海洋解淀粉芽孢杆菌GM-1变异菌株的特性及其抑菌作用研究

Studies on Characteristics and Antibacterial Activity of Mutant Strain of Marine Bacillus amyloliquefaciens GM-1

为了明确海洋解淀粉芽孢杆菌（Bacillus amyloliquefaciens）GM-1自发突变菌株的变异与抗菌作用的关系,进一步获得优良菌种,通过形态学观察、生理生化试验及16S r DNA和gyr B基因序列分析,对变异菌株GM-1-2特性进行分析;以8种植物病菌为供试菌,采用平板对峙法和牛津杯法分别测定变异菌株GM-1-2及其发酵液的抑菌作用;采用硫酸铵沉淀法对变异菌株产生的抑菌物质进行初步分析。结果发现,变异菌株GM-1-2的菌落形态、颜色、菌体大小与原始菌株GM-1存在明显差异,但其生理生化特性、16S r DNA和gyr B序列与解淀粉芽孢杆菌的同源性等均与原始菌株相同。变异菌株GM-1-2及其发酵液对小麦根腐病菌、棉花黄萎病菌、小麦赤霉病菌、大豆菌核病菌、小麦雪腐病菌、甘蓝枯萎病菌、斑点落叶病菌、黄瓜枯萎病菌的菌丝生长均有较强的抑制作用,且抑制作用明显高于原始菌株GM-1,其中菌株的抑菌带宽度提高了0.2-9.0 mm,无菌发酵液的抑制率提高了1.1%-153.7%。变异菌株的抑菌物质主要存在于硫酸铵沉淀中,沉淀后的上清液无明显的抑菌作用。以上结果表明,变异菌株GM-1-2的抑菌活性提高,其产生的抑菌物质与原始菌株存在差异,具有潜在的开发价值。

To make sure of the relationship between the mutation and antibacterial effect of Bacillus amyloliquefaciens GM-1 spontaneous mutants and further obtain excellent strains, such methods as morphology observation of colony, physiological and biochemical reactions, 16S rDNA gene and gyrB gene sequences analysis, were applied for the properties analysis of GM-1-2 mutant. Cylinder-plate method and flat-stand method were used to evaluate the antagonistic effects on 8 plant pathogens of GM-1-2 mutant and its fermentation liquid, respectively. The antagonistic substances of GM-1-2 mutant were preliminarily studied by ammonium sulfate precipitation method. The results showed that there were obvious differences between the mutant and the original strain in the colony morphology, color and cell size. However, no obvious differences were found in the physiological and biochemical reactions and genes （16S r DNA and gyrB） sequences. The mutants showed high inhibitory effects on Bipolaris sorokiniana ,Verticillium dahliae Kleb , Fusarium graminearum , Sclerotinia sclerotiorum, Fusarium nivale （ Fr） Ces, Fusarium oxysporum f. sp. Conglutinans alternate, Fusarium oxysporum f. sp. Cucumerinum. Antibacterial activities ofthe mutant and its fermentation broth were higher than the original strain and its fermentation broth, respectively. The inhibition zone diameter of the mutant increased by 0. 2 -9. 0 m m and the inhibition rate of the fermentation liquid increased by 1.1% -153.7 % . The antibacterial substances of the mutant mainly appeared in the ammonium sulfate precipitation, and the supernatant after precipitation had no obvious inhibitory effects. In a word, the spontaneous mutant GM -1-2 produced different antibacterialsubstances from the original strain GM -1 and showed higher antibacterial activity, which had potential development value.