基于16 S rRNA序列研究华重楼植株可培养内生细菌的多样性

Cultivable bacterial diversity of Paris polyphylla var. chinensis with 16 S rRNA sequence analysis

目的分析华重楼Paris polyphylla健株和茎腐病感病植株内生可培养内生细菌的多样性。方法用牛肉膏蛋白胨培养基分离华重楼茎腐病及健康植株根茎、茎、叶等不同部位的细菌,采用16 S rRNA通用引物27F/1492R进行PCR扩增结合DNA测序技术,对分离自华重楼健株及茎腐病植株不同部位的细菌进行初步鉴定。结果从华重楼的健康植株和感病植株中分离到11属23种细菌。从健株分离到4属11种内生细菌,其中根茎、茎、叶分别分离到9、10和5种内生细菌。从病株分离得到10属14种内生细菌,其中根茎、茎、叶分别分离到11、8和3种内生细菌。华重楼健株的根茎部内生细菌量最高,达2.999×105 cfu/g,叶部内生细菌量较低,为7.32×104 cfu/g,华重楼健株根茎、茎、叶中的芽孢杆菌量最高,比例分别为73.3%、67.1%和81.8%。华重楼病株的茎部内生细菌量最高,达2.817×105 cfu/g,叶部内生细菌量较低,为2.76×104 cfu/g,华重楼病株的根茎、茎、叶部的假单胞菌量最高,比例分别为35.6%、50.3%和60.5%。华重楼健株不同部位的多样性指数和均匀度指数均高于病株。结论华重楼健康植株以芽孢杆菌为优势细菌类群,华重楼茎腐病植株以假单胞菌为优势细菌类群。华重楼健株相比病株具有更为丰富的内生细菌群落多样性。

Objective To analyze the bacterial diversity in stem rot and healthy plants of Paris polyphylla var. chinensis. MethodsBacterial strains were isolated from rhizomes, stems and leaves of the diseased and healthy plants of P. polyphylla var. chinensis using beef extract-peptone medium. Using 16 S r RNA universal primers 27F/1492 R for PCR amplification, combined with DNA sequencing technology to preliminary identification the bacterial strains. Results The bacteria could be divided into 23 microbial species, belonging to 11 genera, isolated from healthy and diseased plants of P. polyphylla var. chinensis. Eleven endophytic bacteria, belonging to 4 genera were contained from healthy plants, in which there were 9, 10, and 5 species isolated from rhizomes, stems and leaves, respectively. Fourteen endophytic bacteria, belonging to 10 genera were contained from diseased plant, in which there were 11, 8, and 3 species isolated from rhizomes, stems, and leaves, respectively. The content of endophytic bacteria in rhizome of healthy plant was the highest, reached up to 2.999 × 10^5 cfu/g, while that in leaf was the lowest with 7.32 × 10^4 cfu/g. The quantities of Bacillus species in rhizome（73.3%）, stem（67.1%）, and leaf（81.8%） of healthy plant were the highest groups, Pseudomonas species in rhizome（35.6%）, stem（50.3%） and leaf（60.5%） of diseased plants were the highest groups. Shannon-Wiener index and evenness index of healthy plant of P. polyphylla var. chinensis were higher than that of diseased plants. Conclusion The dominant group in the healthy plant of P. polyphylla var. chinensis was Bacillus, while the dominant groups in the diseased plant of P. polyphylla var. chinensis was Pseudomonas. The population diversity of the cultivable bacteria in healthy plant of P. polyphylla var. chinensis was more abundant than that of diseased plant.