Biocontrol Activities Of Several Biocontrol Bacteria On Rice Blast And The Substantial Basis

Rice is the world’s main food crop,and more than half of the world’s population uses rice as a main food.The occurrence of rice blast has seriously affected the production of rice.At present,the control of rice blast mainly depends on the selection of disease-resistant varieties and chemical pesticides.The blast fungus is prone to mutation,there are many physiological races,it takes a long time to select resistant varieties,and the application of resistant varieties also has little effect;although traditional chemical pesticide control has been effective,the long-term use of chemical pesticides has a beneficial effect on human and animal health and ecology.Hidden dangers caused by environmental safety.Bio-pesticides are environmentally friendly and non-residue.They are important options for sustainable agricultural development,protecting the ecological environment,and ensuring food safety.This paper is based on the 8 strains of biocontrol bacteria accumulated by the research team in the early stage.By the plate pairing method and evaluation biocontrol activity（genetic stability of the strain,acid and alkali resistance,high temperature resistance,and ultraviolet resistance of the strain fermentation liquid）to determine the active strain;based on this,the secondary metabolites of the strain were enriched through a large amount of fermentations,and the monomer compounds were purified and separated to clarify the material basis of the active strain.The specific results are as follows:1.Identify candidate strainsThe active strain was initially selected by the plate pairing method.In order to obtain an active strain resistant to rice blast,8 strains of biocontrol strains（Stenotrophomonas maltophilia he41,Bacillus subtilis F2,Bacillus subtilis H3,Brevundimonas bullata hd13,Lysinibacillus fusiformis he14,Bacillus amyloliquefaciens D1,Bacillus amyloliquefaciens hd213,Bacillus amyloliquefaciens DEB-2）were screened against the plate.As a result,it was found that strains D1,he41, and he14 inhibited the growth of the three races of Magnaporthe grisea by more than61%,followed by H3,and the inhibition rate>57.6.%,F2 is the lowest,and the inhibition rate>54%,and the other three species have no obvious inhibitory effect.To further clarify the living strain,the active strain is fermented and cultured,and the fermentation medium is added to the basic medium to detect its effect on rice.The inhibition rate of blast fungus,the results showed that when the concentration of the fermentation filtrate reached 32%,the sterile fermentation filtrate of strains D1,he41,he14,and H3 on the mycelium inhibition rate of the three physiological races of Magnaporthe grisea reached over 90%.Significant antibacterial effect,when the concentration of the fermentation broth is 64%,the sterile fermentation filtrate of strains D1,he14,H3,and he41 can prevent the mycelia of Magnaporthe grisea from growing,that is,the inhibition rate reaches 100%,which indicates that the strains D1,H3,he41,he14 has a significant inhibitory effect on the physiological races of rice blast fungus ZB₇,ZC₁₁,and ZG₁.Evaluation of strain biocontrol activity.In order to investigate whether the candidate biocontrol strain has the potential to develop as a biocontrol agent,the strains D1,H3,he41,and he14 were genetically stabilized for activity,UV resistance,high temperature,and acid and alkali resistance.And the greenhouse control experiment of the strain he41,the results showed that（1）strains D1,he41,he14,H3were transferred and cultured 30 times,and all the biocontrol bacteria inhibited the mycelium of the three rice blast mycelia by 70%～Between 100%.（2）After treating the fermentation broth of strains D1,he41,he14,and H3 with the p H 3～11,,the bacteriostatic rate was 51%to 95%.After 6 hours of exposure to UV light（30W）,the bacteriostatic rate of the fermentation broth still reached 75%.Between 95 and 95%,after high-temperature treatment at 121°C,the bacteriostatic rate of the sterile fermentation filtrate of strain he14 was 65%to 82%,while the fermentation liquid of strains D1,he41,and H3 had no inhibitory effect on Magnaporthe grisea Tested the sterile fermentation filtrate of strains D1,he41,he14,H3 with trypsin and proteinase K,and found that some antibacterial protein substances may exist in the fermentation broth of strain D1,he41.Blast prevention and control of leaf blast,when the cell concentration is 4×10 ⁹ cfu/ml,the greenhouse control effect is slightly higher than the biological pesticide Bacillus subtilis control effect,the biological pesticide control effect was 25.27%,and the control effect of the strain he41 was 29.79%.This shows that the strains D1,he41,he14,and H3 all have the potential to develop into biocides.2.Separation of monomer compoundsIn order to clarify the antibacterial material basis of the active strain he41,the monomers of the fermentation product of the active strain he41 were separated.TLC experimental screening and repeated atmospheric silica gel column chromatography were used to isolate 4 monomer compounds and 1 Mixture.Four known compounds were identified by nuclear magnetic resonance（NMR）pop analysis and mass spectrometry（MS）analysis,namely compound 1:pyrrolo[1,2-a]pyrazine-1,4-dione,sixHydrogen-3-（2-methylpropyl）,compound2:3-isobutyl-6-methyl-2,5-piperazinedione,compound 3:3-isobutyl-6-（propane-2-yl）piperazine-2,5-dione,compound 4:3,6-bis-（2-methylpropyl）-2,5-piperazinedione.In order to determine whether the monomer compound obtained was isolated has anti-disease activity.The antifungal activity of the monomer compound and the mixture was screened,and it was found that only compound 1 had a weak antibacterial activity against rice sheath blight,and the mixture had an inhibitory effect on rice blast fungus and brown rot of kiwi fruit.This indicates that the metabolites of strain he41 need to be further explored.