Analysis Of Differentially Expressed Genes In Pathogen Of Poplar Leaf Blight Responsing H₂O₂ Stress

Poplar leaf blight is a leaf disease caused by Alternaria alternata.It is one of the more serious forest diseases in China,and could cause a large number of poplar leaves to fall off in advance and affect the growth and development of trees and wood quality.Revealing the pathogenic genes of Alternaria alternata is of great significance to clarify the pathogenic mechanism and control of poplar leaf blight.In this study,based on Alternaria alternata isolated from leaves,the pathogen of poplar leaf blight was identified by morphology and molecular biology.The differentially expressed genes of Alternaria alternata under hydrogen peroxide stress were identified by transcriptome sequencing.The main results are as follows:（1）The pathogen of poplar leaf blight was isolated from the leaves in Populus simonii×P.nigra.Through morphological identification,it was confirmed that the strain belonged to Alternaria alternata;Koch’s rule proved that Alternaria alternata could infect the leaves of Populus simonii×P.nigra,causing disease spots and even necrosis.The ITS sequence was cloned by PCR.Multiple sequence alignment showed that it was highly similar to the pathogenic fungi of strawberry rot,cucumber rot and tomato rot.Through the construction of phylogenetic tree,it was found that the pathogenic fungus of poplar leaf blight had the closest genetic relationship with Alternaria tenuissima,the pathogenic fungus of Atractylodes black spot.（2）The third generation of Alternaria alternata genome was sequenced by nanopore sequencing technology platform.Through repeated sequence prediction and coding protein gene prediction,a total of 13063 genes were obtained.A total of 724 differentially expressed genes of Alternaria alternata in response to H₂O₂ stress were identified by transcriptome sequencing,including functional annotations of 625 DEGs encoding proteins with 196 up-regulated DEGs and 429 down-regulated DEGs.The results of GO enrichment analysis and KEGG enrichment analysis showed that DEGs were mainly involved in biological processes,molecular functions and cell components.The number of DEGs enriched in synthetic antibiotic pathway was the most,but the enrichment significant level was lower;The number of DEGs enriched in VB6,tryptophan,starch and sucrose metabolism and lysine biosynthesis is small,but the enrichment level is significant.（3）The conserved domains of CAT,POD and SOD families were obtained through Pfam database,and Hmmsearch was used to search the members of protein family.The phylogenetic relationship of different protein families was analyzed through sequence alignment and phylogenetic tree construction.The results showed that the structures of CAT,POD and SOD family of Alternaria alternata were similar to those of maize,Arabidopsis,rice blast pathogen and yeast.（4）Through the analysis of differentially expressed genes,it was found that 11 of the 22CAT,POD and SOD genes were differentially expressed under hydrogen peroxide stress,and45%of the differentially expressed genes were up-regulated and 55%were up-regulated.The expression level of 12 toxin synthesis related genes in poplar leaf blight changed significantly under hydrogen peroxide stress,including 1 up-regulated gene and 11 down regulated genes.In the hydrogen peroxide under the condition of black grape grain mildew toxin synthesis related gene expression level increased by 2.99 times the water treatment control group.Among the 11down regulated genes,there were 5 genes related to aflatoxin synthesis,2 genes related to killer toxin subunit synthesis,and tes toxin synthesisζToxin,Alternaria toxin synthesis,glial toxin biosynthesis protein,etc.the expression level of these genes decreased by 2.7-13.6 times.In this study,through the identification of DEGs of poplar leaf blight pathogen in response to hydrogen peroxide stress,could provide a theoretical basis for the pathogenic mechanism of poplar leaf blight pathogen and the prevention and control of forest diseases.