大肠埃希菌临床菌株优势β-内酰胺酶基因型及其诱导表达与抑制的研究  

Predominant β-1actamase genotypes of Escherichia coli isolates and induction and inhibition mechanisms of β-1actamase gene expression

作　　者：谈潘莉[1] 汪浙炯[1] 孙爱华[2] 严杰[3] 赵金方[1] 

机构地区：[1]浙江中医药大学附属第一医院检验科,杭州310006 [2]浙江医学高等专科学校 [3]浙江大学医学院病原生物学系

出　　处：《中华流行病学杂志》2015年第5期484-490,共7页Chinese Journal of Epidemiology

基　　金：国家自然科学基金（81271893）;浙江省自然科学基金（LY12H19002）;浙江省卫生厅科研项目（2011KYA005）

摘　　要：目的了解浙江地区大肠埃希菌临床菌株优势β-内酰胺酶基因型及其携带模式、β-内酰胺类抗生素诱导β-内酰胺酶基因表达及组氨酸激酶抑制剂氯氰碘柳胺（CLO）抑制其表达的作用。方法采用微量稀释法和E—test检测大肠埃希菌临床菌株对β-内酰胺类抗生素耐药率和最低抑菌浓度（MIC）。采用PCR及其产物测序法检测大肠埃希菌耐药菌株β-内酰胺酶基因型及携带模式。采用实时荧光定量RT-PCR和β-内酰胺酶确证试验分别检测1／4MIC头孢噻肟或青霉素及CLO对大肠埃希菌耐药菌株6一内酰胺酶基因转录和表达的影响。结果浙江地区61．7％（285／462）大肠埃希菌对青霉素、氨苄青霉素、头孢西丁、头孢噻肟和头孢他啶耐药。285株耐药菌株中，TEM和CTX-M基因检出率（83．2％和75．1％）显著高于KPC、SHV和OXA基因（1．4％～10．2％）（P〈0．01），两种以上β-内酰胺酶基因携带率（68．8％）显著高于单基因（31．2％）（P〈0．01），其中61．4％菌株携带TEM＋CTX—M基因（P〈0．01）。除KPC基因外，1／4MIC头孢噻肟和青霉素能诱导89株β-内酰胺酶单基因菌株TEM、CTX．M、SHV和OXAmRNA水平迅速升高（P〈0．01），但可被50～500μg／ml CLO所抑制（P〈0．01）。100gg／ml CLO预处理后，82．8％～85．6％耐药菌株对上述抗生素敏感（P〈0．01），β-内酰胺酶检出率也从95．1％下降至16．1％（P〈0．01）。结论TEM和CTX．M是浙江地区大肠埃希菌临床菌株优势β-内酰胺酶基因型，并以TEM-1＋CTX．M为优势携带模式。低浓度头孢噻肟和青霉素可经细菌二元信号系统上调β-内酰胺酶基因表达，但可被组氨酸激酶抑制剂CLO所抑制。Objective To understand the predominant β-1actamase genotypes and their carrying modes of Escherichia coli isolates in Zhejiang province, and the effects of β-lactam antibiotics on inducing or histidine kinase inhibitor closantel （CLO） on inhibiting the expression of β-lactamase genes. Methods Micro-dilution method and E-test were applied to measure the resistant rate and minimal inhibitory concentration （MIC） in E. coli isolates against β-lactam antibiotics. PCR and sequence analysis of PCR products were conducted to detect the ^-lactamase genotypes and their carrying modes. Real-time fluorescent quantitative RT-PCR and β-lactamase confirmation test were performed to determine the influence of 1/4 MIC penicillin and cefotaxime, and CLO on the transcription and expression of D-lactamase genes in the resistant E. coli isolates. Results Among the 462 E. coli strains isolated in Zhejiang, 285 （61.7%） were resistant to penicillin, ampicillin, cefoxitin, cefotaxim and ceftazidime. In the 285 resistant isolates, the detection rate of TEM or CTX-M β-1actamase gene （83.2% or 75.1% ） was significantly higher than that of KPC, SHV or OXA β-lactamase gene （1.4%-10.2%） （P〈0.01） and the carrying rate of two or more [3-1actamase genes （68.8%） was also significantly higher than that of single 13-1actamase gene （31.2%） （P〈0.01）, and 61.4% of the resistant isolates carded TEM q-CTX-M genes （P^0.01）. Except KPC gene, 1/4 MIC of cefotaxim and penicillin induced a rapid increase of TEM-mRNA, CTX-M-mRNA, SHV-mRNA or OXA-mRNA levels （P〈0.01）, but 50-500 μg/ml CLO inhibited these levels （P〈0.01）. After pre-treatment with 100 μg/ml CLO, 82.8%-85.6% of the resistant isolates became sensitive to β-lactam antibiotics （P〈0.01） , while the detection rate of β-lactamases was also decreased from 95.1% to 16.1% （P〈0.01）. Conclusion TEM and CTX-M are the predominant D-lactamase genotypes in E. coli isolates in Zhejiang and TEM q-CTX-M is the predominant carrying mode

关 键 词：大肠埃希菌 Β-内酰胺类抗生素 耐药性 13-内酰胺酶/基因型/表达 组氨酸激 酶/抑制剂 

分 类 号：R446.5[医药卫生—诊断学]