| Industry Chinese herbal medicine is a strategic emerging industry in China and a vital part of the Chinese medicine and the general health industry.The microbiome is the crucial determinant of the health and productivity of medicinal plants and can directly affect the metabolome of its host,and further thereby affect the quality of medicinal materials.Eucommia ulmoides is a kind of medicinal and edible plant rich in biologically active compounds.It is the main source of E.ulmoides rubber in industrial production and has high medicinal and economic value.However,most of the current researches have focused its genetic breeding,clinical medicine and industrial application,and lacked of the basic understanding of the ecological processes such as the composition of microbiome,the function of specific taxa and community construction.Eighty-one samples(bark,rhizosphere soil,and leaf)from 27 E.ulmoides regions in China were used as research materials.According to high-throughput sequencing technology,gas chromatograph-quadrupole time of flight mass spectrometry(GC-QTOF-MS),chloroplast DNA molecular markers and other methods and means,the study will 1)systematic assessment of the strengths and limitations of the four core approaches to microbiome definition: membership,composition,phylogenetic and functional redundancy,and network connection;2)to reveal the composition and distribution characteristics of microbiome with E.ulmoides bark;3)elucidate the composition,function of core and rare taxa and potential relationship with pharmacologically active components;4)to analyze the effects of bark microbiome,host genetic characteristics,climate and soil physicochemical factors on the pharmacologically active components of E.ulmoides,4)to clarify the relative importance of deterministic and stochastic processes in the construction of bark microbiome,and explore the effects of host genotype,rhizosphere microbiome,climatic factors,soil physicochemical and medicinal active ingredients on microbial community construction.The main results are as follows:(1)Although the core microbiomes defined by different methods have preferences in composition and function,the term refers to a group of microbes that are particularly notable and important for host-associated microbiomes.In terms of the number of compositions,the four definition methods of phylogeny,membership,composition,and network connection contained 274,10,5,and 5 core OTUs.From the perspective of species abundance,phylogeny and network connectivity methods are more general.In contrast,the core microbiomes defined by different methods displayed similarities in function.Therefore,we propose:(I)The definition method of the core microbiome should be selected according to the ecological problems faced;(II)A combination of multiple methods may comprehensively reveal the core microbiome at different levels of the host,and may also facilitate understanding of the ecological and evolutionary processes that govern host-microbe interactions.(2)The bark of E.ulmoides was rich in microbiota,among which fungal community was divided into 12 phyla,39 classes,103 orders,239 families,469 genera and 1850 OTUs.The dominant fungal genus were Sordariomycetes spp.(12.82%).Bacterial community was divided into 36 phyla,77 classes,160 orders,319 families,614 genera and 1884 OTUs.The dominant bacterial genera were Bacillus(12.27%).In general,there was no significant difference in the alpha diversity of microbial communities from different regions,but there was significant difference in the beta diversity.Further analysis revealed that the core microbiota in bark contained11 taxa.Among them,five fungal taxa were Helminthosporium,an unidentified genus of Pleosporales,Fusicolla,Acremonium,and Paraconiothyrium;six bacterial taxa were Nocardioides,Mycobacterium,an unidentified genus of Comamonadaceae,an unidentified genus of Microbacteriaceae,Solirubrobacter,Acetobacter,and an unidentified genus of Acidimicrobiaceae.The rare microbiota of bark contained 599 taxa.Among them,rare fungal microbiota included 281 taxa,such as Lophotrichus,Byssoloma,Distoseptispora,Beauveria,Retiarius;rare bacteria microbiota included318 taxa,such as Actinomyces,an unidentified genus of Lachnospiraceae,Rheinheimera,Saccharopolyspora,Pannonibacter.(3)The core and rare taxa are closely related to the structural stability and network complexity of microbial communities,host fitness,and accumulation of pharmacologically active components.The ecological functions of core taxa involved were mainly endophyte,fungal parasite,plant pathogen,and wood saprotroph,and the phenotypic functions are mainly organotrophic,chemoheterotrophic,mesophilic and thermophilic microorganisms,and the metabolic functions are mainly chemoheterotrophy and aromatic compound degradation.They were also significantly correlated with the content of pharmacologically active ingredients,for example,Pleosporales spp.,Acremonium spp.and Paraconiothyrium spp.were all positively correlated with pinoresinol diglucoside,aucubin and geniposide acid,and Mycobacterium spp.and Microbacteriaceae spp.were positively correlated with the three pharmacologically active components.The ecological functions of rare fungal taxa were mainly wood saprotroph,plant pathogen,animal pathogen,lichenized fungi,and endophyte;rare bacterial taxa were heterotrophic,autotrophic,methylotrophic,organotrophic mesophilic,psychrophilic and thermophilic microorganisms,and their metabolic functions were mainly chemoheterotrophic,fermentation,urea decomposition,and methanol oxidation reactions,methyl nutrition.In addition,our study also found that the core and rare microbiota are key taxa to maintain the network structure and stability of the microbial community,and the network structure of the microbes changed significantly when the core and rare taxa were removed.(4)The genetic diversity analysis showed that there were 12 different haplotypes in all samples of E.ulmoides,and the genetic heterogeneity of population was not significant.The contents of pharmacologically active ingredients in the bark of E.ulmoides varied with the producing regions,the content of pinoresinol diglucosid was14.76-40.07 μg/m L,geniposidic acid was 4.88-33.2 μg/m L,and aucubin was13.99-44.5 μg/m L.Further analysis found that the accumulation of pharmacologically active ingredients of E.ulmoides were dramatically affected by genotype,environmental factors(biological and abiotic factors),and their interaction,and genotype had the greatest impact.Among them,the three pharmacologically active ingredients were significantly correlated with the Lactobacillus,Escherichia,Bacteroides and Faecalibacterium,suggesting that the active ingredients were mainly affected by bark microorganisms.(5)The assembly of microbiome was affected by both stochastic and deterministic factors(β-NTI value > 2,then the deterministic process was dominant;β-NTI value < 2,then the stochastic process was dominant).Further analysis showed that relative humidity,annual mean temperature,available nitrogen,p H and geniposide acid were the main influential factors for the construction of bark microbial community.Fast expectation-maximization microbial source tracking(FEAST)suggested that the rhizosphere soil of E.ulmoides was one of the main sources of microbiota in bark.This study systematically revealed the taxonomic composition,function,construction mechanism of microbiome and potential relationship with pharmacologically active components of bark with E.ulmoides.These results provide comprehensive experimental evidence for artificially synthesizing communities to regulate the assembly composition of microbiome in E.ulmoides,and provide important information for future sustainable agricultural management based on precise microbiome regulation. |
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