌素的生物合成途径。结果 从根际土样中分离得到56株放线菌，分布于6目6科8属，其中链霉菌属和小单孢菌属为优势菌，占比分别为41.0%和33.9%。通过抗真菌筛选获得抑菌活性较好的菌株Streptomyces sp. W444，并从中分离鉴定了3个吲哚类化合物：staurosporine、K252c和streptochlorin。此外，还从菌株Streptomyces sp. W444基因组中定位了星孢菌素生物合成基因簇，并对其生物合成途径进行了推导。结论 泉州湾红树林根际土壤中的放线菌具有多样性，并蕴含着丰富的潜在天然产物资源，从菌株Streptomyces sp. W444中分离鉴定了吲哚咔唑类生物碱staurosporine、K252c和streptochlorin，这为研究泉州湾红树林可培养放线菌的多样性和次级代谢产物研究提供了坚实的基础。

Abstract:

Objective To isolate and identify actinomycetes from two mangrove soil samples in Quanzhou Bay as well as secondary metabolites from the target strain Streptomyces sp. W444 with antifungal activity.Methods The rhizosphere soil samples of two different mangrove plants were collected from Luoyang River in Quanzhou Bay. Actinomycetes were isolated from the soil samples by the dilution plating method. The isolates were classified by the phylogenetic tree constructed based on 16S rRNA gene sequences. The antifungal activities of the isolates were examined by the agar diffusion method. The target strain Streptomyces sp. W444 was subjected to fermentation scale-up for the isolation of secondary metabolites. Furthermore, the biosynthetic gene clusters were analyzed to deduce the biosynthetic pathway of staurosporine.Results A total of 56 strains of actinomycetes were isolated from mangrove soil samples and categorized into 8 genera belonging to 6 families of 6 orders. Among them, Streptomyces and Micromonospora were dominant, with the relative abundance of 41.0% and 33.9%, respectively. Strain Streptomyces sp. W444 exhibited excellent antifungal activity, and three indoles (staurosporine, K252c, and streptochlorin) were isolated and identified from this strain. The biosynthetic gene cluster of staurosporine was localized in the genome of Streptomyces sp. W444 by bioinformatics analysis. The biosynthetic pathway of staurosporine was then proposed.Conclusion Actinomycetes in the rhizosphere soil of mangrove plants in Quanzhou Bay had high diversity and contained potential natural product resources. Staurosporine, K252c, and streptochlorin were isolated from strain W444. These findings lay a solid foundation for studying the diversity and secondary metabolites of cultivable actinomycetes in mangrove forests in Quanzhou Bay.