一株真菌拮抗细菌Z21的筛选与鉴定及其发酵条件优化

Screen, identification and fermentation optimization of an antifungal bacterium Z21

摘要:

【背景】芽孢杆菌属的许多细菌具有抗逆性强、安全等特点，一直以来都是开发新型活性物质的研究热点。【目的】筛选对食品腐败真菌有抑制作用的细菌，将其开发为天然食品防腐剂。【方法】采用平板分离法、平板对峙法、抑制菌丝生长速率法从空气、竹子内生细菌中筛选真菌拮抗菌，通过形态、生理生化特征及16S rRNA基因序列分析等方法对其进行鉴定，利用正交试验确定其最优生长条件。【结果】筛选到一株对6种常见霉菌均有较强抑制作用的细菌Z21。Z21与甲基营养型芽孢杆菌(Bacillus methylotrophicus strain CBMB205T)的相似性最高，且形态特征和生理生化特征与CBMB205T菌株基本相符。Z21最佳发酵培养基配方和培养条件分别为：葡萄糖20.0 g/L、NaNO3 20.0 g/L、MgSO4 3.0 g/L，培养温度为32 °C，培养时间为48 h。【结论】Z21为甲基营养型芽孢杆菌(Bacillus methylotrophicus)，对黑曲霉、康氏木霉、绿色木霉、少根根霉、易脆毛霉、赭绿青霉的生长具有较强的抑制作用且抑菌效果稳定，为广谱真菌拮抗菌。

Abstract:

[Background] Many species of Bacillus with the merits of strong stress resistance and safety for human become a research hotspot in exploiting new active substances. [Objective] To isolate and screen antifungal bacteria and develop its active ingredients as a natural bio-antifungal agent of food. [Methods] These methods of plate culture, plate-confrontation and mycelia growth inhibition were used to isolate and screen antifungal bacteria from air and bamboo endophytic bacteria. Z21 was identified based on morphological, physiological, biochemical characteristics and 16S rRNA gene sequence. The growth conditions of Z21 were through orthogonal experiment. [Results] Z21 showed strong antifungal activity to 6 kinds of fungi. The 16S rRNA gene sequence of Z21 was consistent with that of Bacillus methylotrophicus CBMB205T, and the morphological characteristics and physiological and biochemical characteristics of Z21 were consistent with the CBMB205T strain. The optimum culture conditions for Z21 were 32 °C for 48 h with a medium of glucose 20.0 g/L, NaNO3 20.0 g/L and MgSO4 3.0 g/L. [Conclusion] The Z21 was identified as B. methylotrophicus, it showed strongly and stalely inhibitory activities against Aspergillus niger, Trichoderma koningii, Trichoderma viride, Rhizopus arrhizus, Mucor fragilis, and Penicillium ochrochloron. So it was an abroad-spectrum antifungal bacterium.