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为探究解淀粉芽胞杆菌Bacillus amyloliquefaciens HRH317菌株所产抗菌蛋白物质对禾谷镰孢菌Fusarium graminearum的抑制机理,采用硫酸铵分级沉淀菌株发酵液,DEAE Sepharose Fast Flow柱和Sephadex G-200柱层析分离纯化抗菌蛋白,聚丙烯酰胺凝胶电泳检测抗菌蛋白分子量;并采用平板计数法和扫描电子显微镜观察病原菌孢子萌发和菌丝生长情况。结果表明,硫酸铵饱和度为50%时HRH317菌株发酵液的抑菌活性最大,抑菌圈直径可达2.02 cm,显著大于其它饱和度处理;2次柱层析可纯化出抗菌蛋白,分子量约为36.0 k D。禾谷镰孢菌经该抗菌蛋白处理后12h,病原菌孢子萌发率为55.8%,显著低于对照,且2~12 h间其孢子萌发率平均下降了57.3%;该抗菌蛋白还可造成病原菌菌丝变细、畸形与扭曲,且末端膨大等现象。表明解淀粉芽胞杆菌HRH317菌株产生的抗菌蛋白对禾谷镰孢菌孢子萌发和菌丝发育具有较强的抑制作用。
To investigate the inhibitory mechanism of antimicrobial protein produced by Bacillus amyloliquefaciens HRH317 strain on Fusarium graminearum, ammonium sulfate fractionated fermentation broth, DEAE Sepharose Fast Flow and Sephadex G-200 column chromatography Purified antimicrobial protein and polyacrylamide gel electrophoresis were used to detect the molecular weight of the antimicrobial protein. The spore germination and mycelium growth of the pathogen were observed by plate counting and scanning electron microscopy. The results showed that the antibacterial activity of HRH317 fermentation broth was the largest when the saturation of ammonium sulfate was 50%, and the diameter of inhibition zone reached to 2.02 cm, which was significantly greater than that of other saturation treatment. The antibacterial protein was purified by 2 column chromatography, 36.0 kD Fusarium graminearum 12h after the antibacterial protein treatment, the spore germination rate of pathogenic bacteria was 55.8%, significantly lower than the control, and 2 to 12 h spore germination rate decreased by an average of 57.3%; the antibacterial protein can also cause pathogenic bacteria Silk thinning, deformity and distortion, and the end of swelling and so on. The results showed that the antibacterial protein produced by Bacillus amyloliquefaciens HRH317 had a strong inhibitory effect on the spore germination and mycelium development of Fusarium graminearum.