Antagonistic Activity And Effects Of Bacillus Cereus B-02 On Botrytis Cinerea

The methods of plate culture and agar diffusion bioassay were used respectively to quantify the antimicrobial activity of Bacillus cereus B-02.The result showed that both B-02 and its filtrate had antimicrobial activity aganist Botrytis cinerea.Different concentration of B-02 filtrate played main role in the antimicrobial activity tests.In 24～48h,B-02 showed an inhibition zone against the target fungal by the expansion of its metabolites on plate.The myceliums around the inhibition zone growed abnormally and accumulated in the center of colony with the color deepened to a dark brown.Ammonium sulfate precipitation and ethanol precipitation were used to purify the antifungal substances produced by strain B-02.But the precipitation didn't show any antimicrobial activity.It proved that the antifungal substances were in culture supernatant. The B-02 suspension was heat stable and remained active after sterilization at 121℃for 30 min.In order to study the influence of fermentation conditions to the growth and antimicrobial activity of Ba.cereus strain B-02 against Bo.cinerea,the methods of spectrometer,agar diffusion bioassay and well test were used respectively to quantify the bacterium as well as its antimicrobial activity.Systematic investigations on growth curve of the strain and pH value indicated that NB broth was the optimum liquid media,in which the bacterial population reached the peak in 16h with an inhibition zone about 5mm.The best cultivating conditions were that:50mlNB medium in 250ml flask,with 1%quantity inoculation(v/v),140r/min rotation speed,32℃,pH9.0 and 36h.Under these conditions, the antimicrobial activity of B-02 were increased by 16.2%.The antagonistic mechanism was studied at the level of cellular morphology and cellular physiology.To investigate the antifungal effects of antagonistic bacteria Ba.cereus B-02 on Bo.cinerea,the normal and treated mycelial cells were observed by scanning electron microscope(SEM),transmission electron microscope(TEM) and laser scanning eonfoeal microscope(LSCM).The results showed that the configuration of mycelial cells treated with strain B-02 filtrate was destroyed and there were a lot of unmembranous materials and vacuoles in the cells.Fluorescence intensity of the treated cells and that of control group was different significantly.It indicated that the synthesis of DNA,the mitoehondrial membrane potential and the reactive oxygen quantity of Bo.cinerea hyphae were greatly influenced by Ba.cereus B-02 filtrate.