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【目的】从发病蕉园中的健康香蕉根际筛选能有效抑制香蕉枯萎病病原菌生长的拮抗菌,进一步研究其抑菌机理。【方法】应用双层平板初筛和平板对峙实验复筛具有抑菌效果的拮抗菌;经生理生化试验、16S rRNA基因测序和特异引物扩增对拮抗菌进行鉴定;酸沉淀方法提取拮抗菌发酵液抑菌物质粗提液,基于比色法和HPLC测定粗提液对菌丝蛋白质含量、脂质过氧化、麦角甾醇和果胶酶活性的影响。【结果】筛选获得两株拮抗细菌H-2和H-7,初步鉴定为枯草芽孢杆菌和解淀粉芽孢杆菌,Gen Bank登录号分别为KX791428和KX791430;温室盆栽试验显示,两株拮抗菌对香蕉枯萎病的生防效率分别为59.1%和53.0%;H-2和H-7粗提液处理病原菌菌丝后,因脂质过氧化产生的丙二醛含量显著增加,分别达0.55μmol/L和0.48μmol/L;而蛋白含量、麦角甾醇含量和果胶酶活性均显著下降,其中H-2处理的抑制幅度更大,三项指标分别为0.15 mg/g、1.31 mg/g和0.008 7 U/m L,显著低于对照的0.25 mg/g、1.96 mg/g和0.035 U/m L。【结论】从健康香蕉根际筛选到两株拮抗细菌,两者可能通过增强病原菌菌丝的脂质过氧化和降低细胞代谢产物合成的方式抑制病原菌生长,可为两株拮抗菌的生防应用提供理论依据。
【Objective】 The objective of this study was to screen antagonistic bacteria that could effectively inhibit the growth of banana wilt pathogen from the healthy banana rhizosphere in the banana plantation and to further study its antibacterial mechanism. 【Method】 Antagonistic bacteria with antibacterial effect were screened by double-layer plate screening test and plate confrontation test. Antagonistic bacteria were identified by physiological and biochemical tests, 16S rRNA gene sequencing and specific primer amplification. Antacids were isolated by acid precipitation Liquid antibacterial material crude extract, based on colorimetric and HPLC determination of the crude protein content, lipid peroxidation, ergosterol and pectinase activity. 【Result】 Two strains of antagonistic bacterium, H-2 and H-7, were screened and identified as Bacillus subtilis and Bacillus amyloliquefaciens respectively. Gen Bank accession numbers were KX791428 and KX791430, respectively. Pot experiments in greenhouse showed that two antagonistic fungi wilted banana The biocontrol efficiencies were 59.1% and 53.0%, respectively. After H-2 and H-7 crude extracts were treated with the mycelium of pathogenic bacteria, the content of malondialdehyde (MDA) produced by lipid peroxidation increased significantly by 0.55μmol / L and 0.48μmol / L, while the contents of protein, ergosterol and pectinase decreased significantly, and the inhibitory effect of H-2 treatment was much greater with 0.15mg / g, 1.31 mg / g and 0.008 7 U / m L, significantly lower than the control 0.25 mg / g, 1.96 mg / g and 0.035 U / m L. 【Conclusion】 Two strains of antagonistic bacteria were screened from healthy banana rhizosphere. Both of them may inhibit the growth of pathogenic bacteria by enhancing the lipid peroxidation of pathogenic mycelium and reducing the synthesis of cellular metabolites, which may be the biocontrol application of two antagonistic bacteria Provide a theoretical basis.