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Screening, Identification and Fermentation Conditions Optimization of Antagonistic Strain 6-59 of Bacillus megaterium against Ceratosphaeria phyllostachydis

毛竹枯梢病拮抗细菌分离鉴定及其拮抗物质


从采集的土样中筛选到对毛竹枯梢病致病菌竹喙球菌具有拮抗作用的菌株78株,通过复筛选出1株拮抗活性较高的拮抗菌株6-59。通过形态观察、生理生化试验及16S rDNA的序列分析鉴定该菌株为巨大芽孢杆菌。经有机溶剂萃取和盐析法初步推断该菌株的拮抗物质为蛋白类物质。通过单因素试验和正交试验对菌株6-59摇瓶发酵条件进行优化,最终确定其最佳发酵条件为:蔗糖 2%、蛋白胨 3%、MgSO4·7H2O 0.03%、KCl 0.02%、种龄18 h、培养基初始pH 7.0,250 mL三角瓶装液量50 mL、接种量4%、发酵时间72 h。在此条件下培养得到的菌株6-59的发酵液,其抑菌圈直径可达25.8 mm。

The 78 strains were collected from the soil and were prescreened to have antagonistic activity against Ceratosphaeria phyllostachydis. A strain named 6-59 with a relatively higher antagonistic activity was obtained via re-screening. Then the strain 6-59 was characterized by morphological and culture features observation, physiological and biochemical measurements and 16SrDNA sequence analysis. Based on the data, the strain was initially identified as Bacillus megaterium. Through organic solvent extracting and (NH42SO4 salting out to the fermentation liquid, the antagonistic substance produced by the stain 6-59 was inferred to a protein. Then the fermentation conditions of the strain 6-59 was studied via single factor experiments and orthogonal experiments, and the optimal shaking flask fermentation condition was determined as follows: a medium composed of 2% sucrose, 3% peptone, 0.03% MgSO4·7H2O, 0.02% KCl, seed age 18 h, initial pH of the medium 7.0, medium 50 mL in a 250 mL flask, inoculum size 4% and fermentation time 72 h. In this optimized conditions, the inhibition zone diameter of the fermentation liquid could reach to 25.8 mm.


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