致肾盂肾炎大肠杆菌特异性片段R049聚集性分布的研究

Clustering distribution of the specific fragment R049 of uropathogenic Escherichia coil

目的研究致肾盂肾炎大肠杆菌（UPEC）132特异性片段R049在UPEC菌株和正常人粪便分离的大肠杆菌菌株中的分布情况。方法对20株UPEC和40株粪便分离大肠杆菌采用PCR方法扩增R049片段，对R049阳性菌株检测编码5种毒力因子的基因（papC、fimH、hfy、aer、cnfl），并对其基因组DNA用XbaⅠ酶切，进行脉冲场凝胶电泳分析，继而用毛细管法将DNA片段转移至尼龙膜，利用地高辛标记的R049 ORF探针片段进行Southern杂交，分析杂交条带的分布特征。结果20株UPEC中有8株（40％）、40株粪便分离的大肠杆菌中有3株（7．5％）R049片段扩增阳性，两组差异有统计学意义（P〈0．01）。R049阳性菌株中，R049片段与5种UPEC毒力因子密切相关。Southern杂交结果表明3株粪便分离大肠杆菌R049阳性杂交带分别为150kb、15kb与240kb；8株UPEC中有6株阳性杂交带均为350kb，其余2株分别为280kb和25kb。结论UPEC132特异性片段R049在国内分离的UPEC菌株中具有共有性和聚集性分布的特征。

Objective To study the distribution of the specific fragment R049 of uropathogenic E. coli （UPEC） 132 in UPEC and fecal E. coli strains. Methods The specific fragment R049 was amplified by PCR from 20 UPEC strains and 40 fecal E. coli strains, and 5 genes encoding virulence factors （papC, fimH, hly, aer, cnfl ） were detected from fragment R049 positive strains. Pulse field gel electronphoresis （PFGE） was applied for isolating the Xba Ⅰ restriction fragments of the genomes of fragment R049 positive strains, and then Southern blot was applied for analyzing the distribution features of the positive hybridization bands by digoxin-labeled R049 ORF probe. Results The specific fragment R049 was amplified from 8 of 20 UPEC strains （40%） and 3 of 40 fecal E. coli strains （7.5%）, and statistics analysis showed significant difference （P 〈0.01 ）. The specific fragment R049 was closely related with 5 virulence factors of UPEC in the fragment R049 positive strains. Southern blot showed the sizes of positive bands were 150 kb, 15 kb and 240 kb in 3 fecal E. coli strains, 350 kb in 6 of 8 UPEC strains, and 280 kb and 25 kb in the rest two UPEC strains. Conclusion The specific fragment R049 of UPEC132 possessed the feature of clustering distribution in domestic isolated UPEC strains. [