Raman spectroscopic analysis of the clonal and horizontal spread of CTX-M-15-producing Klebsiella pneumoniae in a neonatal intensive care unit

Nosocomial outbreaks of extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae are an increasing concern in neonatal intensive care units (NICUs). We describe an outbreak of ESBL-producing K. pneumoniae that lasted 5?months and affected 23 neonates in our NICU. Proton pump inhibitor and extended-spectrum cephalosporin exposure were significantly associated with the risk of ESBL-producing K. pneumoniae colonisation and/or infection. Thirty isolates recovered from clinical, screening and environmental samples in the NICU were studied by means of Raman spectroscopy, pulsed-field gel electrophoresis and repetitive extragenic palindromic polymerase chain reaction (rep-PCR). The Raman clustering was in good agreement with the results of the other two molecular methods. Fourteen isolates belonged to the Raman clone 1 and 16 to the Raman clone 3. Molecular analysis showed that all the strains expressed SHV-1 chromosomal resistance, plasmid-encoded TEM-1 and CTX-M-15 β-lactamases. Incompatibility groups of plasmid content identified by PCR-based replicon typing indicated that resistance dissemination was due to the clonal spread of K. pneumoniae and horizontal CTX-M-15 gene transfer between the two clones.     

Molecular epidemiology of extended-spectrum beta-lactamase-producing,fluoroquinolone-resistant isolates of Klebsiella pneumoniae in Taiwan

Strains of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KP) have emerged worldwide. Concomitant ciprofloxacin resistance with ESBL production in K. pneumoniae isolates would severely restrict treatment options. Among 39 (18.5%) of 211 ESBL-KP isolates resistant to ciprofloxacin (MIC, >/=4 micro g/ml), 37 (95%) were high level resistant (MIC, >/=16 micro g/ml). These isolates were also cross resistant to the newer fluoroquinolones, including levofloxacin, gatifloxacin, gemifloxacin, and garenoxacin (BMS 284756). Sitafloxacin was most active against these ciprofloxacin-resistant ESBL-KP isolates with MICs for 67% of the isolates being 相似文献   

First outbreak of multidrug-resistant Klebsiella pneumoniae producing both SHV-12-type extended-spectrum beta-lactamase and DHA-1-type AmpC beta-lactamase at a Korean hospital

PURPOSE: Coexistence of different classes of beta-lactamases in a single bacterial isolate may pose diagnostic and therapeutic challenges. We investigated a spread of Klebsiella pneumoniae isolates co-producing an AmpC beta-lactamase and an extended-spectrum beta-lactamase (ESBL) in a university hospital. MATERIALS AND METHODS: Over a three-month period, a total of 11 K. pneumoniae isolates, which exhibited resistance to cefotaxime, aztreonam, and cefoxitin, were isolated. These isolates showed positive to ESBLs by double disk tests. Minimal inhibitory concentrations (MICs) were determined by broth microdilution testing. All isolates were examined by isoelectric focusing, PCR and sequence analysis to identify bla(SHV) and bla(DHA), and molecular typing by pulsed-field gel electrophoresis (PFGE). RESULTS: All 11 isolates were highly resistant (MIC, > or = 128microg/ml) to ceftazidime, aztreonam, and cefoxitin, while they were susceptible (MIC, < or = 2microg/ml) to imipenem. The bla(SHV-12) and bla(DHA-1) genes were detected by PCR and sequence analysis. PFGE revealed a similar pattern in 10 of the 11 strains tested. CONCLUSION: This is the first outbreak report of K. pneumoniae in Korea which co-produced SHV-12 and DHA-1 beta-lactamase, and we suggest a clonal spread of multidrug-resistant K. pneumoniae at a hospital.     

The first major extended-spectrum beta-lactamase outbreak in Scandinavia was caused by clonal spread of a multiresistant Klebsiella pneumoniae producing CTX-M-15

Between May and December 2005, 64 multidrug-resistant isolates of Klebsiella pneumoniae were detected from patients admitted to Uppsala University Hospital. This represented a dramatic increase in ESBL-producing K. pneumoniae compared to previous years. To investigate the epidemiology and to characterize the resistance mechanisms of the isolates, a study was initiated. Antibiotic susceptibility was determined by means of the Etest and the disc diffusion method. Extended-spectrum beta-lactamase (ESBL) production was identified by clavulanic acid synergy test and confirmed with PCR amplification followed by DNA sequencing. DNA profiles of the isolates were examined with pulsed-field gel electrophoresis (PFGE). All isolates were resistant or exhibited reduced susceptibility to cefadroxil, cefuroxime, cefotaxime, ceftazidime, aztreonam, piperacillin/tazobactam, ciprofloxacin, tobramycin, and trimethoprim-sulfamethoxazole. They produced ESBL of the CTX-M-15 type, and the involvement of a single K. pneumoniae clone was shown. This is the first major clonal outbreak of multiresistant ESBL-producing K. pneumoniae in Scandinavia. The outbreak demonstrates the epidemic potential of enterobacteria containing ESBLs of the CTX-M type, even in a country with a relatively low selective pressure and a low prevalence of multiresistant bacteria.     

Epidemiological study by pulsed-field gel electrophoresis of an outbreak of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae in a geriatric hospital.

Twelve cases of infections caused by extended-spectrum beta-lactamase (ESBla)-producing Klebsiella pneumoniae were reported between August 1991 and March 1993 in the Geriatric Department of the Nimes University Hospital, where these bacterial had not been previously isolated. Restriction profiles of total genomic DNAs cleaved by XbaI and SpeI were compared by pulsed-field gel electrophoresis. The strains that were tested included the 12 isolates from K. pneumoniae-infected patients, strains recovered from rectal swabs of asymptomatic patients in the same ward, and strains isolated in other hospitals in Nîmes at the same time. The restriction profiles of the 12 isolates and those recovered from asymptomatic patients in the same ward were very similar. Over a period of more than 1 year, extended-spectrum beta-lactamases were not detected in K. pneumoniae isolates with restriction patterns different from that of the epidemic strain. It seems, therefore, that there was no transfer of a plasmid or a gene coding for ESBla to strains of K. pneumoniae that were different from the epidemic strain. At the same time, ESBla-producing K. pneumoniae isolates exhibiting restriction endonuclease profiles very different from that of the epidemic strain were isolated from other hospitals in Nîmes. None of these strains caused an outbreak. Pulsed-field gel electrophoresis, which allows precise characterization of strains beyond the species level, is a useful tool for studying the ESBla-producing K. pneumoniae strains involved in nosocomial outbreaks.     

Clonal dissemination of extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae isolates in a Korean hospital

In this study, we investigated the molecular characteristics of extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae isolates that were recovered from an outbreak in a Korean hospital. A new multilocus sequence typing (MLST) scheme for K. pneumoniae based on five housekeeping genes was developed and was evaluated for 43 ESBL-producing isolates from an outbreak as well as 38 surveillance isolates from Korea and also a reference strain. Overall, a total of 37 sequence types (STs) and six clonal complexes (CCs) were identified among the 82 K. pneumoniae isolates. The result of MLST analysis was concordant with that of pulsedfield gel electrophoresis. Most of the outbreak isolates belonged to a certain clone (ST2), and they produced SHV-1 and CTX-M14 enzymes, which was a different feature from that of the K. pneumoniae isolates from other Korean hospitals (ST20 and SHV-12). We also found a different distribution of CCs between ESBL-producing and -nonproducing K. pneumoniae isolates. The MLST method we developed in this study could provide unambiguous and well-resolved data for the epidemiologic study of K. pneumoniae. The outbreak isolates showed different molecular characteristics from the other K. pneumoniae isolates from other Korean hospitals.     

Multiple analysis of a foodborne outbreak caused by infant formula contaminated by an atypicalSalmonella virchow strain

An outbreak of food poisoning involving most autonomous Spanish communities was detected in the first half of 1994. The causative food was infant formula milk contaminated by lactose-fermentingSalmonella virchow. It was not possible to isolate the causative strain from the manufacturer's facilities. During the same period of time, there was a significant increase in lactose-non-fermentingSalmonella virchow strains compared with the same period in previous years. Simultaneously, lactose-non-fermenting strains were recovered from clinical samples from children and from some milk samples that were involved in the outbreak. Therefore, it was speculated that the outbreak might be more extensive than initially thought. The following epidemiological markers were used for typing theSalmonella virchow strains involved in the outbreak: (i) phage typing; (ii) ribotyping, using a set of 20 different endonucleases; and (iii) pulsed-field gel electrophoresis, using three different endonucleases. The most useful markers for this serotype were phage typing and pulsed-field gel electrophoresis, since ribotyping was not able to distinguish all strains tested. The results obtained revealed that the outbreak was caused by at least two strains: one presenting phage type 4-4a and pulsed-field patterns Al or A2 and L + or L–, and another presenting phage type 2 and pulsed-field patterns A1 or A2 and L + or L–. The results indicate that the outbreak was more extensive than initially thought and that the Virchow serotype is very clonal in Spain.     

Molecular epidemiology of Klebsiella pneumoniae producing SHV-5 beta- lactamase: parallel outbreaks due to multiple plasmid transfer.

Over a period of 22 months, 32 patients treated in three independent intensive care units of the Innsbruck University Hospital were infected with extended-spectrum beta-lactamase-producing members of the family Enterobacteriaceae (30 Klebsiella pneumoniae isolates, 1 Klebsiella oxytoca isolate, and 1 Escherichia coli isolate). As confirmed by sequencing of a bla gene PCR fragment, all isolates expressed the SHV-5-type beta-lactamase. Genomic fingerprinting of epidemic strains with XbaI and pulsed-field gel electrophoresis grouped 20 of 21 isolates from ward A into two consecutive clusters which included 1 of 3 ward B isolates. All six K. pneumoniae isolates from ward C formed a third cluster. Stool isolates of asymptomatic patients and environmental isolates belonged to these clusters as well. Additionally, 2,600 routine K. pneumoniae isolates from the surrounding provinces (population, 900,000) were screened for SHV-5 production. Only one of six nonepidemic isolates producing SHV-5 beta-lactamase was matched with the outbreak strains by genomic fingerprinting. Plasmid fingerprinting, however, revealed the epidemic spread of a predominant R-plasmid, with a size of approximately 80 kb, associated with 29 of the 30 K. pneumoniae isolates. This plasmid was also present in the single K. oxytoca and E. coli isolates from ward C and in three nonepidemic isolates producing SHV-5. Our results underline that strain typing exclusively on the genomic level can be misleading in the epidemiological investigation of plasmid-encoded extended-spectrum beta-lactamases. Our evidence for multiple events of R-plasmid transfer between species of the family Enterobacteriaceae in this nosocomial outbreak stresses the need for plasmid typing, especially because SHV-5 beta-lactamase seems to be regionally spread predominantly via plasmid transfer.     

Multilocus sequence typing and antigen gene sequencing in the investigation of a meningococcal disease outbreak

Multilocus sequence typing and antigen gene sequencing were used to investigate an outbreak of meningococcal disease in a university in the United Kingdom. The data obtained showed that five distinct Neisseria meningitidis strains belonging to the ET-37 complex were present in the student population during the outbreak. Three of these strains were not associated with invasive disease, and two distinct strains caused invasive disease, including several fatalities. The initial case of the disease cluster was caused by a strain distinct from that responsible for at least two subsequent cases and two cases remote from the university, which were epidemiologically linked to the outbreak. These observations were consistent with pulsed-field gel electrophoresis data, but the sequence data alone were sufficient to resolve the strains involved in the disease cluster. Interpretation of the nucleotide sequence data was more straightforward than interpretation of the fingerprint patterns, and the sequence data provided information on the genetic differences among the isolates.     

Recurrent Klebsiella pneumoniae mycotic aneurysm in a diabetic patient and emergence of an extended-spectrum beta-lactamase (CTX-M-24)-containing Klebsiella pneumoniae strain after prolonged treatment with first-generation cephalosporins for mycotic aneurysm

A 68-year-old diabetic woman suffered from mycotic aneurysm due to Klebsiella pneumoniae over her abdominal aorta; she received surgical intervention, followed by treatment with first-generation cephalosporins for 6 months. She was hospitalized again 11 months later because of another episode of mycotic aneurysm caused by K. pneumoniae on her thoracic aorta. Fingerprinting generated by pulsed-field gel electrophoresis and infrequent-restriction-site polymerase indicated K. pneumoniae isolates of the identical clonal strain were responsible for these two mycotic-aneurysm episodes. Unfortunately, nosocomial pneumonia developed at the second hospitalization; blood and purposefully sampled feces specimen cultures both grew CTX-M-24-producing K. pneumoniae, which were of the same strain and genetically nonrelated to the K. pneumoniae strain causing mycotic aneurysms earlier. This is the first report on infection due to CTX-M-24-producing K. pneumoniae. It is unclear whether the prolonged use of first-generation cephalosporins in this case selected a strain of enteric organism possessing the ESBL in question, which was capable of passing this ESBL plasmid to the K. pneumoniae strain causing the nosocomial infection. This report suggests that further observation is needed before one can draw a conclusion on the possibility of the selection of ESBL enteric organism by extensive exposure to first-generation cephalosporins.     

Characterization of a nosocomial outbreak caused by a multiresistant Acinetobacter baumannii strain with a carbapenem-hydrolyzing enzyme: high-level carbapenem resistance in A. baumannii is not due solely to the presence of beta-lactamases

From February to November 1997, 29 inpatients at Ramón y Cajal Hospital, Madrid, Spain, were determined to be either colonized or infected with imipenem- and meropenem-resistant Acinetobacter baumannii (IMRAB) strains (MICs, 128 to 256 microg/ml). A wide antibiotic multiresistance profile was observed with IMRAB strains. For typing IMRAB isolates, pulsed-field gel electrophoresis was used. For comparative purposes, 30 imipenem- and meropenem-susceptible A. baumannii (IMSAB) strains isolated before, during, and after the outbreak were included in this study. The molecular-typing results showed that the outbreak was caused by a single IMRAB strain (genotype A). By cloning experiments we identified a class D beta-lactamase (OXA-24) encoded in the chromosomal DNA of this IMRAB strain which showed carbapenem hydrolysis. Moreover, the outer membrane profile of the IMRAB strain showed a reduction in the expression of two porins at 22 and 33 kDa when compared with genetically related IMSAB isolates. In addition no efflux mechanisms were identified in the IMRAB strains. In summary, we report here the molecular characterization of a nosocomial outbreak caused by one multiresistant A. baumannii epidemic strain that harbors a carbapenem-hydrolyzing enzyme. Although alterations in the penicillin-binding proteins cannot be ruled out, the reduction in the expression of two porins and the presence of this OXA-derived beta-lactamase are involved in the carbapenem resistance of the epidemic nosocomial IMRAB strain.     

Characterization of a hospital outbreak of imipenem-resistant Acinetobacter baumannii by phenotypic and genotypic typing methods.

During a 13-month period, 31 patients hospitalized primarily in two intensive care units (ICUs) were either colonized or infected by imipenem-resistant Acinetobacter baumannii. Typing of the isolates by three methods (antibiotyping, biotyping, and pulsed-field gel electrophoresis) revealed that two distinct strains were involved in the first 9 cases of the outbreak and that one of these strains, which had acquired a higher level of imipenem resistance as well as resistance to all aminoglycosides, accounted for 21 of 22 cases in the second part of the outbreak. ICU environmental contamination was recognized as an important reservoir of this epidemic strain. The outbreak ceased only after the ICUs were closed for complete cleaning and disinfection.     

Rapid typing of extended-spectrum β-lactamase- and carbapenemase-producing Escherichia coli and Klebsiella pneumoniae isolates by use of SpectraCell RA

Enterobacteriaceae are important pathogens of both nosocomial and community-acquired infections. In particular, strains with broad-spectrum beta-lactamases increasingly cause problems in health care settings. Rapid and reliable typing systems are key tools to identify transmission, so that targeted infection control measures can be taken. In this study, we evaluated the performance of Raman spectroscopic analysis (RA) for the typing of multiresistant Escherichia coli and Klebsiella pneumoniae isolates using the SpectraCell RA bacterial strain analyzer (River Diagnostics). Analysis of 96 unrelated isolates revealed that RA generated highly reproducible spectra and exhibited a discriminatory power that is comparable to pulsed-field gel electrophoresis. Furthermore, adequate results were obtained for three collections of clinical isolates. RA was able to discriminate outbreak-related isolates from isolates that were not involved in an outbreak or transmission. Furthermore, it was found that the RA approach recognized clones, irrespective of the extended-spectrum β-lactamase type. It can be concluded that RA is a suitable typing technique for E. coli and K. pneumoniae isolates. Combining high reproducibility, speed, and ease-of-use, this technique may play an important role in monitoring the epidemiology of these important nosocomial species.     

High rates of plasmid-mediated quinolone resistance QnrB variants among ciprofloxacin-resistant Escherichia coli and Klebsiella pneumoniae from urinary tract infections in Korea

The aims of this study were to investigate the prevalence of qnrA, qnrB, and qnrS determinants and their molecular characteristics in ciprofloxacin-resistant isolates of Escherichia coli and Klebsiella pneumoniae from urinary tract infections (UTI) in Korea. A total of 202 nonduplicated clinical isolates of ciprofloxacin-resistant E. coli (n = 143) and K. pneumoniae (n = 59) were collected between July 2005 and August 2006. The qnr determinant screening was carried out by PCR amplification of qnrA, qnrB, and qnrS, and all positive results were confirmed by direct sequencing of the PCR products. For qnr-positive strains and their conjugants, antimicrobial susceptibility tests and pulsed-field gel electrophoresis (PFGE) were performed. The qnrB gene was detected in 41 of the 202 isolates. Among 33 of 59 (55.9%) K. pneumoniae isolates showing qnrB, 29 isolates contained the qnrB4 gene, 3 isolates had the qnrB2 gene, and 1 isolate had the qnrB6 gene. All 8 (5.6%) of the qnrB-positive isolates among the 143 E. coli strains possessed the qnrB4 gene. The minimum inhibitory concentrations (MICs) of ciprofloxacin for the transconjugants were 0.03-2 mug/ml, representing an increase of 4- to 256-fold relative to the recipient, E. coli J53Az(r). Resistances to various other antimicrobial agents also were transferred with the plasmid. The PFGE analysis revealed indistinguishable or closely related patterns in several strains and highly diverse patterns in general. QnrB variants, especially the qnrB4 subtype, are highly prevalent in ciprofloxacin-resistant E. coli and K. pneumoniae from UTI in Korea. The emergence of plasmid-mediated quinolone resistance may contribute by several means to the rapid increase in bacterial resistance to these drugs.     

Comparison of screening methods for detection of extended-spectrum beta-lactamases and their prevalence among blood isolates of Escherichia coli and Klebsiella spp. in a Belgian teaching hospital.

Using a set of 33 well-defined extended-spectrum beta-lactamase (ESBL)-producing strains of Escherichia coli and Klebsiella pneumoniae, we compared three screening methods for ESBL detection: (i) a double-disk synergy test, (ii) a three-dimensional test (both the double-disk synergy test and the three-dimensional test were performed with ceftriaxone, ceftazidime, aztreonam, and cefepime), and (iii) the Etest ESBL screen (AB Biodisk, Solna, Sweden), based on the recognition of a reduction in the ceftazidime MIC in the presence of clavulanic acid. In the double-disk test, all four indicator antibiotics scored equally and 31 of the 33 reference strains were recognized. In the three-dimensional test, ceftriaxone was the only satisfactory indicator and 30 ESBL-positive strains were detected by this antibiotic. Both systems produced two false-positive results with cefepime. With the Etest ESBL screen, 15 of 16 TEM-related and 11 of 16 SHV-related ESBL-producing strains scored positive. In 10 cases the clavulanic acid on one end of the strip interfered with the MIC determination for ceftazidime, which was read on the opposite end. This MIC had to be determined with an extra ceftazidime-only strip. No false-positive results were noted. Eighty-six blood isolates of E. coli and Klebsiella species were screened for ESBL expression by the double-disk and three-dimensional tests, both with ceftriaxone. Six strains with suspicious antibiogram phenotypes also gave positive results by the double-disk test. One E. coli strain remained undetected by the three-dimensional test. Identification of the enzymes suspected of being ESBLs by isoelectric focusing (all strains) and DNA sequencing (1 strain) confirmed the screening test results except for one Klebsiella oxytoca strain, which proved to be a hyperproducer of its chromosomal enzyme and which also had a negative Etest score. The five true ESBL producers were all confirmed by the Etest ESBL screen. Pulsed-field gel electrophoresis proved that the E. coli strains were unrelated, but that two of the three K. pneumoniae strains were closely related.     

A Swordless Knight: Epidemiology and Molecular Characteristics of the blaKPC-Negative Sequence Type 258 Klebsiella pneumoniae Clone

In June 2010, a bla(KPC)-negative, ertapenem-resistant ST-258 Klebsiella pneumoniae strain was isolated from a patient in the Laniado Medical Center (LMC). Our aims were (i) to describe its molecular characteristics and resistance mechanisms and (ii) to assess whether the bla(KPC)-negative ST-258 K. pneumoniae clone spreads as efficiently as its KPC-producing isogenic strain. In a prospective study, surveillance of all ertapenem-resistant, carbapenemase-negative K. pneumoniae (ERCNKP) isolates was conducted from June 2010 to May 2011 at LMC (314 beds) and from July 2008 to December 2010 at the Tel Aviv Sourasky Medical Center (TASMC) (1,200 beds). Molecular typing was done by arbitrarily primed PCR, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST). A total of 8 of 42 (19%) ERCNKP isolates in LMC and 1 of 32 (3.1%) in TASMC belonged to the ST-258 clone. These strains carried the bla(CTX-M-2) or the bla(CTX-M-25) extended-spectrum β-lactamase (ESBL) gene. Sequencing of the ompK genes showed a frameshift mutation in the ompK35 gene. The fate of the bla(KPC)-carrying plasmid, pKpQIL, was determined by S1 analysis and by PCR of the Tn4401 transposon, repA, and the truncated bla(OXA-9). Plasmid analysis of the ERCNKP ST-258 isolates showed variability in plasmid composition and absence of the Tn4401 transposon and the pKpQIL plasmid. In addition, the ST-258 clone was identified in 35/35 (100%) of KPC-producing K. pneumoniae isolates but in none of 62 ertapenem-susceptible K. pneumoniae isolates collected in the two centers. Our results suggest that ERCNKP ST-258 evolved by loss of the bla(KPC)-carrying plasmid pKpQIL. ERCNKP ST-258 appears to have low epidemic potential.     

Emergence of rifampin-resistant Rhodococcus equi with several types of mutations in the rpoB gene among AIDS patients in northern Thailand

The antimicrobial susceptibilities of 30 Rhodococcus equi isolates obtained from 30 patients between 1993 and 2001 in northern Thailand were investigated. The MICs showed a tendency toward resistance to various antibiotics but sensitivity to imipenem, minocycline, vancomycin, and teicoplanin (MICs, /=64 micro g/ml) to rifampin. PCR amplification and DNA sequencing of the rpoB gene and molecular typing by pulsed-field gel electrophoresis (PFGE) were performed for eight R. equi isolates from eight AIDS patients with pneumonia or lung abscess caused by R. equi between 1998 and 2001, including one low- and three high-level rifampin-resistant isolates. As a result, two high-level rifampin-resistant strains with PFGE pattern A had a Ser531Trp (Escherichia coli numbering) mutation, and one high-level rifampin-resistant strain with PFGE pattern B had a His526Tyr mutation, whereas one low-level rifampin-resistant strain with PFGE pattern C had a Ser509Pro mutation. Four rifampin-susceptible strains with PFGE patterns D and E showed an absence of mutation in the rpoB region. Our results indicate the presence of several types of rifampin-resistant R. equi strains among AIDS patients in northern Thailand.     

Comparison of pulsed-field gel electrophoresis and randomly amplified DNA polymorphism analysis for typing extended-spectrum-beta-lactamase-producing Klebsiella pneumoniae.

The incidence and transmission patterns of extended-spectrum-beta-lactamase (ESBL)-producing Klebsiella pneumoniae in patients admitted to the intensive care unit (ICU) of a university hospital were investigated over a 3-year period. K. pneumoniae isolates were characterized by antibiotic susceptibility, capsular serotyping, plasmid profiles, and pulsed-field gel electrophoresis (PFGE) of genome macrorestriction patterns with XbaI, and the results were compared with those obtained by typing with the randomly amplified polymorphic DNA (RAPD) patterns. The discriminatory power of RAPD typing was evaluated for three primers. The incidence of isolation of ESBL-producing K. pneumoniae was 2.5 cases per 1,000 admissions to the ICU versus 0.35 cases per 1,000 admissions to other units (relative risk, 7.03; 95% confidence interval, 3.89 to 12.69). Infection developed in 53% of evaluable patients. Thirty-six percent of the cases were possibly acquired in other institutions. Isolates from ICU patients were subdivided into six capsular serotypes and into four clonal groups based on antibiotype, plasmid content, and PFGE and RAPD patterns. Two clones were associated with clusters of cross-infection, involving 5 and 12 patients, respectively. Following implementation of contact isolation precautions, the incidence of nosocomial acquisition of ESBL-producing K. pneumoniae decreased from 0.55 to 0.26 cases per 1,000 admissions (P = 0.03). PFGE and RAPD analysis showed concordant results and comparable discrimination for differentiation between groups of epidemiologically related strains of ESBL-producing K. pneumoniae. More subclonal variants were determined among epidemic clones by PFGE analysis than by RAPD analysis. Both methods are useful for typing K. pneumoniae strains in epidemiological investigations, although RAPD analysis is more efficient.