山瑞鳖源肺炎克雷伯菌的分离鉴定及其药敏特性分析

Isolation,identification and antibiotic sensitivity of Klebsiella pneumoniae from Palea steindachneri

从患病山瑞鳖的肝脏和肺脏分离纯化到1株革兰阴性杆菌GX120222,经细菌培养特性观察和VITEK 2compact全自动微生物鉴定系统对理化特性进行鉴定,结果与肺炎克雷伯菌肺炎亚种（Klebsiella pneumoniae ss pneumonia）特征相符;将该菌16SrRNA基因序列（KP091888）与GenBank中的序列进行BLAST比对并构建系统发育树,结果表明其与肺炎克雷伯菌的16SrRNA基因序列同源性最高（100%）,在系统发育树上与肺炎克雷伯菌肺炎亚种的菌株聚为一簇,且与肺炎克雷伯菌肺炎亚种标准菌株ATCC43816KPPR1（CP009208）的亲缘关系最近,进一步确定菌株GX120222为肺炎克雷伯菌肺炎亚种。人工感染试验结果显示,该菌对供试的山瑞鳖有较强致病性,半数致死量（LD50）为3.8×10-7 CFU/mL;药敏试验显示,GX120222对头孢西叮、舒普深和阿米卡星敏感,对特治星中度敏感,对所测的其他26种抗菌素耐药。上述结果表明,肺炎克雷伯菌GX120222是导致此次山瑞鳖发病死亡的病原菌,且具有多重耐药性。

A strain of gram-negative bacteria,named GX120222,was isolated from the liver and lung of diseased Palea steindachneris.GX120222 was identified as Klebsiella pneumoniae ss pneumoniae according to its cultural characteristics and physio-biochemical properties determined with VITEK 2compact automatic bacteria identification system.The 16 SrRNA gene sequence（GenBank No.KP091888）of GX120222 was compared with that of related sequences in GenBank and the phylogenetic tree was constructed on the basis of the sequences of 16 SrRNAs.The BLAST result showed that GX120222 had the highest homologies（100%）with sequence of K.pneumonia;the phylogenetic tree result revealed that this strain clustered together with strains of K.pneumoniae ss pneumonia and had the closest genetic distances with K.pneumoniae ss pneumoniae standard strain ATCC43816KPPR1,which further confirmed that GX120222 was K.pneumoniae ss pneumoniae.On the other hand,the results of artificial infection experiments revealed that GX120222 had strong pathogenicity to Palea steindachneris,while the median lethal dose（LD50）was 3.8×107 CFU/mL.By testing with thirty antimicrobial agents,GX120222 was found to be with high sensitivity to cefoxitin,sulperazone and amikacin,intermediate sensitivity to tazocin;while it was resistant to twenty-six other antimicrobial agents.These results demonstrated that K.pneumonia GX120222 was the pathogen leading to fatal disease of Palea steindachneris,and it showed multiple resistance to antibiotics.