Study On The Diversity Of Endophytic Fungi And Its Promoting Mechanism In Phoebe Bournei

Phoebe bournei?Hemsl.?Yang,also known as Phoebe,is an evergreen tree of Phoebe in Lauraceae.It is a unique species in China and a national second-class protected plant.In this paper,the healthy Phoebe bournei in Screw case nature reserve of Duyun City,Guizhou Province was used as the experimental material.According to the four seasons of spring,summer,autumn and winter,the samples were taken from Phoebe bournei trees of different tissue parts.The endophytic fungi of Phoebe bournei were isolated and identified by the method of tissue block separation,and identified by morphology and r DNA-ITS.The diversity of endophytic fungi was analyzed.The potential of endophytic fungi biocontrol was evaluated and screened from endophytic fungi with strong antibacterial activity The strains with the ability of dissolving inorganic phosphorus,dissolving potassium and secreting indoleacetic acid?IAA?were selected.The selected growth promoting strains were connected to the seedlings of Phoebe bournei a by pot experiment,and their effects on the growth,physiological and biochemical indexes of Phoebe bournei a were observed and measured,which provided the basis for the effective protection of Phoebe bournei a and the development and utilization of endophytic fungus resources.The specific research results are as follows:?1?389 endophytic fungi were isolated from the roots,stems and leaves of Phoebe bournei in spring,summer,autumn and winter.According to the statistics of the number of endophytic fungi in the tissue of Phoebe bournei,the number of strains isolated from the root is the most,which is 186,accounting for 47.82%of the total number of strains;the number of strains isolated from the stem is 128,accounting for 32.90%of the total number of strains;the number of strains isolated from the leaf is the least,which is 75,accounting for 19.28%of the total number of strains.According to the statistics of the number of endophytic fungi in spring,summer,autumn and winter,98 strains were isolated in spring,accounting for 25.19%of the total number of strains;78 strains were isolated in summer,accounting for 20.05%of the total number of strains;the number of strains isolated in autumn was the most,accounting for 40.10%of the total number of strains;the number of strains isolated in winter was the least,accounting for 14.65%of the total number of strains.The results showed that 389 strains of endophytic fungi belong to 2phylum,4 classes,11 orders,30 genera and 45 species.Among them,Ascomycota is the dominant phylum,accounting for 88.15%;sordariomycetes is the dominant phylum,accounting for 51.67%;diaporthales is the dominant phylum,accounting for 25.97%;the dominant genera of Endophytic Fungi of Phoebe bournei include Diaporthe,accounting for 17.48%;Phyllostica,accounting for 10.28%.?2?The analysis of the diversity of endophytic fungi showed that the Shannon-Weiner diversityindex?H'??Simpson's diversity?D??Margalef'sindex?M?of endophytic fungi in different seasons was:Autumn>spring>summer>winter;The evenness index?E?is:spring>summer>autumn=winter.According to the similarity index?Cs?analysis,the endophytic fungi groups in autumn and spring are the most similar,Cs=0.97,and endophytic true between autumn and winter.The difference of bacterial groups was the largest,Cs=0.34.The Shannon-Weiner diversityindex?H'??Simpson's diversity?D??Margalef'sindex?M?of endophytic fungi in different tissue parts was:root>stem>leaf;the evenness index?E?is leaf>root>stem.The similarity index?Cs?analysis showed that the endophytic fungi group of roots and stems was the most similar,Cs=0.82,and the difference between roots and leaves was the largest,Cs=0.46.?3?The evaluation of biocontrol potential showed that 27 strains of endophytic fungi,accounting for 60%of the total number of endophytic fungi,were screened by using 8 plant pathogens as indicator bacteria.Among them,strains 3-ye-3,3-ccg-2,3-ccg-1,3-hcg-5 and1-cg-18 have obvious inhibitory effects on 8 common plant pathogens,among which 3-ye-3 has the best antibacterial effect,with the antibacterial range of 8 pathogens ranging from 25.20%to85.20%;in the antibacterial experiment of volatile metabolites,only 3-ccg-1 and 3-hcg-5 can inhibit pathogens by producing volatile metabolites;in the liquid metabolism,only 3-ccg-1 and3-hcg-5 can produce volatile metabolites Only 1-cg-18,3-ccg-2,3-ye-3 liquid metabolites can inhibit the pathogenic bacteria.The main metabolites are flavonoids,aldehydes,phenols and organic acids.?4?The evaluation of the growth promoting effect showed that on the basis of 27 endophytic fungi with antimicrobial activity,the strains with strong phosphorus dissolving ability were1-cg-13 and 1-cg-2,and their phosphorus dissolving rates of Ca₃?PO₄?₂were 24.51%and 21.89%,respectively.In addition,strains 1-cg-13 and 3-hcg-5 showed a fast growth tendency to K-feldspar powder.Salkowski colorimetry was used to determine the IAA secretion ability of 27endophytic fungi.Four strains with IAA secretion activity were selected as:1-cg-18,1-cj-4,4-cg-4,4-hby-3,and their IAA secretion was 41.39 ug/ml,40.16 ug/ml,37.68ug/ml,35.75 ug/ml,respectively.?5?Seven strains of endophytic fungi?1-cg-13,1-cg-2,3-hcg-5,1-cg-18,1-cj-4,4-cg-4 and4-hby-3?were selected to connect to the seedlings of Phoebe bournei macrankinensis through pot experiment,and were used to detect the growth and physiological effects on the seedlings of Phoebe bournei macrankinensis.After inoculating 7 endophytic fungi for 90 days,it was found that the hyphae of 7 experimental groups had been successfully colonized in the roots of seedlings through microscopic examination.The results of the effects on the growth and physiology of the seedlings showed that:7 endophytic fungi improved the growth index,chlorophyll,soluble protein,root activity,SOD activity,POD activity,CAT activity,soil nitrogen,phosphorus,potassium nutrient and soil enzyme activity in varying degrees Among them,1-cg-18was the best,the plant height,ground diameter,total chlorophyll and root activity indexes were significantly different?P<0.05?,and the above indexes were increased by 49.78%,47.52%,51.24%and 45.95%respectively.