Resources And Functions Of Dark Septate Endophytes From Chinese Wolfberry Root

Dark septate endophytes(DSE)are a kind of important endophytic fungi,which can infect plant roots,form dark,transseptal hyphae and microsclerotia of different shapes,establish a good symbiotic relationship with plants,promote host absorption of nutrients,enhance host resistance to heavy metals,and promote plant adaptability to stress,which is a research hotspot in recent years.Chinese Wolfberry(Lycium barbarum)is a perennial deciduous shrub growing in northwest China,and it is also a typical representative of medicinal plants on the Silk Road.It is rich in endophytic fungal resources.However,there are few reports on the diversity and function of DSE fungi in Chinese Wolfberry.In this study,root and soil samples of Chinese Wolfberry were collected from two cultivation areas in Yinchuan,Ningxia,and the diversity of endophytic fungi in the root system of Chinese Wolfberry was analyzed by high-throughput sequencing technique;the culturable DSE strain of Chinese Wolfberry root was obtained by tissue isolation method,and its colonization characteristics in the root system of Chinese Wolfberry were observed;the law of DSE colonization of Chinese Wolfberry was investigated,and the effect of soil factors on DSE colonization was discussed.Using Fusarium oxysporumas the target bacteria,the strain DSE with obvious inhibitory effect was screened on Chinese Wolfberry root rot disease and its biocontrol mechanism was discussed.The purpose of this study was to explore the species diversity,ecological distribution and ecological function of DSE fungi in Chinese Wolfberry,and to provide reference for making full use of DSE fungi resources.The main results are as follows:1.The community composition and species diversity of endophytic fungi in the roots of Chinese Wolfberry were studied based on high-throughput sequencing.The results showed that a total of 390,783 valid sequences were obtained,and 712 OTU,were obtained to remove the unidentified endophytic fungi,belonging to 11 phylums,50 orders,94 families and 155 genera.Among them,Ascomycota was the dominant phylum,Hypocreales was the dominant order,and Nectriaceae was the dominant family.The study found that in different soil types,the diversity and richness of endophytic fungi community in the roots of Chinese Wolfberry were slightly higher than that of loam soil,and β diversity analysis showed that the diversity of endophytic fungi in samples H1,H2,H3,S1,S3 was not significantly different,but was a significant difference in sample S2.There were differences in endophytic fungi in Chinese Wolfberry root samples between the two soil types.At the genus level,Fusarium was a landmark group of endophytic fungi in the roots of Chinese Wolfberry in sand.2.It was studied that the community composition and species diversity of DSE in the root system of Chinese Wolfberry.The results showed that 41 DSE strains were isolated from 2 plots.The strains were identified according to morphological characteristics and rDNA-ITS sequence analysis,belonging to 4 orders,6 genera and 8 species,of which 5 species were identified,which were N.radicicola,F.acuminatum,C.cladosporioides,P.cyclamini and A.alternata;the other three were identified as Neonectria sp.and Pleosporales sp.and A.alternata had the highest isolation frequency and was the dominant population in the root of Chinese Wolfberry.3.The staining effects of Quink ink,trypan blue,acid fuchsin,Sudan red Ⅳ and aniline blue on endophytic fungi of Chinese Wolfberry roots were studied.The results showed that Quink ink and Trypan blue had good staining effect,but considering the safety,Quink ink was more suitable for the observation of infection and colonization of endophytic fungi in Chinese Wolfberry roots.4.The colonization of DSE and its relationship with soil factors were studied.The results showed that DSE formed typical structures such as dark color,septum hyphae and microsclerotia in the roots of Chinese Wolfberry.The average DSE colonization rate of the two plots was 48.41%.There were significant differences in DSE colonization rates at different depths of Chinese Wolfberry roots,and the maximum colonization rate appeared in the 20～40 cm soil layer.The results of correlation analysis with soil factors showed that the colonization rate of DSE was significantly correlated with soil alkali-hydrolyzable nitrogen,available phosphorus,organic matter,alkaline phosphatase,pH and sucrase.The results of variance decomposition showed that root depth and soil physical and chemical properties alone explained 63%and 91%of the total variance,respectively.5.Using F.oxysporumas as target bacteria,the inhibitory effect of DSE strain on F.oxysporumas and its control effect on Chinese Wolfberry root rot disease was determined by confrontation culture in dish and pot experiment.The results showed that all the 8 DSE strains had obvious inhibitory effect on the mycelial growth of F.oxysporumas,and the inhibition rate of strain S12 to F.oxysporumas was the highest,with an inhibition rate of 81.47%.When the concentration of fermentation filtrate of strain M1 was 50%,the inhibition rate of spores of F.oxysporumas was the highest,up to 91.48%,and the inhibition rate of volatile substances of strain M1 was the best,and the inhibition rate was as high as 94.10%.The results of pot control experiment showed that the fermentation filtrate of strain M1 and S12 had the best biocontrol effect on Chinese Wolfberry root rot disease,and the control effects were 89.54%and 83.17%,respectively.It showed that DSE had good biocontrol potential and application prospect.