Effect And Mechanism Of Mycorrhizal Fungi On Improving Drought Resistance Of Coelogyne Viscosa(Epiphytic Orchid)

Orchids are one of the largest families of flowering plants and have various life forms.About 70%of orchids are epiphytic orchids,which are the main life forms of orchids in tropical and subtropical regions.Compared with terrestrial orchids,epiphytic orchids face more habitat stress,among which drought is one of the main stress,and seasonal drought makes epiphytic orchids own strong drought resistance.As an important symbiotic partner of orchids,mycorrhizal fungi(OMF)have been considered to play an important role in the ecological adaptation of orchids.In this paper,Coelogyne viscosa,an epiphytic orchid,is used as an experimental plant.On the basis of isolated mycorrhizal fungi,the effect of OMF on the growth and drought resistance under drought stress was clarified,and the main results are as follows:1.Seven strains of endophytic fungi were isolated from the roots of C.viscosa by single peloton separation and tissue mass separation methods.According to morphological and molecular identification,two strains of Tulasnellaceae(hy-111,hy-1),one strain of DSE belonging to Exophiala(HB3-1),one strain of Flammulina(HBS1)and three strains of ascomycetes(cvl-1,C-11,C-22)were identified.2.After the disinfection,sowing and aseptic culture of the seeds of C.viscosa,the sterile seedling culture system of C.viscosa was established.The isolated endophytes were successfully reinoculated to the roots of the sterile seedlings by exploring the symbiotic conditions such as the components of substrate and water content.Mycorrhizal seedlings were obtained after 100 days of culture.3.The fresh weight of seedlings in each group after 100 days of symbiotic culture with 7 strains of fungi was measured,and the change of fresh weight of seedlings compared with before inoculation was calculated.The results showed that DSE HB3-1 significantly promoted the growth of seedlings(P<0.01).And hy-111 and HBS1 showed marginal significance in promoting seedling growth(P=0.065,P=0.068).The contents of POD,SOD,CAT,soluble protein,soluble sugar and proline in the leaves of seedlings on the days of 0,3,6,9 and 12 under drought stress were measured.The results showed that seedlings inoculated with hy-111 and DSE HB3-1 showed significantly higher levels of antioxidant enzymes(POD,SOD,CAT),osmotic regulators(soluble protein and soluble sugar),and lower proline level than the control group.Combined with the change of fresh weight,it is preliminarily assumed that these two fungi could enhance the drought resistance of C.viscosa seedlings.4.The leaves of three groups of seedlings inoculated with hy-111,HB3-1 and noninoculated with fungi(control)were collected on day 6 of drought stress for RNA-Seq and statistically analyzation.GO functional enrichment analysis showed that the DEGs of group hy-111 were significantly enriched in "carbohydrate metabolism","oxidation-reduction process" and "oxidation-reduction activity".The DEGs of group HB3-1 were mainly enriched in "cell polysaccharide metabolism","translation" and "oxidation-reduction process".KEGG pathway enrichment analysis showed that the up-regulated DEGs of group hy-111 were mainly enriched in "plant hormone signal transduction","glycerophospholipid metabolism" and "phenylpropanoid metabolism".In addition,it was found that the"glycosphingolipid biosynthesis-globo and isoglobo series" and "photosynthesis-antenna protein" pathway was up-regulated.The down-regulated DEGs were mainly enriched in pathways such as "carbon metabolism","ribosome",and "amino acid biosynthesis".The up-regulated DEGs of group HB3-1 were significantly enriched in "plant-pathogen interaction","plant hormone signal transduction" and "MAPK signaling pathway-plant".In addition,"glycosphingolipid biosynthesis","carotenoid biosynthesis","ascorbic acid and aldehyde acid metabolism" were also found in up-regulated pathways.Down-regulated DEGs were mainly enriched in "ribosome","carbon metabolism","oxidative phosphorylation".These pathways are closely related to stress resistance,suggesting that both hy-111 and DSE HB3-1 have positive effects on drought resistance of C.viscosa seedlings.In this study,an OMF(hy-111)were isolated from the roots of C.viscosa,which has the ability to promote seedling growth and drought resistance,suggesting that OMF plays an important role in the ecological adaptation of epiphytic orchids.In addition,the discovery of DSE(HB3-1)with similar function to OMF hy-111 suggests that DSE also plays an important role in the stress tolerance of epiphytic orchid.In the future,the role of mycorrhizal fungi and other endophytic fungi in ecological adaptation of orchids should be further explored,which may provide theoretical support to the future protection of orchids,and functional agent development.