A Preliminary Study On The Antibacterial Mechanism Of The Sulfone Series Of Compounds Containing 1,3,4-oxadiazole On The Pathogen Cacao Caerulea Of ​​tea Leaf Spot

Tea leaf spot disease caused by Lasiodiplodia theobromae is a new type of tea foliar disease found in Guizhou province,with a field incidence of 30 to 45%in Huishui county,Guizhou province.Sulfone compounds containing 1,3,4-oxadiazole are widely used in the designing of novel green pesticides because of their high efficiency,low toxicity and broad spectrum of anti-biological activity.Based on the study of the biological characteristics of Lasiodiplodia theobromae,we tested the antifungal activity of 5 sulfone compounds containing 1,3,4-oxadiazole.JHXJZ showed greater antifungal activity against L.theobromae than other sulfone compounds in vitro.The half effective concentration(EC₅₀)of JHXJZ is 3.54μg/mL.The in vivo antifungal test showed that the curative activity of JHXJZ at 50 and 100μg/mL was 14.55%and 41.78%,respectively.The curative activity was comparable to that of tebuconazole at the same concentration（14.08%and 38.50%,respectively）.This master thesis mainly focuses on the antifungal mechanism of JHXJZ against L.theobromae.Light microscopy showed that JHXJZ caused reduced,shortened,distorted,deformed mycelial branches and accumulation of unkown dark substances in the hyphae of L.theobromae.Scanning electron microscopy showed collapsed and shrunk hyphae.Transmission electron microscopy showed blurred cell wall,cell membrane and organelle as well as empty cytoplasm.Besides,fluorescence microscope showed reduced nucleus after staining with 4’,6-diamidino-2-phenylindole,and increased distances of mycelial septa and accumulation of fluorescent substances after staining with calcofluor white.Meanwhile,JHXJZ caused an increase in the permeability of mycelial cell membrane.And high performance liquid chromatography analysis showed that JHXJZ caused a decrease in mycelial ergosterol content.According to padj<0.05 and |log₂^FoldChange|>1,2084 differentially expressed genes were detected in transcriptome sequencing,in which 899 genes were up-regulated and1185 genes were down-regulated.GO enrichment analysis showed that the differential genes between JHXJZ-treated group and control group were mainly annotated in the biological processes of carbohydrate metabolic process,single-organism carbohydrate metabolic process,cellular carbohydrate metabolic process,the cellular components of membrane region,late endosome,late endosome membrane,and the molecular function of catalytic activity,cofactor binding,hydrolase activity,and acting on glycosyl bonds.Among them,16 differential genes were verified by Real Time qPCR（RT-qPCR）.Differentially expressed genes were also enriched in KEGG pathway.We particularly focused on those differentially expressed genes in the sterol biosynthesis pathway of L.theobromae because those genes were closely related to cell membrane components and sterol biosynthesis.Totally,6 genes were up-regulated and 2 genes were down-regulated.We used RT-qPCR to study the expression of the genes in the ergosterol biosynthesis pathway of L.theobromae after treatment of JHXJZ.Overall,farnesyl-diphosphate farnesyltransferase（FDFT1）,squalene monooxygenase（ERG1）,sterol 14alpha-demethylase（CYP51）,Delta-14-sterol reductase（ERG24）and sterol 22-desaturase（ERG5）were up-regulated and lysosomal acid lipase/cholesteryl ester hydrolase（TGL4）was down-regulated.This master thesis provides a basis for the further development of sulfone compounds containing 1,3,4-oxadiazole,and helps the control and management of tea leaf spot disease caused by L.theobromae.