| Orchid mycorrhizal fungi colonies the root cortex cells of orchids and provide nutrients for orchid seed germination and plant growth and development.The mechanisms of promotion and colonization of orchid mycorrhizal fungi are still unclear,thus the study of the symbiosis between orchid mycorrhizal fungi and plants is of great significance for the conservation of rare orchids and the large-scale cultivation of medicinal orchids.In this study,we used Dendrobium officinale as the experimental material,isolated orchid mycorrhizal fungi,analyzed the effect of inoculation of mycorrhizal fungi on the growth of Dendrobium officinale,and further investigate the mechanism of colonization by mycorrhizal fungi.The experimental results were as follows:(1)Dendrobium officinale roots were used as the experimental material for endophytic fungal isolation,Preliminary classification according to morphological characteristics,pigment-producing capacity.A total of 18 fungal strains were obtained.Molecular identification was performed using ITS sequences.The identification of the fungi was combined with sequencing results and morphological characteristics to obtain 16 species of fungi.respectively Trichoderma sp.,Diaporthe sp.,Penicillium oxalicum,Talaromyces ruber,Talaromyces amestolkiae,Fusarium sp.,Paraboeremia sp.,Trichoderma sp.,Sordariomycetes sp.,Trichoderma asperellum,Fusarium solani,Chaetomium sp.,Chaetomium convolutum,Tulasnella sp.and Fusarium oxysporum.The dominant genera were Fusarium and Trichoderma,which account for more than 30% of the total.The obtained fungi were inoculated into plant,and strains E(Penicillium oxalicum),T1-1(Fusarium),YY-1(Fusarium solani),YG-4(Fusarium oxysporum)and YG-5(Tulasnella sp.)could form intracellular mycelial cluster structures.Strain YG-5 promoted the growth of Dendrobium officinale.Under isolated conditions,strain YG-5secretes pectinase and cellulase,but not laccase and xylanase,and has no phosphate solubilization function for organic and inorganic phosphorus.(2)The orchid mycorrhizal fungus YG-5 was inoculated into sterile Dendrobium officinale seedlings to analyze the effects of orchid mycorrhizal fungi colonization on plant growth and biochemical indexes.Inoculation with the mycorrhizal fungus YG-5 increased the dry weight,fresh weight,plant height,leaf number,stem thickness,chlorophyll b and total chlorophyll content of Dendrobium officinale,and the contents of flavonoids,free amino acids,soluble sugars,soluble proteins and malondialdehyde in tissues;in addition,it also increased acid phosphatase and glutamine synthetase activities.Inoculation with mycorrhizal fungi treatment failed to increase the polysaccharide and proline contents and the activities of peroxidase,polyphenol oxidase,acid phosphatase,glutamate dehydrogenase,and glutamate synthase compared to the control group.Overall,inoculation with mycorrhizal fungi promoted the growth and development of Dendrobium officinale and increased the content of some metabolic substances in the tissues.(3)Transcriptome sequencing results showed that after inoculation with mycorrhizal fungi,the number of up-regulated expressed genes was 508 and the number of down-regulated expressed genes was 64;Differential genes are mainly up-regulated in expression.GO enrichment analysis showed that differentially expressed genes were associated with response to stimulus,response to chitin,flavonoid metabolic process,response to organonitrogen compound,acid phosphatase activity,transcription regulator activity,DNA-binding transcription factor activity and calcium ion binding.KEGG enrichment analysis showed that the highest enrichment was in linoleic acid metabolism;the highest enrichment was in transcription factors,followed by plant-pathogen interaction,MAPK signaling pathway-plant.Combined with the transcriptome sequencing results and biological index measurements,it was inferred that inoculation with mycorrhizal fungus YG-5 improved the phototransduction ability and enhanced photosynthesis of plants;promoted the accumulation of flavonoids and other substances,improved the medicinal value of Dendrobium officinale;enhanced the utilization of N and P elements by plants.Inoculation of mycorrhizal fungi enhances the expression of calcium-binding genes and promotes cellular signal transduction,which is involved in carbon and nitrogen metabolism,cytoskeleton production and defense response mechanisms in plants.Mycorrhizal fungi are involved in the MAPK signaling pathway,which activates the expression of resistance genes in plants and enhances their resistance to stress;After invasion,mycorrhizal fungi interact with plants in a complex way,causing a large number of transcription factors to bind to RNA polymerase and promote the transcription of some genes,thus forming a complex plant-mycorrhizal fungal gene interaction network. |
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