Study On Plasmid-mediated Cephalosporinase In Molecular Biology

Objective: To investigate the prevalence and genotype ofplasmid-mediated cephalosporinase (AmpC)β-lactamases andextended-spectrum-β-lactamases(ESBLs) in clinical Escherichia coliand Klebsiella pneumoniae.Methods: 15 strains of cefoxitin-resistant Klebsiella pneumoniae and24 stranis of Escherichia coli were collected from September 2005 toAugust 2006 at the Third Xiangya Hospital, Central South University.Plasmid DNA was extracted and purified by Sangon Plasmid MiniKit.Genotypes and distribution of AmpC and ESBLs were determined bypolymerase chain reaction (PCR) with type-specific primer includingAmpC and ESBLs and sequence analysis. Conjugation experiment wasused to study the transfer of cefoxitin resistance. The homology of theisolates was determined by pulsed field gel electrophoresis (PFGE). Theminimal inhibitory concentrations (MICs) on AmpCβ-lactamases-producing strains were determined by standard agar dilution.Results: The prevalence of plasmid-mediated AmpC enzyme amongKlebsiella pneumoniae isolates was 8.34％(5/60), and that ofEscherichia coli was 8.92％(14/157). These 5 strains of Klebsiellapneumoniae produced DHA-1 AmpC enzyme, and they also producedTEM-1 broad-spectrum enzymes; 4 of them produced CTX-M-3 and 1produced SHV-83 ESBL. These 13 strains of Escherichia coli produced CMY-like AmpC enzymes; one produced DHA-1 AmpC enzyme. Onestrain of Klebsiella pneumoniae and 3 strains of Escherichia coli couldtransfer cefoxition resistance to the recipient. PFGE found these 19strains came from a variety of different clones.Conclusion: In our hospital, 5 strains of Klebsiella pneumoniaeand one strain of Escherichia coli produced DHA-1 AmpC enzyme; 13strains of Escherichia coli produced CMY-like AmpC enzymes; They alsoproduced TEM-1 and CTX-M-3 or SHV-83 ESBL. Plasmid-mediatedcephalosporinase isolates had been found in our hospital, and they couldtransfer resistance to other isotlates. Sensitivity test shows that they areresistant to the most new broadβ-lactamas but sensitive to imipenem.PFGE proved that they originated from different clones.