Abstract: The present study aimed to isolate crude protein, crude lipopeptides, and crude polysaccharides from the cell-free supernatant of Bacillus velezensis A4 through ammonium sulfate precipitation, acid precipitation, and alcoholic extraction. The antimicrobial activities of the crude extracts were investigated through plate confrontation and kiwifruit inoculation experiments. The antibacterial active substances in components were analyzed by liquid chromatograph-mass spectrometer (LC-MS) and SDS-PAGE. The results indicated that crude protein and lipopeptides possessed substantial inhibitory effects on four distinct types of kiwifruit soft rot pathogens. In vitro inhibition assays showed that crude protein had the most potent inhibitory ability against $ Epicoccus\ nigrum $, while crude lipopeptides demonstrated the most significant inhibitory effect on Alternaria alternata. Furthermore, crude protein and lipopeptides effectively suppressed the expansion of kiwifruit lesions caused by pathogens. The LC-MS analysis revealed that the active constituents present in crude lipopeptides consisted of surfactants, iturin, and fengycin, and the results obtained from SDS-PAGE demonstrated that the substances that exhibited antibacterial activities in the crude protein were two specific protein molecules that weighed 20 kDa and 40 kDa. This study provides theoretical basis and technical support for the use of Bacillus velezensis in preventing and controlling plant pathogenic fungi.
Keywords: Bacillus velezensis; cell-free supernatant; antimicrobial substances; isolation
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