Identification Of Bacteriostatic Active Components Of Bacillus Amyloliquefaciens CGMCC15838 And Their Application In Anti-Fungal Diseases Of Wheat

At present,the prevention and control of crop diseases are mainly through chemical means,but chemical agents usually increase the resistance of plant pathogens to chemicals,that will not only cause pesticide residues in the soil,but also disrupt the ecological balance.The biocontrol bacterium Bacillus,as an antagonistic antibacterial for the biological control of crop diseases,can produce metabolites to inhibit the growth of pathogenic bacteria,it can also improve the soil environment,and pathogenic bacteria will not develop resistance.Therefore,related research on the discovery of new bacillus can enrich the resources of biocontrol bacteria in plant diseases,and provides a scientific basis for the molecular design,R&D and application of green control agents.In the previous study,we obtained a biocontrol bacterium CGMCC15838 with strong antibacterial effect,then,a preliminary analysis of its bacterial species relationship was carried out,and its antibacterial active components were also identified and applied.The results of the study are as follows:（1）Identification of biocontrol bacteria CGMCC15838By observing its morphology on the plate,observing its microscopic appearance with an electron microscope after staining,determining the 16S r DNA sequence,and establishing a phylogenetic tree,we determined that the biocontrol bacteria CGMCC15838 is Bacillus amyloliquefaciens.（2）CGMCC15838 has an inhibitory effect on Fusarium graminearumObservation through microscope revealed that after 10 hours,the germination rate of the conidia of F.g in water and LB broth reached 100%and the hyphae were very long,but the conidia in the fermentation broth and fermentation supernatant of CGMCC15838 did not germinate,and the hyphae were malformed.In addition,the content of MDA in the mycelium during the treatment increased by 0.187μmol·ml^-1compared with the control.The above results indicate that the biocontrol bacterium CGMCC15838 can deform and destroy the conidia and hyphae of F.g,thereby inhibiting the germination of conidia and the growth of hyphae,and destroying the cell membrane system of F.g.（3）The inhibitory effect of CGMCC15838 on wheat powdery mildewBy observing the wheat leaves that were smeared or sprayed with CGMCC15838 and were infected with powdery mildew,it was found that the number of ungerminated powdery mildew spores on the wheat leaves after treatment with this strain reached more than 50%.The severity of leaves treated with dd H₂O and LB broth is defined as severe,4%fermentation supernatant is moderate,10%and 6%fermentation supernatant as mild,the fermentation broth(OD₆₀₀=2),fermentation supernatant and triadimefon（1.6 mg/ml）are healthy.（4）The study of antibacterial substances in CGMCC15838 and the determination of their stabilityThe lipopeptide compounds of CGMCC15838 were extracted by acid precipitation and methanol extraction,and the method of filter paper found that its antibacterial rate against F.g reached 65.8%.The lipopeptide compound has good stability at 40-80°C,and the bacteriostatic rate is more than 33%.After UV irradiation for different time,its inhibition rate to F.g was also above 23%,indicating that it has a good tolerance to UV rays.By amplifying the synthetic genes of lipopeptide antibiotics such as sfp,itu A and fen B from the genome of CGMCC15838 and sequencing analysis,it is speculated that there may be surfactant,iturin A and fengyuansu metabolic synthesis operon sequences in its genome,which can metabolize to produce the above-mentioned three types of lipopeptide compounds.