【摘要】 采用组织PCR-末端限制性片段长度多态性分析(T-RFLP)免培养技术,从南药植物高良姜道地产地植株根、根茎、茎、叶4个不同组织总共检测到28个不同的末端限制性片段(T-RFs),说明高良姜内生真菌群体至少组成28个不同的真菌种;不同组织其T-RFLP图谱、T-RFs数目、优势T-RFs组成不同,内生真菌群体Shannon多样性指数和Shannon均匀度指数也随组织而异;采用水蒸气蒸馏法、甲醇抽提-高效液相色谱法测定高良姜主要活性成分挥发油、高良姜素含量,结果显示,该2类活性组分在其根、根茎、茎、叶4个不同组织均有积累,且均以根茎最高;相关性分析显示2类所测活性成分与该宿主内生真菌群体Shannon多样性指数及其均匀度呈极显著的负相关,与325 bp对应于拟盘多毛孢菌属Pestalotiopsis的优势T-RFs呈极显著的正相关。更多还原

【Abstract】 In the present study,terminal-restriction fragment length polymorphism( T-RFLP) technique was applied to assess the diversity and tissue distribution of the fungal endophyte communities of Alpinia officinarum collected from Longtang town in Xuwen county,Guangdong province,China,at which the pharmacological effect of the medicine plant is traditional considered to be the significantly higher than that in any other growth areas in China. A total of 28 distinct Terminal-Restriction Fragment( T-RFs) were detected with HhaI Mono-digestion targeted amplified fungal nuclear ribosomal internal transcribed spacer region sequences( rDNA ITS) from the root,rhizome,stem,and leaf internal tissues of A. officinarum plant,indicating that at least 28 distinct fungal species were able to colonize the internal tissue of the host plant. The rDNA ITS-T-RFLP profiles obtained from different tissues of the host plant were obvious distinct. And the numbers of total T-RFs,and the dominant T-RFs detected from various tissues were significantly different. Based on the obtained T-RFLP profiles,Shannon’s diversity index and the Shannon’s evenness index were calculated,which were significantly different among tissues( P < 0. 05). Furthermore,two types of active chemicals,total volatile oils by water vapor distillation method and galangin by methanol extraction-HPLC method,were examined in the each tissue of the tested plant. Both of tested components were detected in all of the four tissues of the medicine plant with varying contents. And the highest was in rhizome tissue. Correlation analysis revealed there were significant negative correlations between both of the tested active components contents and calculated Shannon’s diversity index,as well as the Shannon’s evenness index of the fungal endophyte communities of the host plant( P = 0,Pearson correlation coefficient≤- 0. 962),and significant positive correlations between both of the tested active components contents and 325 bp dominant T-RF linkage to Pestalotiopsis( P = 0,Pearson correlation coefficient ≥ 0. 975). In conclusion,A. officinarum is colonized by diverse fungal endophytes communities. The diversity of the fungal endophytes was found in the A. officinarum varied with differences of the tissue types of the host plants and was closely correlated with the accumulation of main active components,total volatile oils and galangin contents in the host plant tissue.更多还原