Diversity And Biological Control Of Dark Septate Endophytic Fungi From Lycium Barbarum In Ningxia

Lycium barbarum is a traditional edible herb,at present,the research on L.barbarum mainly focuses on the research and development of antioxidant activity,nutritional components and related functional products at home and abroad,however,the study of its dark septate endophytes fungi was less reported.In this study,dark septate endophytic fungi were isolated from the roots of 8 cultivars of L.barbarum in Ningxia,their diversity and infection characteristics were studied;Taking C.gloeosporioides of Chinese wolfberry as target fungi,we screened for highly active DSE strains with significant inhibitory effect,discussed its bacteriostatic mechanism;The high activity DSE strain NQ8GII4 was used as the research object to study its culture traits,colonization mode and effects on host plants after colonization,the mechanism of its promoting effect is preliminarily revealed;The biocontrol effects of strain NQ8GII4 fermentation broth on pathogenic fungi of Chinese wolfberry,jujube and grape were tested,the potential of its application in biocontrol was discussed.The results are as follows:1?The roots of Ningqi-1,Ningqi-3,Ningqi-5,Ningqi-6,Ningqi-7,Ningqi-8,lycium barbarum var.auranticarpum and Lycium.ruthenicummurr were collected from Ningxia wolfberry cultivation garden.Strains were identified by morphological characteristic and rDNA-ITS sequence,and DSE of roots from L.barbarum were validated by Koch's Rule.The Result showed that DSE live as mircosclerotia in the roots of L barbarum.In this paper,279 DSE were isolated from roots of eight kinds of L.barbarum,belonging to 18 genera,which are rich in species diversity.Fusarium were a common and dominant genus in various breeds.The relative frequency was up to 85%.Monosporascus,Talaromyces and Earliella were firstly reported in L.barbarum.There were significant differences in biodiversity index,Evenness index and Simpson's index of DSE conmunity among different varieties of L.barbarum.DSE of L.barbarum were rich in resources of cultivated and wild varieties of L.barbarum,DSE could form a symbiotic relationship with L.barbarum and enhance the adaptability of wolfberry to the ecological environment.2?The inhibitory effect of endophytic fungi on C.gloeosporioides and the biocontrol potential of anthracnose were determined by indoor bacteriostatic test and in vitro test.The results showed that Fusarium strains NQ8GII4,NQ8GII7,NQ7GII4 and Talaromyces strains NQ6GIII11 had obvious antagonistic effects to C.gloeosporioides.The plate confrontation test showed that four endophytic fungi have strong nutrition and space competitiveness,the inhibition rate of strain NQ8GII4 to pathogenic bacteria reached to 93.43%.According to microscopic observation,endophytic fungi of L.barbarum could entangle and penetrate the hyphae of C.gloeosporioides,which could disintegrate the hyphae of C gloeosporioides.When the concentration of NQ7GII4 culture filtrate was 15%,the inhibition rate of mycelium was 70.75%,the inhibition rate of volatile substances produced by NQ8GII7 to C.gloeosporioides was up to 83.44%.The results of in vitro biocontrol experiment showed that vaccination of antagonistic endophytic fungi on tender fruits and leaves of L.barbarum could effectively inhibit the invasion of L.barbarum anthracnose and the expansion of lesions.The culture filtrate of strain NQ8GII4 10%had the strongest control effect on L.barbarum anthracnose?and the control effect reached to 92.54%and 95.57%,indicating that L.barbarum endophytic fungi had good biocontrol potential and application prospects.3?In order to understand the colonization mode of dark septate endoph>lic fungus NQ8GII4 in L.barbarum and its effect on L.barbarum plants after colonization.The infection behavior and colonization of strain NQ8GII4 in the root of L barbaruo were studied by stereomicroscope,optical microscopy and transmission electron microscopy.The results showed that when the mycelium of strain NQ8GII4 infects the root of L.barbarum for 24 hours,it adsorbs on the surface of the root,after 96 hours of L.barbarum infection by hyphae,hyphae colonized between epidermal cells and epidermal cells;Potted seedlings of L barbarum were treated with 10%fermentation broth,which significantly increased plant height and fresh weight.The results of defense enzymes activity in L.barbarum showed that the activities of PAL,POD and PPO defense enzymes in treatment group were significantly higher than those in control group,it was 1.10 times as much as the control group,indicating that strain NQ8GII4 could colonize L.barbarum root and establish reciprocal symbiosis relationship with host,which can promote host growth.4?In order to understand the growth characteristics of strain NQ8GII4 and its application prospects in production.The rnycelial growth and sporulation of strain NQ8GII4 were measured on four common media.The results showed that oat culture medium was the most suitable medium for rnycelial growth and sporulation.On the 9th day,the diameter of nycelium reached 6.47 cm,which was 1.32 times as much as the control,the sporulation amount was 47.84×105/mL,which was 1.34 times as much as the control.The biocontrol effects of L.barbarum,Jujube and grape on pathogenic fungi?ere studied,In the dish confrontation test,Strain NQ8GII4 can inhibite C.gloeosporioides,Alternaria alternate and Botrytis cinerea.Inhibition rate was 47.00%?65.77%;In vitro control effect test,the highest control effect of NQ8GII4 fermentation broth on pathogenic fungi of L barbarum,Jujube and grape was up to 92.54%.it has the same effect as the chemical agent Pythium.The results showed that strain NQ8GII4 was cultivate simplely,low cost and has good biocontrol effect.It could be used as a germinating strain for the development of microbial pesticides.The strain NQ8GII4 is easy to cultivate and has low cost and good biocontrol effect.It can be used as a germinating strain for the development of microbial pesticides.