Screening Of Antagonistic Bacteria Against Rice Blast In Plateau Soil And Its Antibacterial Mechanism

In order to obtain bacterial strains with antagonistic effect on rice blast fungus,we collected surface of the 5-10 cm soil from seven areas which include Mira Hill,Gongbu Jiangda,Dazi County,Ziyi Mountain,Motuo County Longda Village,Gongga County and Xigaze respectively.A total of 232 different bacterial strains were isolated by soil gradient dilution and plate coating.A strain with strong inhibitory effect on rice blast pathogens was screened by plate confinement.The type of the antagonistic bacteria were determined by physiological and biochemical tests and molecular assays.And the suitable carbon source and environmental conditions were explored,and the suitable environmental factors and nutrient factors for fermentation were studied.On the basis of this,the stability of the fermentation broth and the effect of the antagonistic bacteria and their fermentation broth on the mycelium and spores were explored.In addition,the effective inhibitory substances in the fermentation broth were extracted,and the mechanism of inhibition of antagonistic bacteria was studied.The main conclusions are as follows:A total of 232 bacteria with different morphologies were isolated from seven different sites collected from the Tibetan area.They were screened by screening and plate confrontation.It was found that one of the soil samples in Gongga County had strong resistance to rice blast.Antagonistic effect of bacterial strains,the relative antibacterial efficiency of 90%.The strain was identified as Bacillus atrophaeus by the combination of traditional physiological and biochemical identification and 16 rsDNA molecular identification.OD600 and relative antibacterial efficiency were tested,to determine the optimal growth inhibitory action of the carbon source and nitrogen source for sugar and yeast powder,respectively,the optimum growth temperature was 30 ?-35 ?,and the optimum initial pH was 7-8.The optimum fermentation conditions were as follows: the optimal fermentation temperature was 30 ?-35 ?,the best carbon source and nitrogen source are lactose and sodium glutamate,respectively.The optimum fermentation initial pH was 8.The utilization efficiency of organic nitrogen source is higher than that of inorganic nitrogen source,and the utilization efficiency of disaccharide is higher than that of monosaccharide and polysaccharide.The extraction of antimicrobial substances in fermentation broth was carried out by different methods.It was found that the effective antimicrobial substances were not single and could include macromolecules,low molecular peptides and so on,and had strong polarity.Through the detection of the stability of the fermentation broth,it was found that the antibacterial substance was insensitive to the acid-base environment and ultraviolet radiation.In the range of 40-80 ?,the temperature sensitivity is not high,the antibacterial efficiency is not changed significantly,when the temperature is higher than 100 ?,the relative antibacterial efficiency is significantly reduced.The better stability of the antagonistic bacteria determines the potential for the development of bio-pesticides.Antagonistic bacteria fermentation broth could inhibit the growth of rice blast and the spore germination.Bacteriostatic substances on the pathogen cell membrane damage,increased cell membrane permeability,leading to intracellular electrolyte,protein and sugar leakage,hinder the supply of mycelial nutrition,inhibit the development of pathogens.The treated mycelium undergoes distortion,protoplasm condensation,and enlarged fracture.On the one hand,the treated spores are delayed growth by germination time,on the other hand,the length of the buds was greatly shortened.Eventually leading to the inhibition of pathogen growth.Antagonistic bacteria through the competition of bacteria and pathogens occurrence on nutrition and space,in addition,bacteria secreted antimicrobial substances in the fermentation process,both of them combined made the pathogen growth was inhibited of.However,the types of specific antimicrobial substances and the preparation of biocontrol agents still need further exploration and research.