Study On Physiological Resistance And Transcriptome Of Trichoderma Asperellum T203 Under The Action Of Bursaphelenchus Xylophilus

Trichoderma spp.is widely distributed in humid environments such as forests.wetlands,and farmland.It is an important type of biocontrol fungi and is widely used in forestry disease prevention and control.Pine Wilt Disease(PWD)caused by Bursaphelenchus xylophilus has led to the death of a large number of pine plants and caused a huge loss of forestry resources in my country.Trichoderma asperellum is a commonly used biocontrol strain,which has strong resistance t o p athogenic f ungi,s oil ne matodes and ot her h armful o rganisms.H owever,the physiological changes produced by Trichoderma in the process of responding to the action of B.xylophilus.There are few studies on the mechanism of molecular disease resistance.In order to study the resistance mechanism of T.asperellum under the action of B.xylophilus,this study used three extractants to extract the active metabolites of T.asperellum T203 to determine the virulence of the B.xylophilus.The enzyme activity kit was used to determine the pine wood.After B.xylophilus treatment,the 5 kinds of stress resistance enzyme activities of T.asperellum T203 were studied from the physiological and biochemical aspects.Then.the hypha samples of T.asperellum T203 treated with B.xylophilus for 120 h w ere s ampled a nd a nalyzed b y transcriptome s equencing,and 12 T.asperellum disease r esistance genes i nduced b y B.xylophilus were screened out,and biological information was subsequently used The scientific method analyzed the subtilisin family(peptidase S8)genes.The experiment m ainly draws the following conclusions:1.Three kinds of metabolites effect on B.xylophilus.the suppression rates reached 88.1%.88.75%,and 74.51%after 72 hour s of treatment..Indicating that T.asperellum has a strong inhibitorye ffect on B.xylophilus.D etermination of enzyme activitys howedt hat the five enzyme activities of CAT,SOD,POD,GST and β-1,3GA were significantly higher than that of 0 h in T.asperellum treated with B.xylophilus for 12 h.24 h.48 h.72 h and 120 h.indicating that CAT,SOD,POD plays a key role in the antioxidant function of nematodes,GST plays an important role in the mechanism of detoxification and metabolism,and β-1,3-GAplays a n important role in the mechanism of inducing resistance.2.A total of 508,581,808 clean reads were detected in the 6 samples of this transcriptome sequencing,and a total of 76.3G of sequencing data was obtained.The lowest unique map rate between the transcriptome sequencing data and the reference genome of T.asperellum is higher than 90%.indicating that the T203 strain is closely related to the reference genome,and the accuracy o f gene anno tation is hi gh.In the gene di fferential ex pression analysis.955 differentially expressed genes were screened,of which 372 genes were up-regulated and 584 genes were down-regulated.Using GO.KEGG databases to perform functional annotation and pathway enrichment analysis of di fferentially expressed genes,it w as f ound that genes encoding metabolites such as cell wall degrading enzymes,cell membrane degrading enzymes,antibiotics,etc,t hat expressed differently in the pr ocess of r esisting B.xylophilus.MAPK metabolism The expression of genes related to signal transduction,growth and development in the pa thway was significantly up-regulated.Twelve r esistance ge nes r elated to biological control w ere s elected f rom t he t ranscriptome d ata a nd ve rified b y qR T-PCR.T he r esults showed that the expression trends of these genes were roughly consistent with the analysis of transcriptome data.3.Thirteen s ignificantly up-regulated subtilisin family genes w ere s creened from t he transcriptome data and analyzed by bioinformatics.They were named TaprtP1-TaPrtP13.The analysis results showed that TaprtP1,TaPrtP2,and TaPrtP3 belong to the s ame subfamily.TaPrtP5,TaPrtP6,TaPrtP7 and TaPrtP8 belong t o t he s ame s ubfamily,a nd TaPrtP9,TaPrtP10,TaprtP11,TaprtP12 and TaPrtP13 belong to the same subfamily.The gene structure prediction shows that the gene structure of this family is similar.The a mino acid sequence alignment and the protein tertiary structure prediction show the catalytic triad structure,which can enable the protein to perform its corresponding function.P romoter prediction found the cis-acting elements involved in salicylic acid,growth and development,and stress resistance induction in T.asperellum.