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PubMed1998

Comparison of detection of extended-spectrum beta-lactamases by agar dilution method, E-test ESBL screen and double disk test.

The extended-spectrum beta-lactamases (ESBL) are derived from TEM-or SHV-enzymes. They mediate resistance to broad-spectrum beta-lactams and can cause infectious outbreaks in hospitals. Rapid recognition and diagnosis are important for the clinician to prescribe more effective treatment. In the present study, a group of 52 probable ESBL-producing Klebsiella pneumoniae and Escherichia coli having a suspected resistant antibiogram phenotype were included. The E-test ESBL screen and the double disk test were performed for these isolates for detection of ESBL-producing strains, as compared with the conventional agar dilution method. The agreement between the E-test ESBL screen or the double disk test and the conventional agar dilution method was good and the degree of agreement were 86.5% and 92.3% respectively. The results showed that both the E-test ESBL screen and the double disk test were useful and convenient for detection of ESBLs.

Agar↗
PubMed2001

[Extended-spectrum beta-lactamases (ESBLs) producing bacteria].

TEM- or SHV-type extended-spectrum beta-lactamases(ESBLs) are of clinical concern in Europe and the United States, whereas bacterial strains producing such types of ESBLs had not been reported in Japan for many years. Toho-1, a different type class A ESBL, has been reported in 1995, in which any prototypical enzyme has not been identified so far. At present Toho-1 is the major ESBL in Japan, however, SHV5 alpha has been reported in 1998, followed by TEM-26, SHV-2, and SHV12. More recently, SHV-24, a novel SHV-derived ESBL has also been found. Since Toho-1-type ESBL, AmpC-type beta-lactamase, and class B metallo-beta-lactamase have been widely found in Japan, a novel detection system for ESBLs suitable for this country should be developed.

Bacteria↗
PubMed2001

Comparison of three different methods for the presumptive detection of ESBL production in ceftazidime-resistant strains of Klebsiella pneumoniae.

Twenty-eight (28) strains of ceftazidime-resistant Klebsiella pneumoniae were isolated from blood cultures of in-patients from University Hospital, Kuala Lumpur between March 1995 and August 1996. Three methods were used to detect the production of ESBL enzymes by these strains. These three methods include the double-disc synergy test (DDST), inhibitor-potentiated disc-diffusion test (IPDD) and the E-test ESBL method. All strains could be identified as ESBL producers using the DDST method by a minimum of two beta-lactams and these included either a combination of ceftazidime and ceftriaxone with clavulanate respectively or cefotaxime and aztreonam with clavulanate respectively. Similarly using a combination of either cefotaxime and ceftriaxone with clavulanate or ceftriaxone and aztreonam with clavulanate respectively would have detected all strains as ESBL producers. The IPDD method could also detect for ESBL activity based on combinations of beta-lactam antibiotics with clavulanate respectively. All combinations of beta-lactam antibiotics could detect for ESBL activity in all the strains except a combination of either ceftazidime and aztreonam or cefotaxime and ceftriaxone with clavulanate respectively. The E-Test method using ceftazidime alone and in combination with clavulanate was found to be the most effective method in the presumptive identification of ESBL activity in all the strains.

Ceftazidime↗
PubMed2004

Antimicrobial susceptibility pattern and prevalence of extended spectrum beta-lactamase (ESBLs) producing strains of Klebsiella pneumoniae from a major hospital in New Delhi.

Extended spectrum beta-lactamases (ESBLs) are plasmid mediated enzymes capable of hydrolyzing penicillins, broad spectrum-cephalosporins and monobactams. The ESBL producing K. pneumoniae strains are being reported from around the world including India. The present study was taken up to evaluate the ESBL production and in-vitro susceptibility of K. pneumoniae isolates from a hospital. The bacterial isolates collected during 2003 included 51 K. pneumoniae biochemically confirmed isolates from 395 patients admitted in various wards of a major hospital in New Delhi. The isolates were from pus, wound, pleural fluid, urine and tracheal aspirate of patients attending respiratory, urology and burns wards. Antimicrobial susceptibility was carried out by Kirby Bauer's disc diffusion technique using NCCLS criteria. A screening of ESBL production was done by Double-disc synergy test (DDST) and using E-test ESBL strips. The frequency of resistance among K. pneumoniae for the cephalosporins (cefoxitin, cefuroxime, cefotaxime, ceftazidime, and cefepime) and non-cephalosporins (aztreonam, piperacillin, chloramphenicol and trimethoprim-sulfamethoxazole) were in the range of 39.2-88.0% and 51.0-90.2% respectively. 14 different antimicrobial resistance profiles were recognized ranging from resistance to only four (n=6, 11.7%) to as many as ten (n=9, 17.7%). Among the 51 isolates of K. pneumoniae strains, a total of 36 (70.6%) could be identified as ESBL producers, that correlates with the high frequency of multi-drug resistant K. pneumoniae The study shows alarming rise in ESBL production among K. pneumoniae strains and high rate of resistance to a wide range of cephalosporin and non-cephalosporin group of antimicrobials.

Anti-Bacterial Agents↗
PubMed2007

Analysis of plasmids encoding extended-spectrum beta-lactamases (ESBLs) from Escherichia coli isolated from non-hospitalised patients in Seville.

Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli is an emerging pathogen in the community. We investigated the specific types of ESBLs in E. coli isolates from non-hospitalised patients and the plasmids harbouring these ESBLs. Forty-eight consecutive ESBL-producing E. coli were studied. All the isolates were clonally unrelated. ESBLs were characterised by sequencing. Mobility of the undigested plasmids was investigated both by conventional electrophoresis and by pulsed-field gel electrophoresis. Southern restriction fragment length polymorphism (RFLP) experiments were carried out by digesting plasmids with EcoRI and HindIII. CTX-M-14 was the most frequent ESBL (28 isolates), followed by SHV-12 (16 isolates). Plasmids encoding CTX-M-14 appeared homogeneous both in hybridisation profile and mobility. In contrast, plasmids expressing SHV-12 were more varied in size and hybridisation patterns. Our data suggest that a single type or very closely related plasmids harbouring CTX-M-14 are spreading in the community among unrelated E. coli isolates in our area.

Blotting, Southern↗
PubMed2005

Health and economic outcomes of the detection of Klebsiella pneumoniae-produced extended-spectrum beta-lactamase (ESBL) in a hospital with high prevalence of this infection.

INTRODUCTION: Klebsiella pneumoniae is of high prevalence in hospital infections, mainly in bloodstream infections (BSI), and some produce extended-spectrum beta-lactamase (ESBL). For hospitals with a high prevalence of strains producing this enzyme, there is no reference material to show whether the use of the E-test method for their detection, which can be quite expensive, is actually required. OBJECTIVE: To evaluate the cost-benefit of the disk diffusion and E-test methods for the detection of ESBL-producing K. pneumoniae strains in hospitals where a high prevalence of this resistance mechanism in BSI is found. METHODS: One hundred and eight patients with K. pneumoniae BSI were evaluated retrospectively. ESBL-producing strains were identified by the disk diffusion method and by the E-test method. We estimated the costs of both diagnostic methods based on antimicrobial therapy adequacy. RESULTS: Fifty-two percent of K. pneumoniae infections were due to ESBL-producing strains. The disk diffusion method yielded a positive predictive value (PPV) of 94.7% (95% CI: 88.9-100%) and a negative predictive value (NPV) of 96.1% (CI 95%: 90.8-101.4%) in relation to the E-test. We evaluated cost-effectiveness, i.e., we analyzed the cost of both E-test and disk diffusion methods with carbapenem and cephalosporins, and found that the use of the disk diffusion method accounts for approximately US$3300. CONCLUSIONS: In hospitals with a high prevalence of ESBL-producing strains, the disk diffusion method can be used to detect ESBL-producing K. pneumoniae without compromising the clinical progression of patients with BSI. The E-test showed higher accuracy but this method was more expensive than the disk diffusion method. However, the use of the E-test method was demonstrated to be more cost-effective, as we evaluated cost based on antimicrobial therapy adequacy.

Academic Medical Centers↗
PubMed1997

Risk factors for faecal carriage of Klebsiella pneumoniae producing extended spectrum beta-lactamase (ESBL-KP) in the intensive care unit.

In the course of an outbreak of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KP) in an intensive care unit (ICU), we conducted active surveillance to determine the risk factors for ESBL-KP faecal colonization of patients. We used weekly rectal samples during a four-month period. ESBL-KP was found in the faeces of 72 of 188 (38%) patients, and 42 (58%) of them were colonized within the first week of admission to the ICU. The probability of remaining free of faecal colonization was less than 20% at 30 days of ICU admission. The risk factors associated with ESBL-KP faecal colonization were clinical severity score at admission (P = 0.004), arterial catheterization (P = 0.002), total parenteral nutrition (P = 0.04), urinary catheterization (P = 0.01), mechanical ventilation (P < 0.001), and previous antibiotic therapy (P = 0.04). A logistic regression analysis identified duration of urinary catheterization (OR:3.5; 95% CI 1.2-10.3) and mechanical ventilation (OR:4.6; 95% CI 1.1-19.3) as independent risk factors for ESBL-KP faecal colonization. Our results suggest that in an ESBL-KP prevalent environment, manipulations that facilitate cross-infection are the most relevant in the acquisition of the micro-organism and risk increases throughout hospitalization.

Adult↗
PubMed2005

Treatment of infections with ESBL-producing organisms: pharmacokinetic and pharmacodynamic considerations.

Susceptibility surveillance investigations have demonstrated an increased incidence of ESBL-producing Gram-negative bacilli. Case cohort studies have suggested clinical relevance associated with ESBL-producing Enterobacteriaciae infection. Yet, current laboratory reporting guidelines classify a large percentage of these organisms in the susceptible category. The regulatory agencies Clinical Laboratory Standards Institute (formly NCCLS) and EUCAST, which oversee these guidelines are in the process of re-evaluating the appropriateness of the current classification system. Pharmacokinetic and pharmacodynamic studies examine the relationship between drug exposure (pharmacokinetics), antibiotic potency (MIC), and treatment efficacy. PK/PD studies and analyses have been useful in demonstrating the relevance of increasingly less susceptible pathogens and specific emerging resistance mechanisms. Recent investigations have examined the impact of ESBL-producing Gram-negative bacilli relative to advanced generation cephalosporin pharmacokinetics. Animal model studies suggest that the pharmacodynamic target associated with efficacy in treatment of ESBL-producing organisms is the same as that in therapy against non-ESBL-producing bacteria (50% T>MIC). Simulation of human pharmacokinetics can predict the likelihood of achieving this PD target with specific cephalosporin dosing regimens. In general, the exposure from usual regimens of the advanced generation cephalosporins would not be anticipated to achieve the PD target for many of the organisms currently classified as susceptible. PK/PD analyses should be useful in re-evaluating current susceptibility breakpoints for ESBL-producing organisms and for optimising drug and regimen choice in treatment of these infections.

Animals↗
PubMed2006

Comparison of double disc and three dimensional methods to screen for ESBL producers in a tertiary care hospital.

Extended spectrum beta lactamases (ESBLs) continue to be a major problem in clinical setups world over, conferring resistance to the expanded spectrum cephalosporins. An attempt was made to study ESBL production among Enterobacteriaceae members from a tertiary care center in Chennai. A total of seventy randomly collected isolates of the family Enterobacteriaceae from a tertiary care center was studied for their susceptibility patterns to various antibiotics and detection of ESBL producers by double disc synergy (DDS) test and three dimensional test (TDT). Eighty percent of the isolates were multidrug resistant (MDR) and 20% were ESBL producers. TDT detected 85.7% whereas only 14.2% were detected by DDS. In the present study, a large number of isolates were found to be MDR and ESBL producers. TDTs were found to be better than DDS in the detection of ESBLs. Continued monitoring of drug resistance is necessary in clinical settings for proper disease management.

Anti-Bacterial Agents↗
PubMed2002

Resistance of Klebsiella pneumoniae strains producing and not producing ESBL (extended-spectrum beta-lactamase) type enzymes to selected non-beta-lactam antibiotics.

BACKGROUND: Bacteria of the Klebsiella genus may cause numerous infections in human, which are often treated with beta-lactam antibiotics. The fundamental mechanism of Klebsiella resistance to penicillins or cephalosporins involves the production of enzymes--extended-spectrum beta-lactamases (ESBLs). Because of resistance of many Klebsiella spp. strains to beta-lactams, alternative antibiotic therapy can make use of aminoglycosides and quinolones. MATERIAL/METHODS: The study analyzed the prevalence of ESBL -type enzymes among 256 Klebsiella pneumoniae strains isolated from various clinical materials collected from patients hospitalized between 1997 and 2000. ESBLs were detected by double-disk synergy test (DDST). The prevalence of strains resistant to selected aminoglycosides (gentamicin, amikacin, netilmicin) and quinolones (ciprofloxacin, norfloxacin, nalidixic acid) in the particular years was analyzed. Drug sensitivity was determined by disk-diffusion method according to the recommendations of the National Reference Center for Microbial Drug Sensitivity. RESULTS: During the analyzed time interval, a significant increase of the number of K. pneumoniae ESBL(+) strains was noted: in 1997 - 16.5% (14/85) and in 2000 - 40.4% (22/54) (p<0.001). Among the ESBL(+) strains, an increase of the number of strains resistant to the tested antibiotics, except for nalidixic acid, was demonstrated CONCLUSIONS: A statistically significant increase of multidrug-resistant K. pneumoniae strains, including strains producing ESBLs, was demonstrated in the analyzed material.

Amikacin↗
PubMed2005

[In vitro activity of cefepime against ESBL-positive clinical strains of gram-negative rods].

The aim of this study was to determine an in vitro activity of cefepime against ESBL-positive clinical strains of Gram-negative rods isolated from hospitalized patients. Experiments were performed with 100 ESBL-positive strains of Gram-negative rods isolated from clinical samples in 2004. Strains were identified with the use of automatic ATB Expression system and biochemical ID 32 GN tests (bioMdrieux sa). Extended-spectrum beta-lactamases (ESBLs) were detected by means of disc diffusion methods: the double-disc synergy test (DDST) and the diagnostic disc test (DD, Oxoid Ltd, UK). Susceptibility in vitro of ESBL producers to 4th generation--cefepime was determined with gradient diffusion method Etest (AB Biodisk, Solna, Sweden). MIC value of cefepime was assessed for each strain. Among 100 ESBL-producing strains, 94--belonged to enteric rods and 6--to nonfermentative rods. The greatest number of strains belonged to the species Serratia marcescens (27% of all strains) and next--to the species Enterobacter cloacae (21%). Fourteen strains were susceptible (S) in vitro to cefepime, 12--intermediately susceptible (I) and 74--resistant (R). Application of cefepime in a therapy of infections caused by ESBL-positive strains of Gram-negative rods highly susceptible in vitro to this antibiotic, should be considered.

Cefepime↗
PubMed1998

[Clinical significance of the determination of ESBL (extended- spectrum beta-lactamase) production by gram-negative nosocomial bacteria].

BACKGROUND: Data on the incidence of multiresistant nosocomial bacterial strains of Xanthomonas (Stenotrophomonas) maltophilia, producing in addition to metallo-beta-lactamases also new types of broad spectrum beta-lactamases (ESBL) called for analyses of the production of different types of enzymes. METHODS AND RESULTS: Nosocomial multiresistant strains of Xanthomonas (Stenotrophomonas) maltophilia isolated from patients in 1996 were tested for the production of ESBL by a double diffusion disc test. The production of different types of ESBL was revealed, in particular production of ESBL specific for inactivation of aztreonam which is reversible by the action of clavulanate. The test of ESBL production, proved suitable also for orientation as regards selection of a suitable antibiotic in the treatment of infections caused by multiresistant or naturally resistant bacteria, possibly also the possible synergic action of antibiotics may be detected. The effect of mutual action of different antibiotics with a different inhibitory action of beta-lactamase inhibitors is illustrated when the test of ESBL production is used, where the effect of aztreonam on the inhibition of growth of ofloxacin resistant colonies was proved. CONCLUSIONS: The double diffusion test of production of new beta-lactamases (EBL) in multiresistant Gram-negative nosocomial strains made it possible to detect the possible synergic action of antibiotics in combination with the action of beta-lactamase inhibitors.

Cross Infection↗
PubMed2005

Antimicrobial activity of antibiotics in combination with natural flavonoids against clinical extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae.

Extended-spectrum beta-lactamases (ESBLs) are plasmid-mediated class A enzymes commonly found in the family Enterobacteriaceae, mainly in Klebsiella pneumoniae. Flavonoids have also been reported to possess antimicrobial activity. In this study, the in vitro activities of 18 antibiotics and 12 flavonoids against 20 ESBL-producing K. pneumoniae isolates were evaluated. All of these isolates were susceptible to imipenem and cefmetazole, but were resistant to ampicillin, ampicillin/sulbactam, aztreonam, cefazolin, cefoperazone, cefotaxime, ceftazidime, ceftriaxone, cefuroxime, piperacillin and ticarcillin. Susceptibilities to amikacin, amoxicillin/clavulanate, cefoxitin, ciprofloxacin and gentamicin were variable. Myricetin, a flavonol, inhibited ESBL-producing K. pneumoniae isolates at a high minimum inhibitory concentration (MIC) (MIC(90) value 256 mg/mL), but exhibited significant synergic activity against ESBL-producing K. pneumoniae in separate combination with amoxicillin/clavulanate, ampicillin/sulbactam and cefoxitin. Because of the low-toxic nature of flavonoids, the combination of antibiotics and flavonoids is a potential new strategy for developing therapies for infections caused by ESBL-producing bacteria in the future.

Anti-Bacterial Agents↗
PubMed2001

Properties of multidrug-resistant, ESBL-producing Proteus mirabilis isolates and possible role of beta-lactam/beta-lactamase inhibitor combinations.

At our institution, isolation rates of clinical strains of ESBL-producing Proteus mirabilis increased to 8.8% of all P. mirabilis isolates during the period 1997-1999. To evaluate the susceptibility of ESBL-producing P. mirabilis strains against commonly used drugs, we studied 50 non-duplicated isolates selected on the basis of synergy between clavulanate and beta-lactams (ceftazidime, aztreonam, cefotaxime, and ceftriaxone). The presence of ESBL-coding genes was confirmed by colony hybridization with bla(TEM-1) and bla(SHV-1) probes. Minimum inhibitory concentrations of several antimicrobial agents for each isolate were obtained using the Etest method. All strains were encoding for TEM-derived enzymes. Gene sequencing showed that at least three different genes (TEM-15, TEM-20, and TEM-52) were present. These enzymes have not been previously reported in P. mirabilis. Isolates were characterized by: (a) reduced susceptibility or resistance to third- and fourth-generation cephalosporins (MIC > or = 2 mg/l), (b) resistance to piperacillin that was abolished by tazobactam (MIC > or = 256 vs. < or = 2 mg/l, respectively), (c) multiple antibiotic resistance that included gentamicin, fluoroquinolones and co-trimoxazole. Therapeutic failure and lack of eradication of ESBL-positive P. mirabilis by third-generation cephalosporins has been repeatedly observed both at our Institution and elsewhere. Piperacillin-tazobactam, as well as amikacin and meropenem appear to be important therapeutic options for infections due to multidrug-resistant, ESBL-producing P. mirabilis isolates.

Anti-Bacterial Agents↗
PubMed2002

Surveillance of methicillin-resistant Staphylococcus aureus (MRSA) and Enterobacteriaceae producing extended-spectrum beta-lactamase (ESBLE) in Northern France: a five-year multicentre incidence study.

In order to measure the incidence of methicillin-resistant Staphylococcus aureus (MRSA) and of Enterobacteriaceae producing extended-spectrum beta-lactamase (ESBLE), and to evaluate the impact of the national guidelines for multidrug-resistant bacteria (MDRB) prevention in hospitals of Northern France, a multicentre study was conducted for three months every year starting in 1996, in volunteer hospital laboratories. All clinical specimens positive for MRSA and ESBLE were prospectively surveyed. During the five-year surveillance period, the overall proportion of MRSA was 38.4% in the 28,534 strains of S. aureus, and that of ESBLE was 11.4% in the 6121 strains of Klebsiella pneumoniae and 47.7% in the 2353 strains of Enterobacter aerogenes. The overall incidence rates of clinical specimens positive for MRSA, ESBL-K. pneumoniae and E. aerogenes were 0.84. 0.05 and 0.12/1000 hospital-days (HD), respectively. In the 23 hospitals that participated in the survey every year, the proportion and incidence of ESBLE decreased. Hence, despite recommendations as for isolation precautions, MRSA remains poorly controlled and requires more effective measures.

Anti-Bacterial Agents↗
PubMed2000

Detection of extended-spectrum beta-lactamases in members of the family enterobacteriaceae: comparison of the MAST DD test, the double disc and the Etest ESBL.

A technically simple method-the MAST double disc (MDD) test-for the detection of extended-spectrum beta-lactamase (ESBL) production by bacteria is described. A wide range of ESBL, non-ESBL and Class 1 beta-lactamase-producing isolates was examined. The MDD test, which uses discs containing ceftazidime and a complementary disc containing ceftazidime and clavulanate and a second pair containing cefotaxime and cefotaxime and clavulanate was compared with the standard double disc diffusion test and an Etest method. Both the Etest and the MDD correctly identified 93% of ESBL producers. The MDD is an inexpensive alternative to current methods for the detection of ESBL production.

Anti-Bacterial Agents↗
PubMed2001

Outer membrane protein change combined with co-existing TEM-1 and SHV-1 beta-lactamases lead to false identification of ESBL-producing Klebsiella pneumoniae.

Nine isolates of Klebsiella pneumoniae, obtained from one colonized and eight bacteraemic patients on a paediatric ward, were shown to be identical by PFGE, indicating an outbreak. Screening for extended-spectrum beta-lactamase (ESBL) production using the double-disc synergy test, Etest for ESBLs and agar diffusion tests indicated ESBL production. The isolates showed reduced susceptibility to cefotaxime but not to other third-generation cephalosporins. Molecular studies revealed production of TEM-1 and SHV-1 but no ESBLs were identified. Deficiency in expression of an outer membrane protein (OmpK35) was also observed. These observations led us to postulate that the extremely low level of OmpK35 expression and the co-existence of TEM-1 and SHV-1 resulted in an increased MIC of cefotaxime and the false designation of the isolates as ESBL producers. All the infected infants were treated with either third-generation cephalosporins alone or multiple antibiotics including a third-generation cephalosporin, and recovered and were discharged without sequelae.

Anti-Bacterial Agents↗
PubMed2004

Integron-mediated ESBL resistance in rare serotypes of Escherichia coli causing infections in an elderly population of Israel.

OBJECTIVES: To identify and characterize the aetiology of an outbreak of extra-intestinal multidrug-resistant Escherichia coli infections in elderly patients in Israel. METHODS: Extended-spectrum beta-lactamase (ESBL)-producing clinical isolates of E. coli from extra-intestinal sources were tested for susceptibility to non-beta-lactam drugs, and their serotypes were determined. Restriction enzyme digestion, followed by PFGE of DNA purified from isolates, was used to classify the phylogenetic relationship between them. Plasmid DNA from five isolates of different serotypes was used to transform an E. coli laboratory strain. The plasmids were partially sequenced. RESULTS: E. coli isolates from 86 patients, mostly elderly, were shown to be positive for inhibitor-susceptible ESBLs, and more resistant to cefotaxime than to ceftazidime. Ninety-six per cent of ESBL producers were also resistant to gentamicin, and 100% to trimethoprim/sulfamethoxazole and ciprofloxacin. All isolates belonged to one of five serotypes. PFGE analysis of purified DNA yielded 17 profiles. Sequencing of plasmids isolated from the transformants identified sul1, aac(6')-Ib and bla(CTX-M-2). These genes were embedded in an integron, InS21. CONCLUSIONS: Extra-intestinal infections with ESBL-producing E. coli of different serotypes and probably mixed clonality showed a surprising homogeneity in resistance profiles, with 100% being co-resistant to ciprofloxacin and trimethoprim/sulfamethoxazole, and 96% to gentamicin. Plasmid DNA from three isolates from different serotypes contained integron InS21, previously demonstrated in Salmonella enterica from Argentina. This is the first molecular identification of an ESBL gene and integron in Israel or neighbouring geographical areas.

Aged↗