Mechanism Analysis Of Endophytic Bacteria From Flaveria Bidentis Antagonist Setosphaeria Turcica

Northern Corn Leaf Blight, caused by Setosphaeria turcica, is an important maize leaf fungal disease. Its overspread in spring maize region often causes serious economiclosses. Strain HJ5, which was isolated from the stem of Flaveria bidentis by flat confrontation method, showed antagonistic effects on S. turcica and wide antifungal spectra.Strain HJ5 was identified by physiological and biochemical responses and sequence analysis of 16S rDNA. MEGA5.0 was used to create the phylogenetic tree of the 16S rDNA homologies from Bacillus species in the NCBI website. Phylogenetic tree revealed that the sequence homology between 16S rDNA sequences of HJ5 strain and that of Bacillus subtilis JF411298.1 was 99.62%. HJ5 strains was identified as B. subtilis.In order to improve the antagonistic activity to S. turcica, carbon and nitrogen sources, inorganic salts of the HJ5 fermentation media were optimized. Combined with orthogonal experiment, the media composition and fermentation conditions were also optimized. The optimum media were 1% of glucose, 1% of soy peptone, 0.01% of MnCl2 and primary pH was 8.0. The rotating speed of rocking incubator was 180 r/min. The inoculum and liquid volume in flask were 2% and 50 mL/250 mL, respectively. The cultivating temperature was 28℃. After the optimization, antagonistic activity of HJ5 was nearly doubled.Crude extracellular proteins secreted by HJ5 strain were thermal stability and not sensitive to trypsin and proteinase K. It showed stable activity in wide range of the pH, and the strongest antibacterial activity was showed at pH 8-12, but would be lost under strong acid station.Substrate mycelia of S. turcica inhibited by crude proteins were obvious distorted, its cells became shortened and expanded. the distance between specta decreased, and the mycelia led to be bead-like. Spore germination could be inhibited by the crude proteins, the vesicular structure was formed at the end of the conidium instead of the germ tube, and the cytoplasm in the vesicular structure was thin and contained a large vacuole.