猪圈发酵床中细菌的分离鉴定及其特征研究

Isolation Identification and Characteristics of Bacteria from Sawdust Bed in Pigpen

目的:研究猪圈发酵床分离细菌的系统发育树及生理生化特征。方法:采用R2A培养基对猪发酵床上层垫料进行分离菌株培养,将培养得到的单个菌落培养于固体培养基中,获得纯培养菌株;肉眼观察培养基中细菌的生长情况,电镜下观察细菌的形态、大小和排列特点,采用16S rRNA基因序列扩增、测序,邻接法构建16S rRNA序列的系统发育树,将分离到的猪圈发酵床中的细菌与文献中获知的初期发酵床优势细菌的标准菌株进行生理生化特征的比对。结果:在R2A培养基中分离得到3株细菌SZDIS-1-1、GZDIS-1和SZDIS-2,鉴定为微杆菌属细菌;与文献中获知的4种标准菌株一样,培养得到的3种细菌镜下均为杆状,过氧化氢酶、吲哚试验均为阴性,能够利用麦芽糖作为生长碳源;药敏试验显示,分离得到的3株细菌对阿米卡星、头孢呋辛、利福平、妥布霉素、青霉素和环丙沙星等抗生素比较敏感,其中头孢呋辛抑菌效果最佳。结论:在猪圈发酵床首次分离得到3株菌株,并构建系统发育树。

Objective： To investigate phylogenetic tree and physiological-biochemical characteristic of microbial identification from sawdust bed in a pigpen. Methods： The bacteria were isolated and cultured by R2 A solid media,and cultured individual bacteria colony was further cultured in solid culture medium to obtain pure culture strains. Observing bacteria growth condition in culture medium with naked eye,morphological features,size and sequencing features were observed under electron microscope. Characteristics of the cultured bacteria was analyzed by using 16 S rRNA gene NASBA and sequencing,and buiding phylogenetic tree of 16 S rRNA sequence by neighbor-joining method. Comparing physiological-biochemical characteristics of isolates in the sawdust bed of pigpen and the dominant strain of the type species in reference. Results： Three bacteria strains isolated on the R2 A solid medium,SZDIS-1-1,GZDIS-1 and SZDIS-2,were identified as Microbacterium by gene sequences analysis. Identical with the 4 standard strains from reference,the three cultured bacteria was in the form of rod under microscope. Peroxidase and Benzazole experiment showed negative,it could use malt sugar as growth source. The drug sensitive test showed that the three strains were sensitive to Amikacin,Cefuroxime,Rifampicin,Tobramycin,Penicillin,Ciprofloxacin,of which Cefuroxime showed the best bacteria inhibition effect. Conclusions： Three strains isolated from sawdust bed in pigpen established phylogenetic tree.