BACKGROUND: Increase in Escherichia coli and Klebsiella pneumoniae isolates with extended- spectrum beta-lactamase (ESBL) have been noted recently in Korea. Current NCCLS disk diffusion test is not sensitive to detect ESBL-producing strains. We have more isolates with intermediate or resistance to cefotaxime than to ceftazidime disk test. The aim of this study was to characterize the ESBLs produced by strains isolated from clinical specimens. METHODS: E. coli and K. pneumoniae strains with cefotaxime intermediate or resistance by disk method were tested for ESBL production by double-disk synergy, transfer of resistance by conjugation, and relative hydrolysis of beta-lactams and isoelectric point (pI) of cell sonicate. The types of beta-lactamase gene were determined by PCR. RESULTS: Nine of the 10 E. coli and all of the 18 K. pneumoniae strains tested were synergy test positive. The MIC of cefotaxime ranged from 1.5 to 400microgram/mL, with the inoculum of 10(4) CFU, but was much higher with larger inoculum. Some selected isolates showed that the resistance was transferable and the size of the plasmid was approximately 39 MDa. The MIC of cefotaxime was higher than that of was ceftazidime in majority of the transconjugants. The hydrolytic activity of the sonicate higher for cefotaxime than ceftazidime. TEM- and SHV-type genes were detected by PCR and the pI of the beta-lactamase was 5.9 for 4 TEM-type and 7.8 for one SHV-type. CONCLUSION: Majority of the cefotaxime intermediate or resistant E. coli and K. pneumoniae isolated from the tertiary care hospital are TEM- or SHV- type ESBL producers. The inoculum size significantly affects the MIC value of beta-lactams for the ESBL-producing strains. The ESBL hydrolyzes cefotaxime more actively than ceftazidime, and the ESBL gene is transferable by conjugation.