Extended-spectrum Beta-lactamases Test Research Articles

Prevalence of blaCTX-M and blaTEM Genes in Cefotaxime-Resistant Escherichia coli Recovered from Tertiary Care at Central Nepal: A Descriptive Cross-Sectional Study.

Urinary tract infections (UTIs) are highly prevalent globally, and various antibiotics are employed for their treatment. However, the emergence of drug-resistant uropathogens towards these antibiotics causes a high rate of morbidity and mortality. This study was conducted at the Microbiology Laboratory of Grande International Hospital from November 2021 to May 2022 and aimed to assess the prevalence of UTI caused by Escherichia coli and their antibiotic susceptibility pattern with a focus on extended-spectrum beta-lactamases (ESBLs) and the prevalence of two genes (blaCTX-M and blaTEM) in cephalosporin-resistant E. coli. Altogether, 1050 urine samples were processed to obtain 165 isolates of E. coli. The isolates were identified by colony morphology and biochemical characteristics. Antimicrobial susceptibility tests (ASTs) were determined by the Kirby-Bauer disk diffusion method, and their ESBL enzymes were estimated by the combined disk method (CDM). Two ESBL genes (blaCTX-M and blaTEM) were investigated by polymerase chain reaction (PCR) in cefotaxime-resistant E. coli. Among the 1050 urine samples that were processed, 335 (31.9%) were culture-positive with 165 (49.2%) identified as E. coli. The age group ≥60 years (30.3%) had greater susceptibility to bacterial infections. AST revealed that meropenem was highly effective (95.7% susceptibility), while ampicillin showed the least sensitivity (42.4%). Among the E. coli isolates, 86 were multidrug resistant (MDR) and 10 were extensively drug resistant (XDR). Of these, 46 MDR (96%) and 2 XDR (4%) were ESBL producers. The prevalence of ESBL genes (blaCTX-M and blaTEM) was 49.3% and 54.8%, respectively. The overall accuracy of CDM as compared to PCR for the detection of the blaCTX-M gene was 55.26%. The prevalence of MDR E. coli harboring the blaCTX-M and blaTEM genes underscores the imperative role of ESBL testing in accurately identifying both beta-lactamase producers and nonproducers.

Characteristics and susceptibility pattern of catheter-associated urinary tract infections (CAUTI) bacteria in Indonesia: A study in a national reference hospital of Sumatra region 2020-2021.

Catheter-associated urinary tract infection (CAUTI) is defined as a urinary tract infection associated with catheter placement for more than two consecutive days. Hence, antibiotic resistance in the context of CAUTIs represents a substantial challenge. The aim of this study was to present the characteristics of patients with CAUTI and the susceptibility pattern of CAUTI bacteria in the national reference hospital of the Sumatra region of Indonesia. A cross-sectional study was conducted at H. Adam Malik General Hospital, Medan, Indonesia, from 2020 to 2021, using a total sampling. All CAUTI patients included were on catheterization and diagnosed based on the Centers for Disease Control and Prevention (CDC) guidelines. The patient's urine culture and antibiotic susceptibility test were carried out on the patient's admitted urine sample for further assessment. Identification of bacteria, antibiotic susceptibility test, and the extended-spectrum beta-lactamase (ESBL) test for Escherichia coli and Klebsiella pneumoniae were conducted using the VITEK-2 Compact. A total of 74 CAUTI patients were included in the study, 59.5% were female, 54.1% were 46-65 years old, and a third had cardiovascular disease comorbidities (33.8%). A total of 83 CAUTI-associated bacteria were isolated. The majority were Gram-negative bacteria (74.7%), and the most bacteria isolated was E. coli (31.3%), followed by K. pneumoniae, Enterococcus faecalis, Acinetobacter baumannii, and Enterococcus faecium. The ESBL test was positive mostly in K. pneumoniae (100%) and E. coli (76.9%). CAUTI-associated E. coli was susceptible to tigecycline, meropenem, ertapenem, nitrofurantoin, and gentamicin. The isolated K. pneumoniae was susceptible to tigecycline, meropenem, ertapenem, and amikacin. While E. faecalis showed susceptibility to tigecycline, nitrofurantoin, vancomycin, imipenem, linezolid, ampicillin, piperacillin/tazobactam, amoxicillin/clavulanic acid, ampicillin/sulbactam, and piperacillin.

Prevalence of carbapenem-resistant and extended-spectrum beta-lactamase-producing Enterobacteriaceae in a teaching hospital in Ghana.

Carbapenem-resistant Enterobacteriaceae (CRE) and Extended-spectrum beta-lactamase (ESBL) production among Gram-negative Enterobacteriaceae is an increasing global challenge due to the high morbidity and mortality associated with their infections, especially in developing countries where there are little antibiotic treatment options. Despite these challenges, few studies in Ghana have described the burden of CRE. Therefore, this study aimed to determine the prevalence of carbapenem-resistant Enterobacteriaceae isolated from patients at the Cape Coast Teaching Hospital (CCTH) in the Central region of Ghana. Enterobacteriaceae isolates were collected from April to July 2019 at the bacteriology unit of CCTH using a consecutive sampling method. Isolates were identified by standard microbiological techniques and confirmed using API 20E. Kirby Bauer disc diffusion method was used to determine the antibiogram of isolates. Isolates were also subjected to ESBL testing using the single-disc combination method. Carbapenem-resistant isolates were identified by the Kirby Bauer disc diffusion method and then examined genotypically for the presence of blaKPC-1, blaIMP-1, blaVIM-1, blaNDM-1, and blaOXA-48 genes via polymerase chain reaction (PCR). Of the 230 isolates comprising E. coli (40.9%), Citrobacter spp. (32.6%), K. pneumoniae (9.1%), P. mirabilis (6.1%), P. vulgaris (5.2%), Enterobacter spp (3.5%)., K. oxytoca (2.2%), and Serratia marcenses (0.4%). Most isolates were from urine 162(70.4%) and wound samples. The isolates showed high resistance to ampicillin 171 (74.3%) and cefuroxime 134(58.3%). The prevalence of MDR was 35.2% (81), with E. coli 40(42.6%) being the majority that exhibited MDR. Of the 230 isolates, 113(49.1%) were ESBL producers, with E. coli 54(57.5%) accounting for the majority, while Serratia marcenses was the least. Of the 13 (5.7%) CRE isolates that showed resistance towards carbapenem in the disc diffusion method, 11 showed the presence of the blaNDM-1 gene, while all isolates showed the presence of the blaOXA-48 gene. The prevalence of carbapenem resistance and ESBL-producing Enterobacteriaceae pathogens among patients at the Cape Coast Teaching Hospital is high and alarming. Therefore, it is imperative to consider effective infection prevention and control measures should be implemented at the hospital to prevent the rapid spread of these dangerous organisms.

Fecal Carriage of Multi-drug Resistant Escherichia coli in Healthy Children: a Community-based Study

Introduction: Escherichia coli is enteric Gram-negative bacilli, earlier recognized as non-invasive commensal. Invasive strains of E. coli have been identified and are associated with a variety of human infections. Increasing drug resistance among E. coli has been reported. Multidrug resistance is the ability of an organism to resist at least one agent in three or more antimicrobial categories. The aim of this study was to detect fecal carriage of MDR E. coli isolates among healthy school children.
 Methods: A total of 139 stool samples were collected from Rastriya Secondary School, Purano Tudikhel, Pokhara-1. Isolation, identification, and Antibiotic susceptibility test pattern of E. coli isolates were done by standard microbiological techniques. Further Extended Spectrum Beta-Lactamase test was performed by Combined Disc Test and data analysis was done by using Microsoft Excel.
 Results: A total of 106 E. coli were isolated from the stool samples. MDR E. coli were found to be 8 (7.55%) and 7 E. coli isolates were resistant to Ceftazidime which was further tested for ESBL and 5 isolates were found to be ESBL producers. The resistance of E. coli to Ampicillin, Nitrofurantoin, Ceftriaxone, Ceftazidime, Ciprofloxacin, and Gentamicin was 27.36%, 43%, 10.38%, 6.60%, 6.60%, and 2.83% respectively. Only one isolate was resistant to all antibiotics used except Imipenem.
 Conclusion: E. coli is the main Gram-negative bacteria isolated from the stool sample and identified as the main carrier of antimicrobial resistance in species comparison of Enterobacteriaceae isolated from fecal samples. Therefore, quick surveillance is needed in order to effectively handle such MDR strains. The E. coli population susceptible to all antibiotics used was a much more diverse group than the resistant and ESBL-producing E. coli.

High prevalence of extensively drug resistant and extended spectrum beta lactamases (ESBLs) producing uropathogenic Escherichia coli isolated from Faisalabad, Pakistan.

Urinary tract infections (UTIs) are predominantly caused by uropathogenic Escherichia coli (E. coli). There is rapid increase in antimicrobial resistance in UTIs, also declared as a serious health threat by World Health Organization (WHO). Present study was designed to investigate the antimicrobial resistance status with specific focus on ESBLs and carbapenemases in local uropathogenic E. coli (UPEC) isolates. E. coli isolates were characterized from patients of all ages visiting diagnostic laboratories for urine examination. Demographic data was also recorded for each patient. Antibiograms were developed to observe antibiotic resistance in UPEC using Kirby Bauer disc diffusion technique. Double Disc Synergy test (DDST) was used for phenotypic ESBL test. ESBLs and carbapenemases genes were detected in UPEC using PCR. The PCR results were confirmed by sequencing. The UPEC isolates under study exhibited 78%, 77%, 74%, 72% and 55% resistance against cefotaxime, amoxicillin, erythromycin, ceftriaxone and cefixime, respectively. Resistance against colistin and meropenem was observed in 64% and 34% isolates, respectively. Phenotypic DDST identified 48% isolates as ESBLs producers. Genotypic characterization identified 70%, 74.4% and 49% prevalence of CTXM-1, TEM-1 and CTXM-15 genes respectively. One isolate was observed exhibiting co-existence of all ESBL genes. TEM-1 + CTXM-1 and TEM-1 + CTXM-1 + CTXM-15 + OXA-1 gene patterns were dominant among ESBLs. For carbapenem-resistance, 14% isolates indicated the presence of KPC whereas GES and VIM was detected in 7% and 3.4% isolates, respectively. In conclusion, our results present a high prevalence of extensively drug resistant UPEC isolates with a considerable percentage of ESBL producers. These findings propose the need of continuous surveillance for antimicrobial resistance and targeted antimicrobial therapy.

647. Cefepime Versus Carbapenems for the Treatment of ESBL-producing Enterobacterales among Non-blood Isolates

Abstract Background The incidence of organisms with extended-spectrum beta-lactamases (ESBLs) is increasing. The data for using cefepime in ESBL-producing Enterobacterales infections is conflicting. More favorable outcomes are likely if minimum inhibitory concentrations (MICs) < 2 mcg/mL. The aim of this study is to compare the efficacy of cefepime versus carbapenems for ESBL-producing, non-bloodstream Enterobacterales infections. Methods This study was a single-center retrospective cohort study of patients who received cefepime or a carbapenem for at least 72 hours for the definitive treatment of an ESBL-producing Enterobacterales non-bloodstream infection between Jan. 1, 2011 and Sept. 30, 2021. ESBL production was identified if the isolate either had a positive confirmatory ESBL test from VITEK 2 (bioMérieux) or phenotypic resistance to ceftriaxone. Isolates had to have a MIC < 2 mcg/mL to cefepime or be susceptible to carbapenems. Isolates with a cefepime MIC 4-8 mcg/mL were not included due to likely higher rates of treatment failure. The primary endpoint was clinical failure, defined as persistence of symptoms requiring escalation of therapy or death. Descriptive statistics were used to compare groups. A univariate analysis and odds ratio was calculated for clinical outcomes. Results One hundred patients were included. Twenty-two patients received cefepime and 78 received a carbapenem. Table 1 describes patient characteristics and clinical outcomes. Most patients had a urinary tract infection (UTI) (94%). More patients receiving cefepime were admitted to the intensive care unit (40.9% versus 15.4%). Treatment with cefepime displayed higher rates of clinical failure when compared to treatment with carbapenems (13.6% versus 6.4%). Cohen’s d-test for clinical failure was 0.46, indicating a medium effect. Conclusion Based on our analysis, cefepime may have higher rates of clinical failure when compared to carbapenem treatment for ESBL-producing Enterobacterales. Most of our patients were treated for UTIs and the sample size for both arms were limited, particularly for the cefepime arm. Further research is needed to confirm the role of cefepime as a carbapenem-sparing option in the treatment of these drug-resistant infections. Disclosures Madeline King, PharmD, Shionogi: Honoraria|Tetraphase: speakers' bureau.

Cefixime and cefixime-clavulanate for screening and confirmation of extended-spectrum beta-lactamases in Escherichia coli

IntroductionDetection of Extended-Spectrum Beta-Lactamases (ESBLs) depends on screening for resistance to certain cephalosporins, confirmation with selective ESBL inhibitors, and ESBL genes detection. New tests are required for accurate ESBL detection.AimsTo test the ability of cefixime (CFM) and cefixime-amoxicillin/clavulanate (CFM-AMC) as a screening and confirmatory test for ESBL identification.Methods246 clinical isolates of Escherichia coli were tested by an ESBL screening test, a double-disk synergy test (DDST), a disk replacement test, the Vitek 2 ESBL test, and an ESBL genes test by PCR. CFM ESBL Screening was performed by disk diffusion, while CFM-AMC confirmation was performed by DDST and a disk replacement test.Results246 E. coli clinical isolates from two referral hospitals were collected over 2 years. The mean age ± standard deviation of patients was 43.8 ± 27.7 years and 76.8% were females. Resistance rates to penicillins, first, second, and third generation cephalosporins, and monobactams were very high at 97%, 84%, 100% and 97%, respectively. ESBL screening was positive in 81.3% of isolates, DDST was positive in 74.8%, disk replacement was positive in 79%, Vitek 2 ESBL test was positive in 67.3%, and ESBL genes were detected in 85.8% of isolates (CTX-M 75%, TEM 42.5%, SHV 4.6%). Compared to genotyping, screening with CFM achieved 87.7% sensitivity and 64.7% specificity. CFM-AMC DDST achieved 75.8% sensitivity and 75.4% specificity, and CFM-AMC disk replacement had 73% sensitivity and 70% specificity.ConclusionsHigh prevalence of ESBLs was noted among E. coli isolates, dominated by CTX-M genotype. ESBL screening and confirmation using CFM and CFM-AMC is a new and accurate method for ESBLs detection.

Antibiotic resistance and detection of plasmid mediated colistin resistance mcr-1 gene among Escherichia coli and Klebsiella pneumoniae isolated from clinical samples

BackgroundThe prevalence of antimicrobial resistance (AMR) among Gram-negative bacteria is alarmingly high. Reintroduction of colistin as last resort treatment in the infections caused by drug-resistant Gram-negative bacteria has led to the emergence and spread of colistin resistance. This study was designed to determine the prevalence of drug-resistance among beta-lactamase-producing strains of Escherichia coli and Klebsiella pneumoniae, isolated from the clinical specimens received at a tertiary care centre of Kathmandu, Nepal during the period of March to August, 2019.MethodsA total of 3216 different clinical samples were processed in the Microbiology laboratory of Kathmandu Model Hospital. Gram-negative isolates (E. coli and K. pneumoniae) were processed for antimicrobial susceptibility test (AST) by using modified Kirby-Bauer disc diffusion method. Drug-resistant isolates were further screened for extended-spectrum beta-lactamase (ESBL), metallo-beta-lactamase (MBL), carbapenemase and K. pneumoniae carbapenemase (KPC) production tests. All the suspected enzyme producers were processed for phenotypic confirmatory tests. Colistin resistance was determined by minimum inhibitory concentration (MIC) using agar dilution method. Colistin resistant strains were further screened for plasmid-mediated mcr-1 gene using conventional polymerase chain reaction (PCR).ResultsAmong the total samples processed, 16.4% (529/3216) samples had bacterial growth. A total of 583 bacterial isolates were recovered from 529 clinical samples. Among the total isolates, 78.0% (455/583) isolates were Gram-negative bacteria. The most predominant isolate among Gram-negatives was E. coli (66.4%; 302/455) and K. pneumoniae isolates were 9% (41/455). In AST, colistin, polymyxin B and tigecycline were the most effective antibiotics. The overall prevalence of multidrug-resistance (MDR) among both of the isolates was 58.0% (199/343). In the ESBL testing, 41.1% (n = 141) isolates were confirmed as ESBL-producers. The prevalence of ESBL-producing E. coli was 43% (130/302) whereas that of K. pneumoniae was 26.8% (11/41). Similarly, 12.5% (43/343) of the total isolates, 10.9% (33/302) of E. coli and 24.3% of (10/41) K. pneumoniae were resistant to carbapenem. Among 43 carbapenem resistant isolates, 30.2% (13/43) and 60.5% (26/43) were KPC and MBL-producers respectively. KPC-producers isolates of E. coli and K. pneumoniae were 33.3% (11/33) and 20% (2/10) respectively. Similarly, 63.6% (21/33) of the E. coli and 50% (5/10) of the K. pneumoniae were MBL-producers. In MIC assay, 2.2% (4/179) of E. coli and 10% (2/20) of K. pneumoniae isolates were confirmed as colistin resistant (MIC ≥ 4 µg/ml). Overall, the prevalence of colistin resistance was 3.1% (6/199) and acquisition of mcr-1 was 16.6% (3/18) among the E. coli isolates.ConclusionHigh prevalence of drug-resistance in our study is indicative of a deteriorating situation of AMR. Moreover, significant prevalence of resistant enzymes in our study reinforces their roles in the emergence of drug resistance. Resistance to last resort drug (colistin) and the isolation of mcr-1 indicate further urgency in infection management. Therefore, extensive surveillance, formulation and implementation of effective policies, augmentation of diagnostic facilities and incorporation of antibiotic stewardship programs can be some remedies to cope with this global crisis.

Pediatric Febrile Urinary Tract Infection Caused by ESBL Producing Enterobacteriaceae Species.

Background Over the past decade, drug resistance pattern has worsened for many of the uropathogens due to overuse of antibiotics for empiric treatment. The burden of extended spectrum beta-lactamase (ESBL) producing Enterobacteriaceae associated urinary tract infections (UTI) has become increasingly more common, limiting treatment options among children presenting with febrile UTI. We investigated the burden and correlates of ESBL producing Enterobacteriaceae associated UTI among children and antibacterial resistance pattern. Methods 284 midstream urine specimens were collected using standard aseptic techniques from 284 children who were diagnosed with suspected UTI. Urine culture and bacteria isolation were performed following standard bacteriological techniques. The Kirby-Bauer disk diffusion technique and the double-disc synergy test were used to investigate antibiotic susceptibility and presence of ESBL production. Results UTI was confirmed using a positive urine culture for a relevant pathogen in 96/284 (33.8%) of the cases. Enterobacteriaceae accounted for 75% (72/96) of etiologies of UTI in children. The most frequent Enterobacteriaceae spp. were E. coli, 44.4% (32/72) and K. pneumonia, 27.8% (20/72). The overall multidrug resistance rate was 86.1% (62/72). ESBL-producers accounted for 41.7% (30/72) of the isolated Enterobacteriaceae. ESBL producing K. pneumonia and E. coli isolates accounted for 70% (14/20) and 37.5% (12/32), respectively. History of UTI in the past 1 year (adjusted odds ratio (AoR) = 0.08, 95%CI (0.01 − 0.57)) and medium family wealth index (AoR = 0.03, 95%CI (0.00 − 0.27)) protected from infection with ESBL-producing Enterobacteriaceae. Conclusion. ESBL production was more common in K. pneumonia and appeared to be a major factor contributing drug resistance UTI in children. The findings call for the need to incorporate ESBL testing in the routine clinical practice. The resistance level to commonly prescribed first-line antibiotics observed within Enterobacteriaceae was alarming calling for strengthened antimicrobial stewardship.

Extended Spectrem Beta Lactamases among Multi Drug Resistant Gram Negetive Bacilli Causing Urinry Tract Infection

Extended Spectrum Beta Lactamases (ESBL), the main cause of resistance to broad spectrum β-lactams, among uropathogenic bacteria have increased over time raising a global concern in the therapeutic management of infections caused by these organisms. The study was carried out in Janamaitri Hospital, Kathmandu between December 2012 to May 2013 with an objective to determine the status of ESBL producing Gram negative bacilli isolated from the urine sample, collected from patients suspected of urinary tract infection. Gram negative bacilli isolated were tested for the presence of ESBL by combined disk and antibiotic susceptibility by Kirby Bauer disc diffusion method following Clinical and Laboratory Standard Institute guidelines. Among the total 1105 mid-stream urine samples, 256 Gram negative bacilli were isolated. By screening test using third generation cephalosporins, 156 isolates were screened as ESBL producers and 91 isolates were positive for ESBL test by combined disk method. Among the 91 (35.55%) ESBL producers, 70 (39.32%) Escherichia coli, 16 (44.44%) Klebsiella pneumoniae, and 5 (33.33%) Pseudomonas aeruginosa were found to be ESBL producers. Majority of ESBL producer showed resistance to ampicillin, co-trimoxazole, norfloxacin followed by ofloxacin. imipenem, amikacin and nitrofurantoin seemed to be the agent of choice for urinary tract infections when ESBL producers are susceptible to it. ESBL production found in these Gram negative bacilli with resultant microbial resistance to available cephalosporins and other agents may pose difficulties with the choice of therapeutic options for the treatment of severe infections.
 Keywords: UTI, Extended Spectrum Beta Lactamases, Gram negative bacilli

Widespread antimicrobial resistance among bacterial infections in a Rwandan referral hospital.

BackgroundResistance among bacterial infections is increasingly well-documented in high-income countries; however, relatively little is known about bacterial antimicrobial resistance in low-income countries, where the burden of infections is high.MethodsWe prospectively screened all adult inpatients at a referral hospital in Rwanda for suspected infection for seven months. Blood, urine, wound and sputum samples were cultured and tested for antibiotic susceptibility. We examined factors associated with resistance and compared hospital outcomes for participants with and without resistant isolates.ResultsWe screened 19,178 patient-days, and enrolled 647 unique participants with suspected infection. We obtained 942 culture specimens, of which 357 were culture-positive specimens. Of these positive specimens, 155 (43.4%) were wound, 83 (23.2%) urine, 64 (17.9%) blood, and 55 (15.4%) sputum. Gram-negative bacteria comprised 323 (88.7%) of all isolates. Of 241 Gram-negative isolates tested for ceftriaxone, 183 (75.9%) were resistant. Of 92 Gram-negative isolates tested for the extended spectrum beta-lactamase (ESBL) positive phenotype, 66 (71.7%) were ESBL positive phenotype. Transfer from another facility, recent surgery or antibiotic exposure, and hospital-acquired infection were each associated with resistance. Mortality was 19.6% for all enrolled participants.ConclusionsThis is the first published prospective hospital-wide antibiogram of multiple specimen types from East Africa with ESBL testing. Our study suggests that low-resource settings with limited and inconsistent access to the full range of antibiotic classes may bear the highest burden of resistant infections. Hospital-acquired infections and recent antibiotic exposure are associated with a high proportion of resistant infections. Efforts to slow the development of resistance and supply effective antibiotics are urgently needed.

Prevalence of ESBL in Escherichia coli Isolates Among ICU Patients in a Tertiary Care Hospital

Resistance to common antibiotics is a matter of grave concern in treating infections in hospital settings especially in Intensive Care Units (ICUs). One of the most commonly used and effective group of antibiotics, cephalosporins, exhibit resistance due to production of Extended Spectrum Beta- Lactamases (ESBLs). The prevalence of ESBL producing Escherichia coli (E.coli) has increased throughout the world and is a major cause of treatment failure in ICUs. As per our knowledge studies were not available on the prevalence of ESBL producing E.coli in ICUs of this region. To determine the prevalence of ESBLs among Escherichia coli isolates in ICUs of a tertiary care hospital. A cross sectional study was conducted over a period of 4 years (Sept 2011 to Sept 2015) in the Department of Microbiology, Kalinga Institute of Medical Sciences (KIMS), Bhubaneswar. Consecutive non-duplicate isolates of E.coli recovered from 6800 clinical samples of patients admitted to different Intensive Care Units (ICUs) were subjected to ESBL screening test and then to CLSI recommended Phenotypic Confirmatory Disc Diffusion Tests (PCDDT) for ESBL production determination. Out of 6800 samples, 1038 were E.coli isolates and 452(44%) were resistant to third generation cephalosporins. ESBL producing Escherichia coli among them were 276 (61.1%). Paediatric ICU showed the highest prevalence of ESBL E.coli at 80.9%. The highest prevalence of ESBL E.coli was in urine samples (82.6%) followed by pus (9.8%). The most effective antibiotic for ESBL producers was imipenem (96.7% sensitive), followed by amikacin (88.4%) and piperacillin- tazobactum (87%). This study has highlighted the high prevalence of ESBL producing E.coli in the ICUs of our hospital. An in depth analysis of their antibiogram will be helpful in formulating the antibiotic policy and prevent spread of ESBL strains. It is recommended that ESBL testing should be done routinely to curtail antibiotic resistance and to effectively implement infection control measures.

Evaluation of a chromogenic medium for detection of extended-spectrum-beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae strains.

Because of the emergence and spread of extended-spectrum-beta-lactamase (ESBL)-producing strains which are resistant to many antibiotics, reliable detection of ESBL is very important for infection control. Several chromogenic media have been proposed for the detection of ESBL producers in addition to the conventional phenotypic and genotyping methods. The aim of the present study was to evaluate the performance of Brilliance ESBL agar (Oxoid; Thermo Fisher Scientific, UK), a selective chromogenic agar for the detection of ESBL-producing Escherichia coli (E. coli) and Klebsiella pneumoniae (K. pneumoniae) strains. A total of 237 strains (143 ESBL producers (76 isolates of E. coli and 67 isolates of K. pneumoniae) and 94 non-ESBL producers (44 isolates of E. coli and 50 isolates of K. pneumoniae)) isolated from various clinical specimens were included in the study. Isolates were identified by conventional methods, Phoenix system (Becton Dickinson, USA), and mass spectrometry. ESBL confirmation was performed by phenotypical tests. A 10 microL aliquot of each isolate's 0.5 McFarland suspension was streaked onto Brilliance ESBL agar. All plates were incubated at 37 degrees C for 24 hours and then were interpreted for growth and colony color according to the manufacturer's recommendations. Identification and ESBL test results were used to calculate the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the medium evaluated at 24 hours. The sensitivity, specificity, PPV, and NPV of the medium were 97.9%, 100%, 100%, and 96.9%, respectively, when considering only species specific colored colonies of the isolates. Brilliance ESBL agar could provide a practical alternative to the traditional methods for the identification of ESBL producers.

Extremely high prevalence of multi-resistance among uropathogens from hospitalised children in Beira, Mozambique

A prospective surveillance study was conducted to investigate the epidemiology and patterns of antibiotic resistance among uropathogens from hospitalised children in Beira, Mozambique. Additionally, information regarding determinants of a urinary tract infection (UTI) was obtained. Bacterial species identification, antimicrobial susceptibility testing and extended-spectrum beta-lactamase testing were performed for relevant bacterial isolates. Analysis of 170 urine samples from 148 children yielded 34 bacterial isolates, predominantly Escherichia coli and Klebsiella spp., causative of a urinary tract infection in 29 children; 30/34 isolates (88.2%) from 26/29 children (89.7%) were considered highly resistant micro-organisms (HRMOs). No significant determinants of urinary tract infection with HRMOs were detected when analysing gender, antibiotic use during hospital admission and HIV status. This study shows, for the first time in Mozambique, an extremely high prevalence of HRMOs among uropathogens from hospitalised children with a urinary tract infection.

Evaluation of the VITEK 2 AST-N111 card for detection of extended-spectrum beta-lactamases (ESBLs) in Escherichia coli, Klebsiella pneumoniae, and Klebsiella oxytoca compared to ESBL Etests and combination disk methods

The VITEK 2 AST-N111 card was evaluated for detection of extended-spectrum beta-lactamases (ESBLs) by testing 51 ESBL positive and 50 ESBL negative isolates of E. coli, K. pneumoniae, and K. oxytoca. The occurrence of beta-lactamase genes was confirmed by PCR and sequencing. The advanced expert system (AES) of the VITEK 2 system achieved sensitivity and specificity values of 100% and 96.0%, respectively. The ESBL test of the VITEK 2 AST-N111 card showed a sensitivity of 92.1% and a specificity of 90.0%. Contradictory results obtained with the two VITEK 2 tools could be clarified by combination disk tests in nine of 11 isolates. The combined use of AES and ESBL tests of the AST-N111 card in association with combination disk tests in case of contradictory results seems to be a reliable method for ESBL detection.

Investigation of Extended Spectrum Beta Lactamase Production of Bacteria by Direct Urine Inoculation

Extended spectrum beta lactamase (ESBL) production of bacteria was investigated by direct urine inoculation method in urine samples considered to be infected with Enterobactericeae. Urine samples were aliquoted to two sterile containers. One container was used for microbiological examination and the other one was used for dipstick analyses. ESBL phenotype (both in the direct test and in the standard test) was determined by disc diffusion method. 146 urine samples were tested for phenotypic screening and confirmation tests by direct inoculation method. 110 of them were found to be positive for leukocyte esterase (LE) and nitrite (NIT) with dipstick analyses. 36 of them were found to be positive for LE only with dipstick analyses and containing gram negative bacilles with gram staining. Standard ESBL testing was also performed in parallel. ESBL production was detected in 38 bacteria with direct inoculation method and also confirmed with standart methods. Results of the two tests were in concordance. In conclusion, ESBL production of bacteria could be detected earlier with direct inoculation method compared to standard inoculation method. Use of this method could expedite the initiation of appropriate antimicrobial therapy in hospitals known to have higher rates of ESBL production. Key words: Urinary tract infections, antimicrobial resistance, extended spectrum beta lactamase.

Prevalence, Molecular Characterization, and Phenotypic Confirmation of Extended-Spectrum Beta-Lactamases inEscherichia coli,Klebsiella pneumoniae, andKlebsiella oxytocaat the Radboud University Nijmegen Medical Centre in The Netherlands

The prevalence and molecular types of extended-spectrum beta-lactamases (ESBLs) were determined during a 1-year period in unselected clinical nonduplicate isolates of Escherichia coli (n = 1,738), Klebsiella pneumoniae (n = 436), and Klebsiella oxytoca (n = 208), cultured at the University Medical Centre Nijmegen, The Netherlands. Isolates identified as ESBL producer by the Phoenix automated system were collected prospectively and subjected to molecular analysis for the most common ESBLs TEM, SHV, and CTX-M, as well as OXA and GES. Both the Etest ESBL and double-disk synergy test were performed as confirmatory tests. The estimated prevalence of ESBLs was 2.1% in E. coli, 5.2% in K. pneumoniae, and 2.4% in K. oxytoca. TEM-12 and -26, SHV-5 and -12, and CTX-M groups 1 and 9 were the most frequent ESBLs found. Isolates identified as ESBLs by the Phoenix were confirmed by polymerase chain reaction (PCR) in only 42%. In ESBL PCR-positive E. coli and K. pneumoniae, both confirmatory tests were positive in 95% of the isolates. In 28% of the Etest and 13% of the double-disk synergy test-positive isolates, PCR could not detect any ESBL gene. In these cases, other resistance mechanisms may play a role. Confirmatory tests were unreliable for K. oxytoca. A previously described mutation in the K1 enzyme was detected in one ceftazidime-resistant K. oxytoca. The prevalence of ESBLs in The Netherlands is increasing. The predominant molecular types of ESBLs detected were comparable to other studies. Phoenix ESBL results need to be confirmed as advocated by ESBL detection guidelines.

Evaluation of the Phoenix Automated Microbiology System for Detecting Extended-Spectrum β-Lactamase in Escherichia coli, Klebsiella species and Proteus mirabilis

The aim of this study was to compare the BD Phoenix (Beckton Dickinson Diagnostic Systems, USA) extended-spectrum beta-lactamase (ESBL) test with the Clinical and Laboratory Standards Institute (CLSI) ESBL phenotypic confirmatory test by disk diffusion (CLSI ESBL test) in Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca and Proteus mirabilis. We tested 224 clinical isolates of E. coli, K. pneumoniae, K. oxytoca and P. mirabilis during May 2006 to March 2007. These isolates were examined by the Phoenix and the CLSI ESBL tests simultaneously. For the isolates showing discordant results between the two tests, boronic acid disk test was performed to differentiate AmpC beta-lactamase and ESBL. Among the 224 clinical isolates, 75 and 79 isolates were positive for ESBL by CLSI ESBL test and Phoenix test, respectively. Having detected 4 more isolates as ESBL-producers, Phoenix test showed a 98.2% agreement with a 100% sensitivity and 97.3% specificity compared with CLSI ESBL test. Among the four false positive isolates, three were AmpC-positive but ESBL-negative. The BD Phoenix ESBL test was sensitive and specific, and can be used as a rapid and reliable method to detect ESBL production in E. coli, Klebsiella species, and P. mirabilis.

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 beta-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 beta-lactamases, including 61 different ESBLs, two class A carbapenemases and various species-specific beta-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.

Evaluation of detection of extended-spectrum beta-lactamases among Escherichia coli and Klebsiella spp. isolates by VITEK 2 AST-N029 compared to the agar dilution and disk diffusion methods

A total of 123 clinical Escherichia coli and Klebsiella spp. isolates were included in the study in order to evaluate VITEK 2 AST-NO29 (Nordic) card for detection of extended-spectrum beta-lactamases (ESBL) and to compare the results with genotypic ESBL verification. The results were also compared to alternative phenotypic methods, i.e. agar dilution and disk diffusion. The strains that were ESBL-positive according to AST-N029 were further analysed with the ESBL test card, VITEK 2 AST-N041. Using genotype as reference, Vitek 2 AES had the highest accuracy of the tested methods in classifying the strains as ESBL-positive or -negative (91.1%). When VITEK 2 gave ESBL as the only option for E. coli or K. pneumoniae, 44 of 45 (97.8%) strains had an ESBL gene. VITEK 2 achieved an accuracy of 94.9% and disk diffusion 95.9% compared to the agar dilution method as the phenotypic reference method for the E. coli and K. pneumoniae strains. For the K. oxytoca strains VITEK 2 achieved the highest accuracy (84.0%) of the methods used in this work.