基于高通量测序的明日叶不同组织内生菌群落结构分析

Community structure analysis of the endophyte in different tissues of Angelica keiskei(Miq.)Koidz. based on high-throughput sequencing

  • 摘要: 【目的】分析明日叶不同组织内生菌的群落结构和多样性,为探究明日叶和内生菌互作机制及功能菌株筛选提供参考。【方法】以四川宜宾栽培的药食兼用型明日叶为试材,采用Illumina MiSeq高通量测序技术确定根、茎、叶和果实组织内生真菌和内生细菌群落多样性。【结果】根据97%的序列相似性水平,将内生真菌和细菌的reads分别归类为1023和1871个操作分类单元(OTU)。韦恩图和热图显示明日叶各组织内生真菌和内生细菌群落结构差异性较高,种群丰度存在一定的差异。α多样性分析发现明日叶的叶内生真菌种类数最多,根内生真菌群落多样性最低,果实内生真菌群落丰富度最低;而根含有内生细菌种类数最多,茎内生细菌群落多样性最低,果实内生细菌群落丰富度最低。β多样性分析表明,组织特征对明日叶内生真菌或内生细菌群落结构有显著影响。明日叶4个不同组织中的优势真菌门和细菌门分别为子囊菌门(Ascomycota)和变形菌门(Proteobacteria)。根、茎、叶和果实中内生真菌优势属分别为SerendipitaParamycosphaerella、多臂菌属(Trichomerium)和Apiotrichum,内生细菌优势属分别为罗尔斯通菌属(Ralstonia)、罗尔斯通菌属、甲基杆菌属(Methylobacterium)和弧菌属(Vibrio)。线性判别分析(LEfSe)表明,除根外,明日叶的茎、叶和果实中内生真菌重要差异指示种分别是Paramycosphaerella、苍白尾孢属(Pallidocercospora)和横断孢属(Strelitziana);茎、叶和果实中内生细菌重要差异指示种分别是罗尔斯通菌属、甲基杆菌属和亮杆菌属(Leucobacter)。经FUNGuild和COG功能预测发现,不同组织内生菌群功能丰富,除具有固氮、促生和抑菌等功能的菌群外,还存在较大比例未知功能菌群。【结论】明日叶中蕴含丰富多样的内生菌群,在不同组织中的群落结构、多样性、优势菌属及差异指示种存在保守性与差异性,组织特征差异是影响内生菌群的重要因素;不同组织内生菌群生态功能亦差异较大,且大量为未鉴定出种属和未定义的功能菌群。

     

    Abstract: 【Objective】The community structure and diversity of endophyte in different tissues of Angelica keiskei (Miq.)Koidz. was analyzed,to provide a theoretical basis to explore the interaction mechanism and functional strain screening of A. keiskei and endophyte.【Method】 The medicinal and edible plant A. keiskei,which was cultivated in Yibin, Sichuan,was taken as the object. The diversity of endophytic fungi and bacteria in the root,stem,leave and fruit tissues was examined by high-throughput sequencing using Illumina MiSeq.【Result】According to the 97% sequence similarity, endophytic fungal and endophytic bacterial reads could be indentified into 1023 and 1871 operational taxonomic units (OTU),respectively. A Venn diagram and a heatmap showed that there were fairly high community structure difference between endophytic fungi or endophytic bacteria in the four different parts of tissues of A. keiskei,and the abundances of populations differed. According to α diversity analysis,the variety of endophytic fungi in the leaves of A. keiskei was the most,the diversity of endophytic fungal community in the roots was the lowest,and the abundance of endophytic fungal community in the fruits was the lowest. The variety of endophytic bacteria in the roots of A. keiskei was the most,the diversity of endophytic bacterial community in the stems was the lowest,and the abundance of endophytic bacterial community in the fruits was the lowest. β diversity analysis showed that the characteristics of tissues had a significant effect on the community structure of the endophytic fungi or endophytic bacteria in A. keiskei. Ascomycota and Proteobacteria were the dominant fungal phylum and bacterial phylum in the four different parts of tissues of A. keiskei respectively. The dominant genera of endophytic fungi in the roots,stems,leaves and fruits were SerendipitaParamycosphaerellaTrichomerium and Apiotrichum respectively,and the dominant genera of endophytic bacteria were RalstoniaMethylobacterium and Vibrio respectively. Linear discriminant analysis showed that,the most important biomarkers of the endophytic fungi in the stems,leaves and fruits were ParamycosphaerellaPallidocercospora and Strelitziana respectively. The most important biomarkers of endophytic bacteria in stems,leaves and fruits were RalstoniaMethylobacterium and Leucobacter respectively. FUNGuild and COG functional predictions showed that the function of endophyte in different tissues was abundant,and there was a large proportion of unknown functional endophyte in addition to those that could fix nitrogen, promote growth and inhibit bacteria.【Conclusion】 There are abundant and diverse endophytes in A. keiskei. The community structure,diversity,dominant genera and biomarkers in different tissues are conservative and different,and the difference of tissue characteristics is an important factor affecting the endophyte. The ecological functions of endophyte in different tissues are also quite different,and a large number of species and functional endophyte are not identified.

     

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