生物有机肥中微生物的分离鉴定及液体菌剂的构建

Isolation and Identification of Microorganisms in Bio-organic Fertilizers and Construction of a Liquid Microbial Agent

【目的】了解生物有机肥中微生物的种类及各种微生物之间的关系,并借此构建一种高效便捷的液体菌剂。【方法】采用平板划线法从肥料中分离出菌种,并通过菌落观察、革兰氏染色法及16SrDNA基因同源序列比对方法对其中的微生物进行鉴定,同时运用菌饼法和平板涂布法对各菌种之间的拮抗作用及培养条件进行研究。【结果】分离得到4种菌落,分别为植物乳杆菌(Lactobacillus plantarum)、枯草芽孢杆菌(Bacillus subtilis)、地衣芽孢杆菌(Bacillus licheniformis)和巨大芽孢杆菌(Bacillus megaterium),这4种菌之间均无拮抗作用。地衣芽孢杆菌:枯草芽孢杆菌:巨大芽孢杆菌:植物乳杆菌按1∶1∶1∶1添加到不同pH的牛肉膏蛋白胨培养基液体培养基中进行培养,并用稀释涂布平板法测定有效活菌数,最终得到最适pH为7.0。【结论】从生物有机肥中分离得到4种菌种,它们之间均无拮抗作用,采用等比例的菌种配比,在pH为7.0的牛肉膏蛋白胨液体培养基进行培养,各菌种都能很好地生长,从而得到一种高效的液体菌剂。

【Objective】The study was conducted to understand the species of microorganisms in bio-organic fertilizer and the relationship among various microorganisms and to construct an efficient and convenient liquid microbial agent.【Method】Bacteria were isolated from fertilizer by streak plate method, and identified by colony observation, Gram Staining Method and 16 SrDNA homologous gene sequence comparison method. At the same time, the antagonistic effects and culture conditions of bacteria were studied by using the hypha cake method and the spread-plate method.【Result】Four bacterial colonies, namely, Lactobacillus plantarum, Bacillus subtilis, Bacillus licheniformis and Bacillus megaterium, were isolated and showed no antagonistic effect. When adding B. licheniformis, B. subtilis, B. megaterium, L. plantarum at the ratio of 1∶1∶1∶1 into beef extract-peptone liquid medium of different pH for cultivation the number of living bacteria was determined by spread-coat plate method and the optimuml pH was 7.0.【Conclusion】There was no antagonistic effect among the four species of bacteria that isolated from bio-organic fertilizer. When the bacterial colonies were prepared at equal ratio and added into the beef extract-peptone liquid medium with pH of 7.0, the strains could grow well and a kind of efficient liquid microbial agent was obtained.