Background: Among Gram-negative pathogens in Korea, the incidence of resistance to thirdgeneration cephalosporins is becoming an ever-increasing problem. This study was designed to determine the prevalence of third-generation cephalosporins-resistant Escherichia coli and Klebsiella pneumoniae isolates from patients in a tertiary care hospital in Busan, Korea, and to characterize the mechanism of resistance.

Methods: A total of 710 E. coli and 237 K. pneumoniae non-duplicate isolates were collected from patients in Kosin Medical Center in 1999. Antimicrobial susceptibilities were tested by the disk diffusion method. Extended-spectrum β-lactamase (ESBL) production was determined by the double disk synergy test. MICs were determined by the agar dilution method. Searches for bla_TEM , bla_SHV , and bla_CMY genes in cefotaxime-resistant or intermediate isolates were performed by PCR amplification. PCR products were used to determine the sequence of resistance genes by the dideoxy-chain termination method.

Results: Seven percent of E. coli and 25% of K. pneumoniae isolates were resistant to cefotaxime. Among the isolates with decreased susceptibility to cefotaxime, 69% (18/26) of E. coli and 80% (20/25) of K. pneumoniae isolates showed positive results in double disk synergy test. Banding patterns of PCR amplification showed that the bla_TEM , bla_SHV , and bla_CMY genes were harboured by 71% (20/28), 86% (24/28) and 14% (4/28) of isolates with decreased susceptibility to cefotaxime, respectively. Seventy-one percent (20/28) of the isolates contained more than two types of βlactamase genes. Nucleotide sequence analysis of PCR products revealed that bla_SHV-12 and bla_TEM-1b were the dominant types of β-lactamase gene. In addition, we also identified bla_TEM-52 , bla_SHV-5 , and a new ESBL gene named bla_TEM-17b .

Conclusions: Third-generation cephalosporins-resistant E. coli and K. pneumoniae are wide spread in Kosin Medical Center, Busan, Korea. Most of the isolates with decreased susceptibility to cefotaxime had bla_TEM and/or bla_SHV , and some isolates harboured bla_CMY genes that may confer resistance against cephamycins. The spread of these β-lactamase genes could compromise the future usefulness of third-generation cephalosporins for the treatment of infections caused by E. coli and K. pneumoniae. (Korean J Clin Microbiol 2002;5(1):06-14)