Studies On The Mechanism Of Dry Mycelium Of Penicillium Chrysogenum-induced Plant Resistance

Controlling plant disease has been an important issue in agricultural production.Traditional chemical control of plant disease with pesticides that suppress or kill pathogens directly has been made great contribution to agricultural production.However,adverse effects of pesticides on environmental and human health have been caused worldwide attention,leading to the exploring of alternatives that accord with sustainable agriculture.Plants have an innate immune system that recognizes pathogens and responds byactivating specific defenses that are directed against the invader,after which a systemic defense response is often triggered in distal plant partsto quickly and intensively protect plant against subsequent invasion.The systemic defense response is called induced resistance and protects plant from wide range invaders.Elicitors that activate induced resistance including pathogens and molecules derived from microbes and plants,and can act as alternative to traditional chemical pesticides.Dry mycelium of Penicillium chrysogenum comes from penicillin industrial production,which includes mycelium,metabolites and fermentation residues,has been shown to protect plants from diverse disease,indicating the ability of dry mycelium of Penicillium chrysogenum induced plant defense responses.However,the mechanism of how dry mycelium of Penicillium chrysogenuminduces plant resistance is not clear,the effect and mechanism of dry mycelium of Penicillium chrysogenumprotects plants from viral diseases is remain unknown.What's more,the effective component in dry mycelium of Penicillium chrysogenumremains unclear.In this work,the dry mycelium of Penicillium chrysogenum-induced plant defense responses were studied by biochemical and molecular biological method,as well as systemic resistance against viral disease induced by the effective extracts derived from dry mycelium of Penicillium chrysogenum in tobacco.Furthermore,putative peptides derived from dry mycelium of Penicillium chrysogenum that capable of inducing plant defense responses were separated and purified using methods of analytical chemistry.The main results are as followed:(1)The effect and mechanism of dry mycelium of Penicillium chrysogenum-induceddefense responses in plant were investigatedThe water extracts of dry mycelium of Penicillium chrysogenum(DMP)-induced defense responses and signaling pathways were studied in this work.Transcripts of four genes involving in the synthesis of secondary metabolites,and the contents of phenolic compounds,scopoletin and lignin,were rapidly induced in DMP-treated BY-2 cell suspensions derived from Nicotiana tabacum L.cv.Bright Yellow-2,indicating a DMP-induced resistance in BY-2 cells.Transcripts of salicylic acid(SA)-dependent gene PR-la were rapidly induced after DMP-treatment,revealing the activation of SA-dependent signaling pathway.The expression of jasmonic acid(JA)-responsive gene PDF1.2suggested that DMP might induce distinct signaling pathways simultaneously.(2)The effective extracts that derived from DMP-induced resistance in plant were investigatedThe Polysaccharide derive from DMP(PsD)and Peptides derive from DMP(PDMP)-induced defense responses in tobacco BY-2 cells were studied.Transcripts of defense response genes and the contents of defense-related secondary metabolites were rapidly induced in PsD/PDMP-treated tobacco BY-2 cell suspensions,which indicated that PsD and PDMP act as elicitors capable of inducing resistance in plant.(3)The effect and mechanism of PDMP-induced systemic resistance in tobacco was studiedPDMP-induced systemic resistance in Nicotiana glutinosa was investigated in this work for further understanding of PDMP-induced resistance in plant systemic resistance.Transcripts of defense related genes in PDMP-treated leaves and systemic leaves of N.glutinosawere rapidly up-regulated after PDMP treatment,less numbers and smaller size of the TMV lesions were detected in PDMP-treated plants compared with that in control plants,indicating that PDMP induced systemic resistance in N.glutinosa.The detection of genes involved in defense signaling pathways and hypersensitive response,as well as the detection of callose deposition in TMV-inoculated leaves revealed that PDMP induced defense responses through the SA-dependent signaling pathway on the on hand,referring to the interactions between elicitor and plant;On the other hand,PDMP activated priming through the JA/ET-dependent signaling pathway,resulting in faster and more intensive expression of hypersensitive response,which represented a more effective resistance against TMV,referring to the interactions between elicitor,primed plant and pathogen.(4)The effect of PDMP protecting susceptible tobacco plants from TMV were investigatedTo comparing the similarities and differences of PDMP-induced resistance in resistant and susceptible tobacco plants,the spread of TMV in N.benthamiana plants was studied using a TMV vector that carrying the GFP gene mediated by agrobacterium.The cell-to-cell movement and long-distance movement of TMV vector were restricted in PDMP-pretreated plants,indicating that PDMP induced resistance against TMV in N.benthamiana plants.The results of the expression of defense-related genes and callose deposition revealed that the primed deposition in PDMP-pretreated leaved played a central role in resistance against TMV.This work revealed the mechanism of PDMP-induced resistance against TMV in susceptible tobacco plants,indicated that JA/ET-dependent priming playing a key role in PDMP-induced TMV resistance both in resistant and susceptible tobacco plants.The primed callose deposition is the key factor in PDMP-induced resistance against TMV in susceptible tobacco plants,while faster and more intensive expression of hypersensitive response primed by PDMP plays an important role in resistance tobacco plants that inoculated with TMV.This work also provided foundation for further study on the regulatory mechanism of callose deposition in primed tobacco plants interacted with TMV.(5)Established the separation and purification system for peptides derived from DMPThe peptides derived from DMP were further separated and purified using size-exclusion chromatography and high performance liquid chromatography.The results indicated that the peptides derived from DMP were comprised of a series of small molecular peptides with similar hydrophobicity.Above all,this work revealed the key substance of DMP,as well as the mechanisms of DMP-and PDMP-induced resistance in tobacco.Furthermore,the established of the separation and purification system for peptides derived from DMP provided the possibility for further investigation of novel peptide elicitor in DMP.This work provided theoretical foundation for application of DMP and its effective extracts in agricultural production.