Diversity Of DSE In Roots Of Tussilago Farfara And Its Inoculation Effect With PGPR

Dark septate endophytes(DSE)can widely colonize in the root of host plants and have no disease on plants,and can promote the growth of plants.Tussilago farfara L.is a herb of Compositae,which has strong medicinal value and application prospect.In this study,the root soil was collected from Lujiakou village and Baima village in Pingshun County,Changzhi City,Shanxi Province,and North Campus of Changzhi college of Luzhou District,Changzhi City.BIOLOG ECO technology was used to study the functional diversity of microbial community in the sample plot of T farfara L..The physical and chemical analysis and enzyme activity determination of root soil were carried out to analyze the effects of continuous cropping and artificial cultivation on soil properties and microbial diversity;endophytic fungi were isolated and purified from the roots of T farfara L.by trapping and isolation method,and identified by morphological,molecular biology and phylogenic tree analysis;the ability of phosphate solubilization and IAA production of the isolated strains,as well as their tolerance to Na Cl and PEG6000 were determined;their growth status under different nutritional components was detected.Meanwhile,the strains with high IAA production efficiency were isolated from the rhizosphere soil of T farfara L.,and the related growth-promoting characteristics were assessed,including the ability to produce indole-3-acetic acid,siderophore and the phosphate solubilization capacity.The strains were identified by morphology,physiology-biochemistry,and 16 S r DNA sequence and phylogenic tree analysis.The suspected DSE strains isolated and rhizosphere bacteria with high IAA production were inoculated in the rhizosphere of T farfara L.seedlings respectively to observe the growth status of seedlings and determine the soil microbial diversity.The above results as a theoretical foundation for the collection,protection and utilization of DSE microbial resources and the study of functional flora of T farfara L.,and have important theoretical significance for the enrichment and development of DSE community.The main results are as follows:1.Typical DSE infection structure was observed in the roots of T farfara L.,mainly including septal hyphae and microsclerotia.2.Through the analysis of BIOLOG ECO microplate,the AWCD curves of Lu Jiakou plot,Bai Ma plot and Lu Zhou plot showed an upward trend,but the utilization ability of single carbon source of microorganisms in Lu Jiakou plot was quite different from the others.When reached 7 days,the community diversity and species homogeneity of LZ were significantly different from those of LJK and BM(P < 0.05),and the dominance index of the three plots was significantly difference(P < 0.05).The main carbon sources of the three plots were sugars,amino acids and acids.Lu Jiakou plot had the strongest utilization ability of D,L-α-glycerol and itaconic acid,Bai Ma plot had the strongest utilization ability of D-cellobiose and L-threonine,and Lu Zhou plot had the strongest utilization ability of liver sugar and L-serine.The contents of organic matter and total nitrogen in LZ plot were significantly different from the other two plots,the contents of total phosphorus in LJK plot were significantly different from the other two plots,and the p H of the three plots were significantly difference(P < 0.05).It can be seen that the continuous planting and artificial planting have certain influence on the soil properties and microbial diversity.3.Three IAA producing bacteria strains KD17 B,KD6B and KD26 B were isolated from the rhizosphere soil of T farfara L.Their IAA producing capacities were 31.38,31.78 and32.13 mg/L,respectively.KD17 B was Klebsiella oxytoca,KD6 B was Enterobacter cloacae ss dissolvens and KD26 B was Klebsiella aerogenes.4.A total of 19 endophytic fungi were isolated from the roots of T farfara L,belonging to 5 genera.Among them,5 strains of Phoma sp.,5 strains of Pleosporales sp.and 5 strains of Alternaria sp.,accounting for 26.32 %;three strains were Paraphoma sp.,accounting for15.79 %;one strain was Didymella sp.,accounting for 5.26%,and has the highest similarity with Didymella pinodella.5.Four endophytic fungi KDY2,KDY6,KD6 and KD14 had different ability to produce IAA and phosphate.KD14 had the strongest ability to produce IAA(22.49 mg/L)and KD6 had the strongest ability to dissolve phosphate(839.20 mg/L).Under the stress of Na Cl and PEG6000,the growth ability of different strains was difference,and the contents of MDA,soluble protein and SOD activity were difference.6.The effects of different carbon sources on the growth of four endophytic fungi KDY2,KDY6,KD6 and KD14 were different.The best carbon and nitrogen sources of KDY2 were soluble starch and diammonium hydrogen phosphate,KDY6 was rhamnose and peptone,KD6 was soluble starch,urea and ammonium sulfate,KD14 was soluble starch and tryptone7.Four DSE showed different compatibility with three PGPR,endophytic fungi KDY2 had compatibility with KD26 B and KD17 B,KDY6 was all rejected with three PGPR strains,KD6 had compatibility with KD17 B,KD14 had compatibility with KD26 B and KD17 B.The suspected DSE strain and PGPR were inoculated into the rhizosphere of T farfara L.seedlings through single or double inoculation.Among them,KD6 and KD14 had a greater effect on seedling growth,which could significantly increase leaf area and leaves.Quantity,and can increase the colonization rate of DSE in the root system.In addition,all inoculation treatments increased the diversity of rhizosphere microorganisms of T farfara L.,and the main trend was double inoculation > single inoculation > no inoculation.