红海榄内生细菌多样性及其抑制鱼类致病菌活性研究

Diversity of cultivated marine bacteria and antibacterial activity of endophytic bacterial in Rhizophora stylosa

关键词:  红海榄, 内生细菌, 物种多样性, 副溶血弧菌, 抑菌活性

Key words:   Rhizophora stylosa, endophytic bacterial, diversity, Vibrio parahaemolyticus, antibacterial activity

摘要:

红海榄为红树科红海属植物,广泛分布于热带海岸的红树林中,为真红树植物的典型代表。该研究以广西山口红树林国家级自然保护区红海榄为对象,应用稀释涂布法和三线法从红海榄各组织中分离出17株内生细菌,通过各菌株形态特征观察及利用16S rRNA序列基因和韦恩图分析其内生细菌多样性。结果表明:17株内生细菌分属3个门5科7属8种,Micromonospora和Mangrovibacter属为其优势类群。进一步分析发现红海榄根茎叶等组织的内生细菌类群差别较大,仅有1个相同菌属,17株菌中有3株菌与已有细菌物种典型菌株的全长16S rRNA基因相似性低于97%,代表着潜在的新属或新种。运用纸片法研究8株内生细菌发酵液的乙酸乙酯提取物对鱼类致病菌副溶血弧菌活性抑制效果,发现3株内生细菌(H003、H013、H009,浓度5 mg·mL^-1)的代谢产物对副溶血弧菌具有较强的抑菌活性,其抑菌圈直径分别达到(8.4±0.07)、(8.2±0.07)、(8.3±0.14)mm。该研究结果表明红海榄中具有较好的内生细菌多样性和抑菌活性,为今后研究其内生细菌的化学多样性及其应用提供了重要的物质基础。

Abstract:

Rhizophora stylosa is an evergreen tree with prop roots, and a common tree found in tropical coastal mangrove forests, widely distributed in tropical coastal mudflats. As a typical representative of the mangrove, R. stylosa has rich endophytic diversity with little research. The study objects of R. stylosa were collected from Shankou National Nature Reserve, Guangxi, China. From stem, root, leaf and other tissues, seventeen endophytes were isolated with the dilution coating method and the three-wire method. We Observed their morphological characteristics, 16S rRNA and Venny Chart were utilized to analyze the diversity of their endogenous bacterial. Depending on the results of diversity analysis, we could find that these strains have different morphological characteristics and they could classified into three doors, seven genera, five families and eight species. Further analysis showed that endophytic bacteria groups of R.stylosa root, stem, leaf and other organizations vary different, only one same genus was Mangrovibacter plantisponsor. Micromonospora and Mangrovibacter genus were its dominant bacteria, the proportion was 29.4% and 17.6%, respectively. Among them, the most endophytic bacteria within the root, the proportion was 47.1%. Seventeen strains belong to three doors, wherein the most number of bacteria were Proteobacteria, accounting for 52.9%, then Actinomycetes door accounted for 41.2%. In those isolated strains, three strains of bacterial species with the full-length 16S rRNA gene similarity less than 97%, may be possible new genera or new genus and had potential research value. Vibrio parahaemolyticus was a zoonotic bacteria caused some damage to uman health, severe cases even death. To test anti Vibrio parahaemolyticus activity of the crude extract of strains isolated, we used filter paper method, and the results indicated that three strains(H003, H009, H013, 5 mg·mL^-1)had a strong antibacterial activity against Vibrio parahaemolyticus, their inhibition zone could reach(8.4±0.07),(8.2±0.07), and(8.3±0.14)mm, respectively. Endophytic bacteria of Rhizophora stylosa were genetically diverse and most of them showed strong inhibition effect against Vibrio parahaemolyticus. The results provides the essential material basis for the further study of its chemical diversity and utilization of resources.