2种基质种植下石斛根系内生菌多样性分析

Diversity analysis of endophytic microbes in roots of Dendrobium nobile(Lindl.)(Orchidaceae)on two substrates

【目的】阐明不同基质种植下石斛根系内生菌菌群的差异性,为进一步研究内生菌组成差异对石斛药效成分积累的影响、优化种植条件提供参考依据。【方法】选取仿野生贴石种植于丹霞石块和火山石块的2年生石斛根系,进行16S rRNA V4区域和ITS 2区域的高通量测序,分析2种基质种植下石斛根系内生细菌和内生真菌的次生代谢产物、群落组成结构、alpha多样性、beta多样性和功能预测。【结果】火山石与丹霞石的石斛根系次生代谢产物中倍半萜糖苷类物质Dendromoniliside C or isomer(288-p)和Dendromoniliside D or isomer(123-n)差异显著(P<0.05,下同)。群落结构分析显示,内生细菌和内生真菌的组成在门水平和属水平上均无明显差异,其中,内生细菌在门水平上均以变形菌门(Proteobacteria)相对丰度最为富集,在属水平上均以鞘脂单胞菌属(Sphingomonas)相对丰度最为富集;内生真菌在门水平上均以子囊菌门(Ascomycota)为绝对优势门,在属水平上均以暗球腔菌属(Phaeosphaeria)为优势菌群。主成分分析反映火山石与丹霞石样本组间差异明显,基于ANOSIM组间差异检验下的Unweighted UniFrac指数、Bray Curtis指数和Jaccard指数均进一步表明不同基质种植下石斛内生菌的组间差异远大于组内差异。功能预测结果显示,在2种基质下石斛根系中,内生菌真菌营养型以病理腐生营养型(Pathotroph-Saprotroph)和腐生营养型(Saprotroph)差异最显著,在主要功能通路中真菌寄生—植物病原—植物腐生菌功能群(Fungal Parasite-Plant PathogenPlant Saprotroph)和未定义腐生菌功能群(Undefined Saprotroph)差异显著,均为丹霞石显著高于火山石。【结论】2种基质种植环境显著影响石斛根系内生菌群的次级代谢产物、菌群相对丰度、优势菌群和群落功能。同时也提示,石斛根系内生真菌相较于内生细菌,对不同种植基质条件更敏感。

【Objective】This study was conducted to clarify the diversity of endophytic flora in roots of Dendrobium nobile(Lindl.)(Orchidaceae)cultivated on two different substrates,so as to provide reference for investigating the influence of differences in endophytic composition on the accumulation of effective medicinal components of D.nobile,as well as optimizing planting conditions.【Method】Two-year D.nobile root samples planted on Danxia stone and volcanic stone substrates were selected for high-throughput sequencing of the V4 region and ITS2 region of 16S rRNA.The secondary metabolites,microbial composition,alpha diversity,beta diversity,and function prediction analysis of endophytic bacteria and endophytic fungi in D.nobile roots were performed.【Result】Dendromoniliside C or isomer(288-p)and Dendromoniliside D or isomer(123-n),two representative sesquiterpene glucosides,were significantly different in roots of D.nobile cultivated at Danxia stone and volcanic stone(P<0.05,the same below).Microbial composition analysis showed no significant difference at phylum and genus levels between the two substrate groups.Proteobacteria was the most abundant bacteria at phylum level and Sphingomonas was the most abundant bacteria at genus level.Ascomycota was the absolutely dominant fungal phylum,Phaeosphaeria was the predominant fungal genus.Principal component analysis reflected great differences between volcanic stone and Danxia stone,and the inter-group differences were larger than the intra-group differences based on Unweighted UniFrac index,Bray Curtis index and Jaccard index in ANOSIM test.Furthermore,according to the results of fungal function prediction,in the D.nobile roots of volcanic stone and Danxia stone,the difference of Pathotroph-Saprotroph and Saprotroph among endophytic fungi was the most significant,and there was a significant difference between Fungal Parasite-Plant Pathogen-Plant saprotroph and Undefined Saprotroph.Both were significantly higher in Danxia stone than volcanic stone.【Conclusion】The two substrate planting environments significantly affect the secondary metabolites,relative abundance,dominant microbial community,and community function of the endophytic microbiota in D.nobile roots.At the same time,it also suggests that endophytic fungi in the root system of D.nobile are more sensitive to different planting substrate conditions compared to endophytic bacteria.