Cloning and Characterization of ESBLs TEM-141 / 中国药房

ABSTRACT

OBJECTIVE:To clone,prokaryotic express and characterize the TEM-type ?-lactamase produced by Enterobacter cloacae clinical isolate EC002. METHODS: The drug susceptibility of Enterobacter cloacae clinical isolate EC002 was detected by agar double dilution,double disk screening and confirmatory test were employed to detect the ESBLs. The isoelectric point (pI) of enzyme was detected by isoelectric focusing electrophoresis (IEF),the genes were coded by PCR amplification enzyme,and the prokaryotic expression and phenotype of the TEM-type ?-lactamase were detected. RESULTS: Enterobacter cloacae EC002 were resistant to most of the ?-lactamases. Positive results were noted for the phenotype identification and plasmid conjugation test. IEF showed that Enterobacter cloacae EC002 produced two ?-lactamases with pI value at 8.7 and 5.4 respectively,which were confirmed to be CTX-M-22 and a new TEM-subtype ?-lactamase by DNA sequencing,and the phenotype of the expressed enzyme of the cloned strains was non-ESBLs. The TEM-type ?-lactamase was named as TEM-141 by GenBank. CONCLUSION: The TEM-141 produced by Enterobacter cloacae EC002 was a new type of plasmid-mediated broad-spectrum ?-lactamase.