Korean J Nosocomial Infect Control.  2003 Dec;8(2):83-93.

Molecular Characterization of Extended-spectrum beta-lactamase(ESBL) in Clinical Isolates of Klebsiella pneumoniae

• Wie SH
• Kim SY
• Kim SI
• Kim YR
• Yoo JH
• Shin WS
• Kang MW

Abstract

BACKGROUND: ESBL-producing Klebsiella spp. are increasing worldwide, and infections with ESBL-producing organisms are usually hospital-acquired and common infections with ESBL-producing organisms include urinary tract infections, peritonitis, cholangitis, intraabdominal abscesses and nosocomial pneumonia. We studied the phenotypic and genotypic characteristics of ESBL-producing K. pneumoniae, which were isolated from the patients in St. Vincent Hospital.
METHODS
Susceptibility to antibiotic was determined by standard disk diffusion and agar dilution methods for all 22 strains of K. pneumoniae. PCR amplifications were performed with primers specific for TEM. SHY, and CMY-1 genes, and the DNA of the amplified products were sequenced. Total DNA was extracted from the isolates restricted with XbaI, and fingerprinted using pulsed-field gel electrophoresis. Crude preparations of beta-lactamase were obtained by sonications and used for characterization of beta-lactamase by isoelectric focusing.
RESULTS
The MIC90 values for ceftazidime and aztreonam were 128 microgram/mL and 64 microgram/mL, respectively. The MIC90 values for cefotaxime and sulperazon were 8 microgram/mL and 4microgram/mL, respectively, and that for cefoxitin was 1.0 microgram/mL, which is much lower than the value for third-generation cephalosporins. The MICs for ciprofloxacin. cefepime, and imipenem were less than 1 microgram/mL for all organisms, which is within the susceptible range. Isoelectric focusing studies demonstrated three beta-lactamases with pls of 5.. (TEM-1), 7.6 (SHV-2a), and 8.2 (SHV-12). The presence of blaSHV and blaTEM genes was confirmed by specific PCRs and DNA sequencing analysis. but blaCMY-1 genes were not found. According to DNA sequencing analysis, 21 K. pneumoniae strains produced SHV-12 ESBL and one strain produced SHV-2a ESBL.
CONCLUSIONS
Our results suggest that the resistance of K. pneumoniae from clinical isolates to extended spectrum beta-lactam antibiotics may be due to the production of SHY-type ESBL. This approach in detecting and characterizing ESBL will be valuable for the control of infection and antibiotics use in medical institution.

Keyword

Extended-spectrum beta-lactamase; Klebsiella pneumoniae; TEM; SHV

MeSH Terms

Abscess
Agar
Anti-Bacterial Agents
Aztreonam
beta-Lactamases
Cefotaxime
Cefoxitin
Ceftazidime
Cephalosporins
Cholangitis
Ciprofloxacin
Dermatoglyphics
Diffusion
DNA
Electrophoresis, Gel, Pulsed-Field
Humans
Imipenem
Isoelectric Focusing
Klebsiella pneumoniae*
Klebsiella*
Peritonitis
Pneumonia
Polymerase Chain Reaction
Sequence Analysis, DNA
Sonication
Urinary Tract Infections
Agar
Anti-Bacterial Agents
Aztreonam
Cefotaxime
Cefoxitin
Ceftazidime
Cephalosporins
Ciprofloxacin
DNA
Imipenem
beta-Lactamases