| Plant endophytic fungi are considered to be one of the biocontrol factors with great application potential.A large number of studies have shown that in addition to producing antibacterial substance and competing with pathogen for nutrients and space,antagonistic endophytic fungi can also act as elicitors and induce plant’s defense responses to improve disease resistance,but the mechanism of its action is still unclear.In this study,the antagonistic endophytic fungus Fusarium nematophilum NQ8GⅡ4 isolated in the previous stage was used as the material to explore the colonization ability and growth-promoting effect of the NQ8GⅡ4 strain on the host and non-host plants.F.oxysporum and Fusarium oxysporum f.sp.Lycopersici Snyder et Hansen was used as the pathogen target.The pot experiment,determination of defense enzyme activity,and histochemical staining were conducted to study the induced disease resistance and physiological and biochemical mechanism of NQ8GⅡ4 strain on host plants(Lycium barbarum)and non-host plants(tomato).Finally,on the basis of the above,through transcriptome sequencing analysis,the molecular mechanism of NQ8GⅡ4 strain-inducing disease resistance in host plants(L.barbarum)and non-host plants(tomato)was explored from the gene level.The results of the study are as follows:1.The biological methods were conducted to determine the growth-promoting potential of the NQ8GⅡ4 strain.Using a transformed strain of NQ8GⅡ4-GFP expressing the green fluorescent protein(GFP)to inoculate in six plants(L.barbarum,tomato,wheat,zucchini,lettuce,and celery)by the pouring-root method,and the colonizing ability of NQ8GⅡ4-GFP in host and non-host plants was monitored using confocal laser scanning microscopy.Meanwhile,the effects of different concentrations of NQ8GⅡ4-GFP inoculant on the growth of L.barbarum seedlings were examined.The results showed that the concentration of gibberellin produced from wild-type NQ8GⅡ4 strain reached up to 16.6 μg/mL at 5 days,and it also had certain abilities to solubilize phosphorus,produce siderophores,cellulase,and protease.Furthermore,the NQ8GⅡ4-GFP strain was shown to colonize the roots ofL.barbarum,tomato,and wheat,but was unable to colonize celery,lettuce,and zucchini.Root colonization rates indicated a significant difference among plants,with the highest colonization rate of 93.9%for the host plant L.barbarum,91.84%for the non-host plant tomato,and only 21.43%for wheat.The pot experiment showed that different inoculation concentrations had different effects on the growth indexes of L.barbarum seedlings,of which 1×105～1×106 CFU/mL spore solution could significantly promote the growth of L.barbarum seedlings.2.In order to study the control effect and mechanism of the NQ8GⅡ4 strain on tomato Fusarium wilt,the face-to-face-culturing method was used to determine the inhibitory effect of the NQ8GⅡ4 strain on the growth of F.oxysporum mycelium,and its pot control effect on tomato Fusarium wilt was determined.The Defense enzyme activity assay,histochemical staining,and defense gene expression were used to analyze its mechanism of action.The results showed that preinoculation with the NQ8GⅡ4 strain had a 63.00%inhibition rate on tomato F.oxysporum mycelium.The pot control effect of NQ8GⅡ4 strain on tomato Fusarium wilt reached 76.85%.Inoculation of NQ8GⅡ4 strain can increase the activities of defense enzymes such as POD,SOD,and CAT in tomato leaves,remove the accumulation of reactive oxygen species in tomato leaves under F.oxysporum stress and reduce the degree of membrane lipid peroxidation and cell death;the NQ8GⅡ4 strain induces defense-related gene expression of tomato roots such as PR1a,GLUA,PAL,NPR1,and Basic CHI in the roots improving the disease resistance of tomato.3.The biological control and physiological and biochemical mechanism of NQ8GⅡ4 strain on L.barbarum root rot were studied through plate confrontation culture,greenhouse pot biocontrol experiments,the defense enzyme activity assay,and histochemical staining.The results showed that pre-inoculation with the NQ8GⅡ4 strain could effectively inhibit the growth of mycelium of F.oxysporum,and the inhibition rate could reach 70.07%;Under the stress of F.oxysporum,the NQ8GⅡ4 strain promoted the growth of L.barbarum,and the agronomic indexes increased by 10.81%-80.42%compared with the single-inoculated pathogen group(P).Compared with the blank control,the NQ8GⅡ4 group(E)inoculated alone increased the activities of defense enzymes such as SOD,PAL,CAT,POD,and PPO in L.barbarum leaves by 13.24%-208.27%;The activities of POD,SOD,CAT and PAL in the challenge inoculation group(E+P)increased by 19.73%-78.56%,while the MDA content decreased by 27.50%.Inoculation of the NQ8GⅡ4 strain could remove the accumulation of reactive oxygen species in the leaves of L.barbarum,reduce the degree of membrane lipid peroxidation and cell death,and improve the disease resistance of L.barbarum.4.In order to study the molecular mechanism of NQ8GⅡ4 strain-induced disease resistance of L.barbarum,the tissue culture seedlings of L.barbarum inoculated with NQ8GⅡ4 strain(+E)and uninoculated group(-E)for 3 and 7 days were used as materials,and transcription analysis was performed by Illumina high-throughput sequencing technology.Effect of inoculation of NQ8GⅡ4 strain on the gene expression profile of L.barbarum.The results showed that the Q30 of the sequencing samples were all greater than 92.65%,the average GC content was above 43.08,and the quality was qualified.Compared with the uninoculated treatment,there were 2130 differentially expressed genes(DEGs)of the NQ8GⅡ4 strain infected for 3 days,of which 1935 were up-regulated DEGs and 198 were down-regulated DEGs.There were 2098 DEGs of the NQ8GⅡ4 strain infected for 7 days,including 1302 up-regulated DEGs and 796 down-regulated DEGs.GO enrichment analysis found that the DEGs at 3 and 7 days were mainly enriched in immune system regulation process,antioxidant activity,and electron carrier activity;KEGG enrichment analysis showed that the DEGs were mainly enriched in pathways with plant hormone signal transduction,plantpathogen interaction,and phenylpropanoid biosynthesis.Transcription factor analysis showed that AP2/ERF,WRKY,bZIP,NAC,and MYB family members involved in plant stress resistance were all induced and expressed to varying degrees. |
