| Plant endophytes are widely distributed in nature,and they can promote the growth of host plants or protect plants from pests and diseases,and have important application potentials.In this study,endophytic fungi targeting the devastating apple Glomerella Leaf Spot pathogen Colletotrichum fructicola were isolated and identified with a combination of tissue separation and confrontation co-culture method.Moreover,the antagonistic activity,indoor disease control efficacy and phytotoxicity of the fermentation products of the strain ferments,and the pathogenic characteristics of the strains were comprehensively evaluated,and the following main results were obtained:1.Based on confrontation co-culture assay,four antagonistic fungal strains were isolated from healthy garden plants such as begonia and papaya,which had good antagonistic activity toward Colletotrichum fructicola,namely lf,lq,ye and ccza,respectively,and the inhibition effects of the four antagonistic strains on PDA colony growth of 1104-7 in co-culture assays were 48.51%,38.06%,48.51% and 32.09%,respectively,and the inhibition effects of PDB broths of the four antagonistic strains on PDA colony growth of 1104-7 were 53.96%,74.74%,58.23%,and 49.21%,respectively.For lq strain,strong antagonistic activity exists in both the organic and inorganic extraction fractions of the ethyl acetate extract of PDB fermentation broth,whereas activities mainly exist in the organic phase for lf,ye and ccza.2.PDB fermentation broths of the four antagonistic strains all inhibited conidial germination and infection structure differentiation of strain 1104-7,but the inhibitory characteristics were different.lq fermentation broth completely inhibited conidia germination,lf fermentation broth did not affect conidia germination but strongly inhibited germ tube elongation,and ccza significantly inhibited conidia germination and appressoria simultaneously.In indoor disease control experiment,lf and lq had good protective effects on fruits,lq and ye had good therapeutic effects on fruits,and pretreatment with fermentation broth of all four strains delayed symptom development on apple leaves.3.Except for ye strain that is weakly pathogenic to apple fruit,none of the four strains caused disease to apple leaves or fruits.PDB fermentation broth of the lq strain could induce tobacco leaf necrosis,but had no effect on tomato leaves,while the PDB fermentation broths of lf,ye and ccza strains had no effect on tobacco and tomato.ye and lf fermentation broths significantly reduced wheat seed germination and elongation.4.Based on morphological characteristics and ITS molecular identification,lf strain was identified as Eutypella scoparia,lq strain as Epicoccum layuense,ye strain as Peroneutypa scoparia,and ccza strain as Phaeosphaeria fuckelii.5.Using Czapek as the base medium,the carbon-nitrogen source fermentation conditions were optimized for antagonistic metabolite production of the lf and lq strains.It was found that nitrogen sources such as yeast powder and ammonium peptone sulfate,as well as carbon sources such as maltose and mannitol significantly increased the production of antagonistic substances in lf strain.Nitrogen source of yeast powder and carbon source of soluble starch significantly increased the yield of antagonistic substances in lq strain.In summary,four candidate biocontrol bacteria with obvious antagonistic activity against Glomerella Leaf Spot pathogen were identified through systemic screening,and their bioncontrol traits in terms of antagonisitc properties,disease prevention effects,and phytotoxicity.This study built up new biocontrol resources against Colletotrichum disease on apple,which laysa good foundation for developing novel biological control strategies against the disease. |
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