Rapid Genotyping Of ESBL Gene In Escherichia Coli And Drug Resistance Prediction

Objective:Extended spectrumβ-lactamases(Extended-spectrumβ-Lactamases, ESBLs) is a derivative of serine proteases, which can hydrolyze penicillin, broad-spectrum and ultra-broad-spectrum cephalosporins and monocyclicβ-lactam antibiotics,β-lactamase, leading to resistance to many antibiotics, infections have high mortality rate, and producing extended spectrumβ-lactamase spread of bacteria can easily pass, leading to widespread dissemination of antibiotic resistant bacteria, have done great harm. Therefore, the clinical need for early detection of bacteria producing ESBLs, as soon as possible to make the results of clinical drug sensitivity. The Central Hospital of Dalian, in order to understand Escherichia coli producing extended spectrumβ-lactamases and ESBLs bacteria drug susceptibility genotype, the use of capillary electrophoresis (CE), which compared with traditional slab gel electrophoresis, with the separation speed, high resolution advantages, can use this technique for detection of ESBLs producing E. coli genotypes, gas chromatography (GC) has a strong ability to multi-component separation, mass spectrometry (MS) has a strong qualitative characteristics, gas chromatography mass spectrometry with a combination of the two more significant advantages can be qualitative and quantitative analysis of metabolites in complex bacterial components. Clinical and Laboratory Standards Institute(CLSI) recommended antibiotic susceptibility test provides the sensitivity of the results, the need for at least 18 hours of training process is not conducive to the early diagnosis of anti-infective therapy. Try to use gas chromatography mass spectrometry (GC-MS) to achieve rapid identification of ESBLs producing strains of Escherichia coli susceptibility results.Methods: From November 2005-2006, Dalian Central Hospital in February identified 120 clinical isolates of Escherichia coli strains producing ESBLs, the application software to analyze the overall drug susceptibility whonet extracted plasmid of E.coli bacteria, resistant to ESBLs Drug gene six pairs of primers(TEM,SHV,CTX- M-Ⅰ,CTX-M-Ⅱ, CTX-M-Ⅲ,CTX-M-Ⅳ) do polymerase chain reaction (PCR), amplification products by capillary electrophoresis Genotyping of law to do. Escherichia coli cultivation process of adding antibiotic ceftazidime, using GC-MS method for the determination of intracellular metabolites in bacteria levels, the use of cluster analysis PCA for pattern recognition, combined with multivariate analysis, analysis of drug susceptibility of bacteria, only made This is a drug that the prediction of ceftazidime, but other drugs may be so.Results: ESBLs Escherichia coli from Dalian Central Hospital are 100% sensitive to meropenem and imipenem, relatively more sensitive to the antibiotic cefoxitin of Cephamycin class,enzyme inhibitors pull piperazine amoxicillin/tazobactam and cefoperazone/sulbactam, amikacin aminoglycosides.40 strains of Escherichia coli producing ESBLs have 1 strain TEM(2.5%),1 strain SHV(2.5%),7 strains CTX-M-Ⅰ(17.5%),7 strains CTX-M-Ⅱ(17.5%),2 strains CTX-M-Ⅲ(5%), 4 strains CTX-M-Ⅳ(10%). More than a total of 18 genotypes (46.15%), and the most was CTX-M and TEM-or SHV co-exist; addition, there is an amplification results were negative. All strains of cells can be found within the compounds of ionic compounds, at least 1,500 pieces of information, 57 strains resistant, 5 strains were false positive; 47 in the sensitive strains,8 strains were false positive; 16 in susceptible two were false positives.Overall, the research methods used basically GC-MS method can be sensitive, resistant and sensitive strains in separate.Conclusion:The established capillary electrophoresis can be more quickly identify the genotypes of Escherichia coli producing ESBLs, GC-MS methods by the intracellular bacterial content of the analysis of metabolites can reach the situation of rapid identification of bacterial susceptibility purpose Analysis of the clinical rapid susceptibility provides a new approach.