一株黄瓜枯萎病拮抗细菌的筛选鉴定及抑菌作用初探

Screening and Identification of Antagonistic Bacteria against Fusarium Wilt of Cucumber and Preliminary Study on Its Antifungal Effect

【目的】筛选黄瓜枯萎病的高效拮抗细菌并研究其抑菌作用。【方法】采用平板对峙法从设施温室土壤样品中筛选拮抗细菌,对其中抑菌率最高的菌株进行生理生化试验及16S rDNA测序分析;利用平板倒扣法、凹玻片法以及HPLC等方法对其抑菌作用进行初步研究。【结果】共筛选得到8株拮抗细菌(XN-1~XN-8),其中抑菌率最高的为XN-3。在NCBI数据库的分析比对结果显示:XN-3与解淀粉芽孢杆菌(Bacillus amyloliquefaciens)16S rDNA序列的同源相似性最高(99%)。XN-3能产生有抑菌作用的挥发性物质。XN-3可使病原菌菌丝卷曲,长度变短,显微形态出现菌丝膨大、弯曲不均,分枝菌丝生长受阻,有厚垣孢子生成;其胞外物质使病原菌孢子萌发率降低89.01%。HPLC色谱图显示:峰A-2中的成分有明显的抑菌作用。【结论】分离得到的拮抗细菌XN-3为解淀粉芽孢杆菌,其对黄瓜枯萎病病原菌具有较强的抑菌作用,可以通过胞外物质抑制菌丝生长及孢子萌发来实现抑菌。该结果为进一步研究其生防机制奠定了基础。

[Purpose]To screen high effective antagonistic bacteria of cucumber fusarium wilt and to study its antimicrobial activity.[Methods]The antagonistic bacteria were screened from green-house soil samples by plate confrontation method,physiological and biochemical tests and 16S rDNA sequencing analysis were performed on the strains with the highest bacteriostatic rate,the antifungal effects were studied by plate inversion method,hollow glass and HPLC.[Results]A total of eight strains of antagonistic bacteria(XN-1-XN-8)were screened,and XN-3 had the highest inhibitory rate.The results of analysis and comparison in NCBI database showed that XN-3 had the highest homo-logy similarity with 16S rDNA sequence of Bacillus amyloliquefaciens(99%).XN-3 could produce volatile compounds which had antifungal effect.XN-3 could make the mycelia curl,shorten their length,and the microscopic morphology of mycelia expand and bend unevenly,the growth of branch-ing mycelia was blocked,and chlamydospore was generated,its extracellular substance reduced the spore germination rate by 89.01%.HPLC chromatogram showed that the components in peak A-2 had obvious antifungal effect.[Conclusion]The isolated antagonistic bacterium XN-3 was Bacillus amyloliticus,which had a strong antifungal effect on Fusarium oxysporum.It could inhibit mycelial growth and spore germination by extracellular substances,laying a foundation for further study on its biocontrol mechanism.