Prevalence of extended-spectrum β-lactamases in isolates of the Enterobacter cloacae complex from German hospitals

In 2002, 119 isolates of the Enterobacter cloacae complex were collected randomly from 11 German laboratories nationwide. Antibiotic susceptibilities were tested by disk-diffusion tests according to CLSI guidelines, and MICs were determined using Etests. PCRs were performed to amplify all TEM and SHV, and most CTX-M and OXA beta-lactamase genes. PCR products were sequenced to identify the precise extended spectrum beta-lactamase (ESBL) types. Isoelectric focusing (IEF) and PM/PML Etests were used to confirm production of the respective ESBLs. According to susceptibility tests and CLSI criteria, 49 (40%) isolates were resistant to extended-spectrum cephalosporins. Seven (5.8%) isolates were positive in at least one of the PCR assays. Sequencing identified production of TEM-1 beta-lactamase genes by three (2.9%) isolates, and ESBL genes of the CTX-M and SHV beta-lactamase families by five (4.2%) isolates. IEF confirmed the production of beta-lactamases in the expected pI ranges of the respective ESBLs, and four of the five ESBL-producers were detected using the PM/PML Etest. All ESBL-producing isolates showed co-resistance to sulphonamides.     

Risk-factors for emerging bloodstream infections caused by extended-spectrum β-lactamase-producing Escherichia coli

Risk-factors for bloodstream infections caused by extended-spectrum β-lactamase (ESBL)-producing Escherichia coli were investigated using an exploratory case-double control study in which 43 cases (70% producing CTX-M enzymes) were compared with: (i) 86 patients with bacteraemia caused by non-ESBL-producing E. coli ; and (ii) 86 hospitalised patients. Previous follow-up as an outpatient, urinary catheterisation and use of oxyimino-β-lactams or fluoroquinolones were independent risk-factors for ESBL-producing E. coli among patients with E. coli bacteraemia, and previous use of oxyimino-β-lactams or fluoroquinolones were also independent risk-factors among hospitalised patients. These findings may help in identifying patients at greater risk for bloodstream infection caused by ESBL-producing E. coli in endemic areas.     

Occurrence of extended-spectrum β-lactamase-producing Salmonella enterica in northern Spain with evidence of CTX-M-9 clonal spread among animals and humans

Among the 1233 Salmonella enterica isolates obtained in two Spanish hospitals, five isolates (0.4%) (serovars: Virchow, four; Livingstone, one) had the phenotype of an extended-spectrum β-lactamase (ESBL) producer. The genetic characterization of the ESBL of S. enterica Livingstone revealed a bla _SHV-2 gene. The bla _CTX-M-10 gene in a phage-related genetic environment was found in one S. enterica Virchow isolate, and the bla _CTX-M-9 gene within the In60 integron was found in the three remaining Virchow isolates. These three isolates presented indistinguishable or closely related pulsed-field gel electrophoresis patterns among themselves and also as compared with the two other bla _CTX-M-9-containing isolates previously obtained from animals. ESBL production is an emerging mechanism of resistance in S. enterica in the two studied hospitals.     

The use of antibiotic-containing agars for the isolation of extended-spectrum β-lactamase-producing organisms in intensive care units

MacConkey agar containing either cefotaxime 1.0 mg/L or ceftazidime 1.0 mg/L was evaluated for use in screening for extended-spectrum beta-lactamase (ESBL)-producing organisms. The media were evaluated using known ESBL-positive and -negative strains and 630 clinical specimens over a 6-month period. All Enterobacteriaceae isolated were identified and screened for ESBL production by phenotypic methods. In total, 14 ESBL-producing organisms were detected in the clinical samples. All known ESBL-positive strains were also detected. The use of both screening plates was required to detect all ESBLs.     

Extended-spectrum and CMY-type β-lactamase-producing Escherichia coli in clinical samples and retail meat from Pittsburgh, USA and Seville, Spain

Infections due to Escherichia coli producing extended-spectrum β-lactamase (ESBL) or CMY-type β-lactamase (CMY) are increasingly observed in non-hospitalized patients. The origin of these organisms is uncertain, but retail meat contaminated with E. coli may be a source. In the present study, clinical information and strains collected from patients infected or colonized with ESBL-producing and CMY-producing E. coli at hospitals in Pittsburgh, USA and Seville, Spain were investigated. Retail meat purchased in these cities was also studied for the presence of these organisms. Twenty-five and 79 clinical cases with ESBL-producing E. coli and 22 cases and one case with CMY-producing E. coli were identified in Pittsburgh and Seville, respectively. Among them all, community-acquired and healthcare-associated cases together constituted 60% of the cases in Pittsburgh and 73% in Seville. Community-acquired cases were more common in Seville than in Pittsburgh (49% vs. 13%; p   <0.001). ESBL-producing and CMY-producing E. coli isolates were commonly recovered from the local retail meat. In particular, 67% (8/12) of retail chickens in Seville and 85% (17/20) of those in Pittsburgh contained ESBL-producing and CMY-producing E. coli isolates, respectively. Among the ESBL-producing isolates, CTX-M and SHV were the most common ESBL types in both clinical and meat isolates. Approximately half of the ESBL-producing and CMY-producing E. coli isolates from meat belonged to phylogenetic groups associated with virulent extra-intestinal infections in humans. Community and healthcare environments are now significant reservoirs of ESBL-producing and CMY-producing E. coli . Retail meat is a potential source of these organisms.     

Risk-factors for acquisition of extended-spectrum β-lactamase-producing Escherichia coli among hospitalised patients

Between 1996 and 2002, 103 hospitalised patients yielding one or more clinical isolates of extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-EC) were identified. A significant increase was observed in the incidence of ESBL-EC colonisation or infection during the study period (1.65 episodes/100 000 patient-days in 1996 to 12.6 episodes/100 000 patient-days in 2002; p 0.01). Infection developed in 70 (68%) patients (75 episodes), with surgical site (44%) and urinary tract (17%) infections being the most frequent. Pulsed-field gel electrophoresis showed extensive clonal diversity among the isolates. A case-control study and multivariate analysis identified female gender (OR 2.1; p 0.01), use of a nasogastric tube (OR 3.5; p 0.001) and previous antibiotic therapy (OR 3.9; p < 0.001) as independent variables associated with acquisition of ESBL-EC. The study demonstrated a progressive increase in the number of ESBL-EC isolates in a non-epidemic setting. Most cases of ESBL-EC colonisation or infection occurred in hospitalised patients exposed to invasive procedures and antibiotic pressure.     

Bactericidal activity of three β-lactams alone or in combination with a β-lactamase inhibitor and two aminoglycosides against Klebsiella pneumoniae harboring extended-spectrum β-lactamases

Objective: To study the bactericidal activity of β-lactam antibiotics (imipenem, cefepime, cefpirome) alone or in combination with a β-lactamase inhibitor (sulbactam) in the presence or absence of aminoglycoside (amikacin or isepamicin) against Klebsiella pneumoniae strains producing extended-spectrum β-lactamases (ESBLs).
Methods: We characterized 10 strains by means of analytic isoelectric focusing and pulsed-field gel electrophoresis. The ESBLs produced by these strains were derived from either TEM (TEM-1, TEM-2) or SHV-1. The killing-curve method was used for this bacterial investigation. Bacteria (final inoculum 5×10 ⁵ CFU/mL) were incubated with antibiotics at clinical concentrations obtained in vivo.
Results: All the combinations with cefepime or cefpirome + sulbactam were bactericidal, with a 4 log₁₀ decrease being obtained within 6 h without regrowth at 24 h, whereas imipenem alone, and combinations, gave a bactericidal effect within 6 h. The two cephalosporins alone decreased the inoculum of 4 log₁₀ at 6 h but regrowth was observed at 24 h. When the aminoglycoside was added, this bactericidal effect was obtained within 3 h with amikacin and within 1 h with isepamicin.
Conclusions: Cefepime + sulbactam or cefpirome + sulbactarn may be an alternative to imipenem for the treatment of patients with ESBL-producing K. pneumoniae. Aminoglycosides are often associated in nosocomial infections due to ESBL-producing K. pneumoniae: isepamicin acted faster than amikacin, but both worked well. To conclude, it may be prudent to avoid extended-spectrum cephalosporins as single agent when treating serious infections due to ESBL-producing K. pneumoniae. Addition of a β-lactamase inhibitor such as sulbactam ± aminoglycoside is advisable to avoid failure of treatment.     

Evaluation of the Vitek-2 extended-spectrum β-lactamase test against non-duplicate strains of Enterobacteriaceae producing a broad diversity of well-characterised β-lactamases

The Vitek-2 extended-spectrum β-lactamase (ESBL) test was assessed using a collection of 94 ESBL-positive and 71 ESBL-negative non-duplicate isolates of Enterobacteriaceae. These isolates produced a wide diversity of well-characterised β-lactamases, including 61 different ESBLs, two class A carbapenemases and various species-specific β-lactamases. ESBL detection was performed using (i) the conventional synergy test as recommended by the Comité de l'Antibiogramme de la Société Française de Microbiologie, (ii) the CLSI phenotypic confirmatory test for ESBLs, and (iii) the Vitek-2 ESBL test. For Escherichia coli and klebsiellae, the sensitivity/specificity values were 97.3%/96.9% for the synergy test, 91.8%/100% for the CLSI phenotypic confirmatory test, and 91.8%/100% for the Vitek-2 ESBL test. For other organisms, the sensitivity/specificity values were 100%/97.4% for the synergy test, 90.5%/100% for the CLSI phenotypic confirmatory test, and 90.5%/100% for the Vitek-2 ESBL test. The Vitek-2 ESBL test seemed to be an efficient method for routine detection of ESBL-producing isolates of Enterobacteriaceae, including isolates producing AmpC-type enzymes.     

TEM-24 extended-spectrum β-lactamase-producing Enterobacter aerogenes: long-term clonal dissemination in French hospitals

Objective To investigate interstrain relatedness of TEM-24-producing Enterobacter aerogenes clinical strains isolated between 1993 and 1998 in 10 French hospitals from nine areas by pulsed-field gel electrophoresis (PFGE) and plasmid patterns.
Methods Fifteen TEM-24 - producing strains and a set of 16 control strains having various other antibiotic resistance phenotypes were genotyped by PFGE. Plasmid DNA from TEM-24 - producing strains and transconjugants was analyzed .
Results Analysis of Xba I macrorestriction patterns revealed only minor variations, and showed that all 15 TEM-24 - producing strains were closely related. Some isolates originating from distant areas had indistinguishable patterns . According to their clustering correlation coefficients, they were also genomically distant from the control strains . Two plasmid patterns were observed in TEM-24-producing strains, one of them in 13 of the strains. Large plasmids of 85 kb encoding TEM-24 β-lactamase were present in all isolates and, in all except one strain, could be transferred with high frequency by conjugation .
Conclusions These results confirm that the spread of the TEM-24 extended-spectrum β-lactamase in France was essentially due to the dissemination of a single clone .