多药耐药肺炎克雷伯菌gyrA基因新亚型的发现

New Subtypes of GyrA gene in Multidrug-resistant Klebsiella pneumoniae Strains

目的调查多药耐药肺炎克雷伯菌中喹诺酮类耐药相关基因的存在与变异状况。方法收集绍兴市人民医院2007年10月-2009年6月住院患者标本中分离的多药耐药肺炎克雷伯菌共20株,采用聚合酶链反应及序列分析的方法分析1种染色体介导的喹诺酮类耐药相关基因(gyrA基因)和5种质粒介导的喹诺酮类耐药相关基因〔qnrA、qnrB、qnrS、aac(6′)-Ⅰb、qepA基因〕。结果 gyrA基因83位密码子17株菌(85.0%)有突变,87位密码子7株菌(35.0%)有突变,各分为4种突变方式;SX007株存在同义突变的gyrA基因新亚型,SX001、002、003、006、017株同时存在有义突变和同义突变的gyrA基因新亚型;另检出aac(6′)-Ⅰb-Cr基因2株(10.0%),qnrB基因1株(5.0%),qnrS基因4株(20.0%);qnrA和qepA基因未检出。结论该组多药耐药肺炎克雷伯菌gyrA基因突变率高,这是喹诺酮类耐药的主要原因,其他耐药基因阳性率比较低。

OBJECTIVE To investigate the distribution and variety of quinolone-resistance genes in multidrug-resistant Klebsiella pneumoniae strains.METHODS From Oct 2007 to Jun 2009,20 strains of multidrug-resistant K.pneumoniae were collected from Shaoxing People′s Hospital of China.One kind of chromosome-mediated quinolone-resistance gene（gyrA） and 5 kinds of plasmid-mediated quinolone-resistance genes（qnrA,qnrB,qnrS,aac（6′）-Ⅰb-Cr,and qepA） were analyzed by PCR and verified by DNA sequencing.RESULTS In gyrA,17 strains（85.0%） contained mutation in the 83rd coden（4 kinds of mutative modes） while 7 strains（35.0%）contained mutation in the 87th coden（4 kinds of mutative modes）.In addition,genotype of gyrA on strain SX007 was a new subtype（same-sense mutation,GenBank accession number GQ286174）,while genotypes of gyrA on strain SX001,SX002,SX003,SX006,and SX017 were 5 new subtypes（all contained missense mutation and same-sense mutation synchronously,GenBank accession number GU060624-GU060627 and GU060629）.Moreover,2 strains were aac（6′）-Ⅰb-Cr positive（confirmed by DNA sequencing and genomic comparison with sequence registered in NCBI）（10.0%）,1 strain was qnrB positive（5.0%）,and 4 strains were qnrS positive（85.0%）.However,qnrA and qepA could not be detected.CONCLUSIONS The fact that the mutation rate of gyrA gene is 85.0% in this group of K.pneumoniae which plays the main role in quinolone resistance,but the positive rates of other resistance genes are low.