Antagonistic Effect Of Bacillus Subtilis CHY On Jujube Gum Anthrax And Its Antagonistic Substance

With the improvement of environmental and food safety requirements,a new biopesticide, human and environmental friendly and good control effect,has attract a lot of scholars’s attention.For the reason of the characteristic of widely distributed,non-pathogenic, environment friendly,strong resistance,produce a variety of antimicrobial substances and broad-spectrum antimicrobial activity etc.,therefore,attract a large number of scholars to conduct research.There have a lot of biocontrol agents applied in field trials.In addition,biocontrol agents has access to broad-spectrum,highly efficient biological bacteria is still the prevention of plant protection research focus.In order to obtain an excellent antibacterial strains with broad-spectrum antimicrobial activity,this paper studied systematically Bacillus CHY,isolated from the soil of American ginseng planting in Shanxi Liuba,on plant pathogenic fungi and the antagonism of resistance,the main contents as follows:i)Bio-control bacterial CHY was identified as Bacillus amyloliquefaciens on the basis of morphological, physiological and biochemical characters as well as the sequence analysis of 16S rDNA.The results of the antagonistic tests by plate confrontation methods showed different degrees antibacterial activity to plant pathogenic fungi with broad-spectrum antimicrobial activity,the inhibited rate of Colletotrichum gloeosporioides reached 80.15%.ii)Study the antimicrobial substances of bio-control bacterial CHY,Colletotrichum gloeosporioides as target. Oxford cup experimental results show that the antibacterial substances mainly secreted into extracellular.Its sterile fermentation liquor can inhibit the hyphal growth and spore germination of the plant pathogen Colletotrichum gloeosporioides,and the sterile fermentation liquor has a certain temperature, light and alkaline stability.Further using chromatography,HPLC,mass spectrometry and SDS-PAGE to analyze.The MS result of lipopeptide showed that bio-control bacterial CHY can generates iturin and surfactin.The molecular weight of antimicrobial proteins was identified with SDS-PAGE,the result showed that only one stripe on the electrophoresis gel, between 66 kD and 45 kD,approach 55 kD.iii)Optimization of fermentation conditions of bio-control bacterial CHY based on the inhibition zone diameter and the amount of bacteria(lgcfu/mL).The result of the method of single factor showed that when the soluble starch,peptone were used as carbon, nitrogen, inorganic salts,respectively,antimicrobial activity and the amount of the strain of bacteria than other experimental groups.By orthogonal method(L9-34)experiments showed that the optimum fermentation conditions for this strain is seed age 24 h,inoculum 2%,pH 7.5, temperature 35 ℃;Response surface methodology results show that soluble starch 23.66 g/L, peptone was 12.86 g/L,the bio-control bacterial CHY sterile broth inhibition zone diameter of 25.39 mm, significantly higher than the base fermentation medium(inhibition zone diameter 5.56 mm).iiii)Plant experiments with fresh jujube to verify the control effect of bio-control bacterial CHY show that the control effect of protective group and cure group were 73.36%, 47.04 %,respectively,after treatment 7 d,the control effect of protective group was significantly higher than the cure group.The control effect of protective group and cure group were 42.31%,32.58%,respectively,after treatment 14 d.And in this process,and the defense enzyme activity of PAL,CAT,PPO,POD that the protective group than the cure group.The research shows that the bio-control bacterial CHY has the well application potential, provide basis for the biocontrol agents that low cost and can effective control of plant pathogenic.