| Bacillus licheniformis W10 is a biocontrol bacterium screened from plant rhizosphere.It can well inhibit Botrytis cinerea,Monilinia fructicola and Phomopsis amygdali,etc.,and has significant control effect on these diseases in the field.Our previous reaearch found that W10 could produce an extracellular antifungal protein with subtilisin activity,so we named it W10-Sp1(subtilisin protein 1).This tesearh explored the broad-spectrum anti-pathogen effect of W10-Sp1 on various plant pathogenic fungi and bacteria.In order to further understand the mechanism of it,treatments of W10-Spl dealed with P.amygdali and tobacco were used to analyze the different expressing genes according to the transcriptomics,which was helpful to understand the mechanism of W10-Spl anti-pathogen and inducing plant disease resistance.The results will lay the foundation for the application of W10-Sp1.Confrontation culture tests showed that W10-Sp1 had obvious inhibitory effect on a variety of plant pathogenic fungi.It inhibited mycelial growth and spore germination,caused the abnormal hyphal growth,or made mycelium thicker and deformed.Besides that,it could inhibit the growth of a variety of pathogenic bacteria.The study also found that whether peach branches were treated with W10-Spl before or after inoculation,all of them had significant inhibiyory effects on peach shoot blight.Peach fruit were treated by W10-Spl first and then inoculated M.fructicola which could also significantly inhibit the occurrence of peach brown rot.These results suggested that W10-Sp1 had the effect of anti-pathogen,and might induceplant disease resistance at the same time.Further treatment of tobacco with W10-Sp1 showed resistance to B.cinerea,and the optimal efficiency was inoculating B.cinerea after 12 h of treatment.The differentially expressed genes of P.amygdali after W10-Spl treatment for 72 h were analyzed.Differentially expressed genes were mainly enriched in carbon metabolism,amino acid metabolism and secondary product metabolism pathways through KEGG metabolic pathway.Thus,W10-Sp1 might affect the growth of P.amygdali by inhibiting P.amygdali’s energy production,protein degradation,and affecting the synthesis of secondary products.Among them,subtilisin plays important roles in fungal spore formation,protein degradation and autophagy.W10-Sp1 inhibites the expression of multiple subtilisins of P.amygdali that might be an important reason for the destruction morphology and growth of P.amygdali.In addition,acetyl-CoA acetyltransferase(GME8738_g)is differentially up-regulated in the metabolism of propionate,"glyoxylic acid and dicarboxylic acid metabolism" and fatty acid metabolism pathways.W10-Sp1 might regulate the secondary metabolism of P.amygdali through regulating acetyl-CoA acetyltransferase.Analysis of the molecular mechanism of W10-Sp1 inducing plant disease resistance at the transcriptome level showed that the differentially expressed genes of tobacco treated by W10-Sp1 were mainly enriched in "photosynthesis-antenna protein",biosynthesis of secondary metabolites,phytohormone signal transduction,"interaction between plants and pathogenic bacteria",and "starch and sucrose metabolism" pathways.These pathways have an important relationship with the growth and disease resistance of tobacco.The key genes of the "photosynthesis-antenna protein" metabolic pathway were up-regulated,and hormones related to tobacco growth and development,such as auxin,cytokinin,gibberellin and other related genes in the phytohormone signal transduction pathway,all showed different degrees of up-regulation.These indicated that W10-Sp1 might promote the growth of tobacco by inducing photosynthesis of tobacco and the synthesis of hormones such as auxin.In the plant hormone signal transduction pathway,the negative regulator JAZ and the transcription factor MYC2 that regulate jasmonic acid(JA)signaling pathway,were significantly down-regulated.In the salicylic acid(SA)signaling pathway,the positive regulator NPR1 showes up-regulated expression,and the degree of up regulation of the transcription factor TGA was much more significant.These results indicate that W10-Sp1 might activate the tobacco JA signaling pathway and salicylic acid signaling pathway.W10-Sp1 possibly be respectively recognized by Fls2,Pto,and RIN4 to activate PTI and ETI in plants to play its inducing disease resistance. |
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