蜘蛛抱蛋属植物内生细菌的多样性及其抗菌活性筛选

【背景】药用植物中蕴含多样性丰富的内生菌资源,这些微生物产生的多种新型物质在制药领域表现出较好的应用前景。【目的】研究蜘蛛抱蛋属(Aspidistra Ker-Gawl.)植物内生细菌的多样性,探索药用植物内生细菌在药用活性产物方面的开发潜力,以期发现具有抗菌活性的次级代谢产物。【方法】对9种13株新鲜的蜘蛛抱蛋植物进行表面消毒,采用5种分离培养基分离内生细菌;根据菌落形态特征排除重复菌株,并测定其16S rRNA基因序列,构建系统进化树分析内生细菌多样性;将菌株分别用2种培养基发酵,使用耻垢分枝杆菌(Mycobacterium smegmatis ATCC 700044)、水稻白叶枯菌(Xanthomonas oryzae PXO99A)、白色念珠菌(Candidaalbicans ATCC 10231)、肺炎雷伯菌(Klebsiella pneumoniae ATCC 700603)和耐药粪肠球菌HH22(Enterococcus faecalis HH22)5种检定菌对分离菌株的发酵液进行抑菌活性筛选。【结果】从植物组织中分离得到了234株内生细菌,根据形态初步排重得到156株植物内生细菌;基于16S rRNA基因序列构建的系统进化树显示它们分属于3门10目22科29属,其中链霉菌属(Streptomyces)、芽孢杆菌属(Bacillus)、微杆菌属(Microbacterium)、类芽孢杆菌属(Paenibacillus)和根瘤菌属(Rhizobium)的菌株广泛地分布在不同种的蜘蛛抱蛋植株中,且占据一定优势;发现可能的潜在新分类单元6个;156株内生细菌中38株菌的发酵液具有抑菌活性,初筛阳性率为23.7%。【结论】蜘蛛抱蛋植物组织中含有种类多样的内生细菌,它们可能是抗菌生物活性次级代谢产物的有效来源。

Endophytic bacteria diversity of Aspidistra Ker-Gawl. and their antimicrobial activities

Background] Medicinal plants constitute the huge diversity of endophytic bacteria. These microbes have potential to synthesis of numerous novel compounds that can be exploited in pharmaceutical industries. Objective] For the purpose of exploring the pharmaceutically potential of the endophytes in medicinal plants and finding antimicrobial secondary metabolites, endophytic bacteria diversity of Aspidistra Ker-Gawl. was investigated. Methods] After surface sterilization, the tissue fragments of 13 plants, which were classified into 9 different species, were inoculated onto 5 diverse isolation media. Duplicate isolates were eliminated by colony properties. Based on 16S rRNA gene sequencing results, Neighbour-Joining phylogenetic tree was constructed to analyze the endophytic bacteria diversity in Aspidistra spp. All the non-duplicated isolates were fermented with two kinds of media. The antimicrobial activities of the endophytic motablisms were detected with 5 testing strains, including Mycobacterium smegmatis ATCC 700044, Xanthomonas oryzae PXO99A, Candida albicans ATCC 10231, Klebsiella pneumoniae ATCC 700603 and resistant strain Enterococcus faecalis HH22. Results] A total of 234 endophytic bacteria were isolated from different plant tissues and 156 non-duplicated strains were confirmed after colony properties checking. These endophytic bacteria strains were classified into 3 phyla, 10 orders, 22 families and 29 genera. Notably, Streptomyces sp., Bacillus sp., Microbacterium sp., Paenibacillus sp. and Rhizobium sp. were the predominant species in Aspidistra spp., and 6 potential novel species were found. The fermentation broth from 38 isolates (23.7%) showed antimicrobial activities. Conclusion] The data of this study showed an unexpected diversity of endophytes inside the Aspidistra tissues, and suggested that endophytic bacteria of the Aspidistra tissues are promising sources of new natural secondary metabolites with favorable antimicrobial activities.