弗氏柠檬酸杆菌的分离鉴定与PCR-SSCP分析

摘要

摘要利用营养琼脂、MaC培养基从草鱼肠道中分离到3株细菌,暂时编号为TC-1、TC-2和TC-3,通过形态学观察、生理生化特征、药敏试验、动物试验、构建系统发育进化树及PCR-SSCP分析等系统鉴定,结果表明3株菌株均为弗氏柠檬酸杆菌(Citrobacter freundii),其中TC-2菌株对小鼠、斑马鱼有致病性；3株菌均对头孢噻肟、头孢曲松、洛美沙星、诺氟沙星等多种药物敏感；系统发育分析表明,3株弗氏柠檬酸杆菌16S rDNA序列与DSM 30039模式株同源性分别为99.59%、99.47%和99.53%,且位于系统发育树的同一分支；进一步采用V3区PCR-SSCP分析结果显示弗氏柠檬酸杆菌SSCP图谱中菌株间带型存在差异.%The bacteria were isolated from intestine in grass carp ( Ctenopharyngodon idellus ) using nutrient agar and MaC medium, designated temporarily as TC-1, TC-2 and TC-3, and identified with morphological observation, physiological and biochemical characteristics, drug sensitive test, animal test, established phylogcnctic tree, as well as PCR-SSCP analysis. The results showed that all 3 strains were Citrobacter freundii (Cf). Animal tests showed that TC-2 strain was lethal to mice and zebrafish, respectively. Drug sensitivity test analysis revealed that 3 strains were sensitivity to cefotaxime, ceftriaxone, lomefloxacin, norfloxacin, and many others. The phylogenetic analysis showed that 16S rDNA sequence of the 3 strains had homoiogy to the typical strain DSM 30039 at 99. 59% , 99. 47% and 99.53% respectively, and they located on the same branch of the phylogenelic tree. Further analysis results adopting V3 region of PCR-SSCP showed there were differences among SSCP atlases of Cf strains.