Molecular characterization of extended-spectrum beta-lactamases produced by clinical isolates of Klebsiella pneumoniae and Escherichia coli from a Korean nationwide survey

To determine the prevalence and genotypes of extended-spectrum beta-lactamases (ESBLs) among clinical isolates of Klebsiella pneumoniae and Escherichia coli, we performed antibiotic susceptibility testing, pI determination, induction testing, transconjugation, and DNA sequencing analysis. Among the 509 isolates collected from 13 university hospitals in Korea, 39.2% produced ESBLs. ESBL-producing isolates were detected in every region in Korea. A total of 44.6% of the isolates produced both TEM- and SHV-type ESBLs, and 52% of ESBL-producing isolates transferred resistance to ceftazidime by transconjugation. The ESBLs were TEM-19, TEM-20, TEM-52, SHV-2a, SHV-12, and one new variant identified for the first time in Korea, namely, TEM-116. TEM-1 and SHV-12 were by far the most common variants. TEM-1, TEM-116, and SHV-12 showed a high prevalence in K. pneumoniae. Two isolates (E. coli SH16 and K. pneumoniae SV3) produced CMY-1-like beta-lactamases, which play a decisive role in resistance to cefoxitin and cefotetan, as well as TEM-type enzymes (TEM-20 and TEM-52, respectively). Using MIC patterns and DNA sequencing analysis, we postulated a possible evolution scheme among TEM-type beta-lactamases in Korea: from TEM-1 to TEM-19, from TEM-19 to TEM-20, and from TEM-20 to TEM-52.     

Survey of Extended-Spectrum β-Lactamases in Clinical Isolates of Escherichia coli and Klebsiella pneumoniae: Prevalence of TEM-52 in Korea

Two hundred ninety isolates of Escherichia coli were investigated for the production of extended-spectrum beta-lactamases (ESBLs). Fourteen (4.8%) of the 290 strains were found to produce ESBLs. Each of the 14 strains produced one or two ESBLs, as follows: 10 strains produced TEM-52, 1 strain produced SHV-2a, 1 strain produced SHV-12, 1 strain produced a CMY-1-like enzyme, and 1 strain expressed SHV-2a and a CMY-1-like enzyme. Another two strains for which the MICs of ceftazidime and cefoxitin were high, were probable AmpC enzyme hyperproducers. Because of the high prevalence of TEM-52 in E. coli isolates, we further investigated the TEM-type ESBLs produced by Klebsiella pneumoniae in order to observe the distribution of TEM-52 enzymes among Enterobacteriaceae in Korea. All TEM enzymes produced by 12 strains of K. pneumoniae were identified as TEM-52. To evaluate the genetic relatedness among the organisms, ribotyping of TEM-52-producing E. coli and K. pneumoniae was performed. The ribotyping profiles of the organisms showed similar but clearly different patterns. In conclusion, TEM-52 is the most prevalent TEM-type ESBL in Korea.     

转座子tnpU基因在多重耐药革兰阴性杆菌中的分布情况

目的研究转座子tnpU基因和β-内酰胺酶在多重耐药革兰阴性杆菌中的分布情况。方法用纸片扩散初筛试验、扩散确证试验进行超广谱β-内酰胺酶（ESBLs）表型检查,头孢西丁三维试验进行AmpC β-内酰胺酶的表型检查,纸片协同试验筛选产金属β-内酰胺酶（MBL）菌株;用多重聚合酶链反应（PCR）技术扩增转座子tnpU基因,并进行DNA测序;用MIC药敏法分析多重耐药革兰阴性杆菌的药物敏感性。结果转座子tnpU的总检出率为25．5％。在各菌种的检出率分别为大肠埃希菌占6.3％（3株）、肺炎克雷伯菌占8．3％（1株）、鲍曼不动杆菌占33-3％（20株）、铜绿假单胞菌占43．2％（16株）;β-内酰胺酶表型检测中,ESBLs的检出率最高,转座子tnpU基因阳性的菌株大多数B．内酰胺酶表型为阳性;转座子tnpU基因阳性菌株对抗生素的耐药率显著高于转座子阴性菌株（P〈0．05）。结论转座子tnpU基因在非发酵菌中的分布较广泛,可能在多重耐药机制中起重要作用。     

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 characteristics of extended-spectrum beta-lactamases in Korean isolates of Enterobacteriaceae

Extended-spectrum beta-lactamases (ESBLs) in gram-negative organisms have been implicated as the enzymes responsible for resistance to oxyimino-cephalosporins. The incidence of ESBL-producers in Korean isolates of Escherichia coli and Klebsiella pneumoniae were in the range of 4.8-7.5% and 22.5-22.8%, respectively. The ESBL-producing isolates revealed variable levels of resistance to cefotaxime, ceftazidime and aztreonam. They also showed the elevated MIC values of non-beta-lactam antibiotics. SHV-12 and SHV-2a were the enzymes most frequently found in K. pneumoniae strains, but TEM-52 was the most prevalent in E. coli isolates. About 15% of ESBL-producing isolates of Enterobacteriaceae produced CMY-1 enzyme, which conferred resistance to cephamycins such as cefoxitin as well as oxyimino-cephalosporins. Thus, the most common types of ESBLs in Korea are TEM-52, SHV-12 SHV-2a, and CMY-1.     

Prevalence of plasmid-mediated AmpC beta-lactamases in Escherichia coli and Klebsiella pneumoniae in Korea

Cefoxitin-resistant Escherichia coli and Klebsiella pneumoniae are relatively prevalent in Korea, suggesting dissemination of plasmid-mediated AmpC beta-lactamases. In this study, 238 isolates of cefoxitin-resistant E. coli and K. pneumoniae (not including subspecies ozaenae and rhinoscleromatis) were collected in 2003 from 16 Korean hospitals. The prevalence of plasmid-mediated AmpC beta-lactamases was determined by PCR. The AmpC gene alleles detected in E. coli and K. pneumoniae were bla(DHA-1), 10 (8.6%) and 93 (76.2%); bla(CMY-1)-like, 14 (12.1%) and 2 (1.6%); and bla(CMY-2)-like, 38 (32.7%) and 1 (0.8%) isolates, respectively. The genes identified were bla(DHA-1), bla(CMY10)-like, and bla(CMY-2)-like, and a new variant, bla(CMY-18). Plasmidmediated AmpC gene allele-positive isolates were present both in large city and in small province hospitals, as well as in isolates from outpatients. The proportions of plasmid-mediated AmpC gene-positive isolates were similar in both expanded spectrum beta-lactamase (ESBL)-producing and -nonproducing isolates. In conclusion, DHA-1, CMY-2-like, and CMY-10-like plasmid-mediated AmpC beta-lactamase-producing K. pneumoniae and E. coli isolates are widely disseminated in both large city and small province hospitals. Absence of bla(CMY-1) and detection of a novel variant of bla(CMY-2), bla(CMY-18), indicate continued evolution of the prototype genes. Similar proportions of plasmid-mediated AmpC gene-positive isolates in both ESBL-producing and -nonproducing isolates suggest unhindered future spread of these resistances.     

Prevalence of newer beta-lactamases in gram-negative clinical isolates collected in the United States from 2001 to 2002

Newer beta-lactamases such as extended-spectrum beta-lactamases (ESBLs), transferable AmpC beta-lactamases, and carbapenemases are associated with laboratory testing problems of false susceptibility that can lead to inappropriate therapy for infected patients. Because there appears to be a lack of awareness of these enzymes, a study was conducted during 2001 to 2002 in which 6,421 consecutive, nonduplicate clinical isolates of aerobically growing gram-negative bacilli from patients at 42 intensive care unit (ICU) and 21 non-ICU sites across the United States were tested on-site for antibiotic susceptibility. From these isolates, 746 screen-positive isolates (11.6%) were referred to a research facility and investigated to determine the prevalence of ESBLs in all gram-negative isolates, transferable AmpC beta-lactamases in Klebsiella pneumoniae, and carbapenemases in Enterobacteriaceae. The investigations involved phenotypic tests, isoelectric focusing, beta-lactamase inhibitor studies, spectrophotometric assays, induction assays, and molecular analyses. ESBLs were detected only in Enterobacteriaceae (4.9% of all Enterobacteriaceae) and were found in species other than those currently recommended for ESBL testing by the CLSI (formerly NCCLS). These isolates occurred at 74% of the ICU sites and 43% of the non-ICU sites. Transferable AmpC beta-lactamases were detected in 3.3% of K. pneumoniae isolates and at 16 of the 63 sites (25%) with no difference between ICU and non-ICU sites. Three sites submitted isolates that produced class A carbapenemases. No class B or D carbapenemases were detected. In conclusion, organisms producing ESBLs and transferable AmpC beta-lactamases were widespread. Clinical laboratories must be able to detect important beta-lactamases to ensure optimal patient care and infection control.     

Prevalence of plasmid-mediated AmpC beta-lactamases in a Chinese university hospital from 2003 to 2005: first report of CMY-2-Type AmpC beta-lactamase resistance in China

The aim of this study was to investigate the prevalences of plasmid-mediated AmpC beta-lactamases (PABLs) in isolates of Escherichia coli and Klebsiella spp. from a university hospital in China. A total of 1,935 consecutive nonrepeat clinical isolates of Escherichia coli, Klebsiella pneumoniae, and Klebsiella oxytoca were collected between January 2003 and July 2005. The isolates with cefoxitin zone diameters less than 18 mm (screen positive) were selected for PCR of the bla(AmpC) genes and sequencing. Fifty-four (2.79%) isolates harbored PABLs, as demonstrated by PCR and isoelectric focusing. Sequence analysis revealed the presence of bla(DHA-1) and bla(CMY-2) genes. The Southern blot hybridization studies confirmed that bla(CMY-2) and bla(DHA-1) were located on plasmids. Based on species, PABLs were detected in 4.29% (29 isolates of DHA-1 and 1 isolate of CMY-2) of K. pneumoniae, 1.91% (11 isolates of DHA-1 and 12 isolates of CMY-2) of E. coli, and 3.03% (1 isolate of DHA-1) of K. oxytoca isolates. In contrast to our anticipation, the occurrence rate of DHA-1-producing K. pneumonia significantly decreased (P < 0.01), from 7.54% in 2003 to 2.72% in 2004. The results of random amplified polymorphic DNA analysis indicate that the prevalences of DHA-1-producing K. pneumoniae and CMY-2-producing E. coli strains were not due to epidemic strains. In conclusion, DHA-1 was the most prevalent acquired AmpC beta-lactamase in this collection of isolates from a medical center in China, and DHA-1-producing K. pneumoniae was the most prevalent bacterium harboring a PABL. To the best of our knowledge, this is the first report of CMY-2-type AmpC beta-lactamases in the Chinese mainland.     

儿童感染肺炎克雷伯菌产AmpC酶和ESBLs的检测及耐药性分析

目的了解广州地区儿童感染肺炎克雷伯菌产质粒介导的AmpC酶和超广谱β-内酰胺酶（ESBLs）的情况及其耐药特征,为临床合理用药提供参考依据。方法采用标准纸片扩散法检测ESBLs,头孢西丁三维试验法检测AmpC酶,K—B纸片法测定肺炎克雷伯菌对抗菌药物的敏感性。结果共检出248株肺炎克雷伯菌,其中46株产AmpC酶,阳性率为18．5％;157株产ESBLs,阳性率为63．3％;同时产AmpC酶和ESBLs菌株阳性率为18．1％。产AmpC酶肺炎克雷伯菌对第三代头孢菌素高度耐药。耐药率达80％～100％;对头孢吡肟、含酶抑制剂复合药的耐药率也在56．5％～93．5％之间：但对环丙沙星、阿米卡星的耐药率则在30％以下,对亚胺培南全部敏感。产ESBLs菌株对头孢菌素、含酶抑制剂复合药的耐药率也较高,在50％-91．7％之间,但对阿米卡星、环丙沙星、亚胺培南仍高度敏感。ESBLs阴性肺炎克雷伯菌对所测抗生素的敏感率均在81．2％以上。产酶菌株耐药率明显比非产酶菌株高。结论广州地区儿童感染肺炎克雷伯菌产ESBLs和AmpC酶的状况已十分突出：产酶菌株对常用抗生素的耐药率较高;碳青霉烯类抗生素可作为治疗产AmpC酶和／或ESBLs肺炎克雷伯菌感染的经验用药。     

Development of a multiplex PCR and SHV melting-curve mutation detection system for detection of some SHV and CTX-M beta-lactamases of Escherichia coli, Klebsiella pneumoniae, and Enterobacter cloacae in Taiwan

Infection by extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae has been increasing in Taiwan. Accurate identification of the ESBL genes is necessary for surveillance and for epidemiological studies of the mode of transmission in the hospital setting. We describe herein the development of a novel system, which consists of a multiplex PCR to identify bla(SHV), bla(CTX-M-3)-like, and bla(CTX-M-14)-like genes and a modified SHV melting-curve mutation detection method to rapidly distinguish six prevalent bla(SHV) genes (bla(SHV-1), bla(SHV-2), bla(SHV-2a), bla(SHV-5), bla(SHV-11), and bla(SHV-12)) in Taiwan. Sixty-five clinical isolates, which had been characterized by nucleotide sequencing of the bla(SHV) and bla(CTX-M) genes, were identified by the system. The system was then used to genotype the ESBLs from 199 clinical isolates, including 40 Enterobacter cloacae, 68 Escherichia coli, and 91 Klebsiella pneumoniae, collected between August 2002 and March 2003. SHV-12 (80 isolates) was the most prevalent type of ESBL identified, followed in order of frequency by CTX-M-3 (65 isolates) and CTX-M-14 (36 isolates). Seventeen (9%) of the 199 clinical isolates harbored both SHV- and CTX-M-type ESBLs. In contrast to Enterobacter cloacae, the majority of which produced SHV-type ESBLs, E. coli and K. pneumoniae were more likely to possess CTX-M-type ESBLs. Three rare CTX-M types were identified through sequencing of the bla(CTX-M-3)-like (CTX-M-15) and bla(CTX-M-14)-like (CTX-M-9 and CTX-M-13) genes. The system appears to provide an efficient differentiation of ESBLs among E. coli, K. pneumoniae, and Enterobacter cloacae in Taiwan. Moreover, the design of the system can be easily adapted for similar purposes in areas where different ESBLs are prevalent.     

Prevalence and characterization of extended-spectrum beta-lactamases in Klebsiella pneumoniae in Algiers hospitals (Algeria)

AIM OF THE STUDY: To determine the prevalence and the diversity of extended-spectrum beta-lactamases (ESBLs) in 196 Klebsiella pneumoniae clinical isolates collected from three hospitals in Algiers. MATERIALS AND METHODS: Antibiograms were done on Mueller-Hinton agar plates with the disc-diffusion method and MICs were determined by agar-dilution method. Mating experiments were performed in agar medium. Plasmid DNA was extracted by the alcalin-lysis method. Total DNA was extracted with a Qiagen mini kit and screened for bla(TEM) and bla(CTX-M) genes by PCR. Linkage of bla(CTX-M) genes with insertion sequence ISEcp1B and class 1 integrons was investigated by PCR. PCR products were sequenced by the Sanger method. The epidemiological relationships between ESBL-producing K. pneumoniae isolates were analyzed by ERIC-PCR. RESULTS: Thirty-nine (19.9%) isolates were found to produce ESBLs belonging to CTX-M-1 group and TEM penicillinases (CTX-M-3, CTX-M-15 and TEM-1). ERIC-PCR analysis showed that the isolates are genetically unrelated. The bla(TEM) and bla(CTX-M) genes as well as aminoglycosides and sulfonamides resistance determinants were found located in self-transferable plasmids of approximately 85 kb. The class 1 integrons and the insertion sequence ISEcp1B were present in the isolates and in their transconjugants. ISEcp1B was found genetically linked to the bla(CTX-M) genes and located 127bp upstream, with the presence of the V and W sequences. CONCLUSION: The study revealed a high rate of ESBL-producing K. pneumoniae in Algerian hospitals, resulting from horizontal dissemination of mobile bla(CTX-M) genes.     

Molecular epidemiology of Enterobacteriaceae isolates producing extended-spectrum beta-lactamases in a French hospital

In 2002, 80 isolates of Enterobacteriaceae producing extended-spectrum beta-lactamases (ESBLs) were collected from infected patients in our hospital. Enterobacter aerogenes was the most common bacterium isolated from all specimens (36.5%). The ESBLs were predominantly (90%) TEM derivatives (TEM-24, TEM-3). Pulsed-field gel electrophoresis highlighted that E. aerogenes, Klebsiella pneumoniae, and Citrobacter koseri had a clonal propagation.     

Modified double-disc test for detection of extended-spectrum beta-lactamases in Escherichia coli and Klebsiella pneumoniae

This study evaluates the performance of a modified double-disc test (MDDT) for the detection of extended-spectrum beta-lactamases (ESBLs) in clinical isolates of Escherichia coli and Klebsiella pneumoniae. Ninety-six isolates of E. coli and 40 K. pneumoniae are studied for ESBL production by the National Committee for Clinical Laboratory Standards (NCCLS) combination disc tests and MDDT A total of 112 (82%) isolates (80 [83%] E. coli, 32 [80%] K. pneumoniae) were positive for ESBL by MDDT compared to 102 (75%; 72 [75%] E. coli and 30 [75%] K. pneumoniae) by the NCCLS method. In 10 (7.4%) isolates, ESBLs were detected only by MDDT Twenty-four (17.6%) isolates were negative for ESBL by both methods. The protocol described in this study provides a more sensitive approach than does the NCCLS method for ESBL detection in E. coli and K. pneumoniae.     

Accelerated detection of extended-spectrum beta-lactamases in clinical isolates of Enterobacteriaceae

A prospective study is carried out to evaluate the performance of a protocol for the accelerated detection of extended-spectrum beta-lactamases (ESBLs) in clinical isolates of Escherichia coli, Klebsiella pneumoniae and other Gram-negative bacteria. A modified double-disc test (MDDT) is incorporated in a Gram-negative template for routine susceptibility testing. The MDDT identified accurately ESBLs in all isolates subsequently confirmed as ESBL-producers by the standard Clinical Laboratory Standards Institute (CLSI) combined disc method. Of 1213 isolates tested, 98 (8%) were positive for ESBLs by MDDT and 95 (7.8%) were positive by the CLSI method. ESBLs were detected in 48 (7.8%) E. coli, 21 (8%) K. pneumoniae, 12 (5.8%) Proteus mirabilis, 13 (18.8%) Providencia stuartii and four (6.8%) Enterobacter cloacae isolates. Time required for ESBL detection by the MDDT method was one day. The protocol described provides a simple, rapid and low-cost method for early detection of ESBLs in Gram-negative bacteria.     

Clinical variables associated with the isolation of Klebsiella pneumoniae expressing different extended-spectrum β-lactamases

Clinical variables associated with the isolation of Klebsiella pneumoniae expressing different extended-spectrum beta-lactamases (ESBLs) were studied. Clinical records of patients with ESBL-positive K. pneumoniae isolates between 1989 and 2003 (n = 80) were reviewed retrospectively. Patients with SHV- and TEM-type ESBLs were identified more frequently in the intensive care units (67% and 78%, respectively), whereas those with CTX-M ESBLs were found in medical wards (52.2%) or were outpatients (17.4%) (p <0.01). The absence of urinary or central catheters was associated with CTX-M-10 (p 0.013 and p <0.01, respectively). Central catheter-related infections and secondary bacteraemia were associated more frequently with SHV- and TEM-type ESBLs, whereas urinary tract infections were associated with CTX-M-10. Previous aminoglycoside use was associated particularly with SHV-type ESBLs (p <0.01), whereas amoxycillin-clavulanate and oral cephalosporins were associated with CTX-M-10 (p <0.01 and p 0.050, respectively). The frequency of adequate empirical treatment was low (22%), and 61% of patients were treated according to the susceptibility testing results. Mortality (22%) and related mortality (14%) did not differ statistically according to the type of ESBL. Different ESBL types in K. pneumoniae were associated with different clinical variables, and this should be taken into account in current and future epidemiological scenarios.     

High rate of fatal cases of pediatric septicemia caused by gram-negative bacteria with extended-spectrum beta-lactamases in Dar es Salaam, Tanzania

Extended-spectrum beta-lactamases (ESBLs) were present in high proportions of Escherichia coli (25% [9 of 36]) and Klebsiella pneumoniae isolates (17% [9 of 52]) causing pediatric septicemia at a tertiary hospital in Tanzania. Patients with septicemia due to ESBL-producing organisms had a significantly higher fatality rate than those with non-ESBL isolates (71% versus 39%, P = 0.039). This is the first report of the CTX-M-15 genotype of ESBLs on the African continent and the first observation of SHV-12 genotype in an isolate of Salmonella enterica serotype Newport.     

Occurrence of extended-spectrum beta-lactamases in members of the genus Shigella in the Republic of Korea

A nationwide survey was carried out in Korea to assess the prevalence of Shigella strains producing extended-spectrum beta-lactamases (ESBLs). From 1991 to 2002, 5,911 clinical strains were isolated and screened for resistance to extended-spectrum cephalosporins. Twenty of the Shigella isolates were ESBL positive, based on the synergistic effects between clavulanate and selected beta-lactams (ceftazidime and cefotaxime). Nucleotide sequence analysis of these isolates revealed that they harbored bla(TEM-19) (eight isolates), bla(TEM-15) (five isolates), bla(TEM-52) (six isolates), bla(TEM-17) (one isolate), bla(TEM-20) (one isolate), and bla(CTX-M-14) (three isolates). All the ESBL-encoding genes in this study were carried in conjugable plasmids. Thus, TEM-19, TEM-15, TEM-52, and CTX-M-14 beta-lactamases can be considered common Korean ESBL types in Shigella sonnei and are probably transmitted through interspecies spread between medical facilities and the community in Korea. This is the first report of the presence of TEM-17, TEM-19, and TEM-20 in Korea and in S. sonnei.     

Outbreak of infection caused by Enterobacter cloacae producing the novel VEB-3 beta-lactamase in China

Over a 4-month period from November 2002 to February 2003, 27 ceftazidime-resistant or cefotaxime-resistant nonrepetitive Enterobacter cloacae isolates were collected from 27 patients hospitalized at HuaShan Hospital, Shanghai, People's Republic of China. The Etest did not detect extended-spectrum beta-lactamases (ESBLs) in those 27 isolates; however, screening by the NCCLS ESBL disk test and confirmatory tests detected ESBLs in 4 of 27 isolates and PCR detected ESBLs in 23 of 27 isolates. The majority of ESBL producers exhibited the same repetitive extragenic palindromic PCR pattern but harbored different ESBL genes. CTX-M-3 was the most prevalent ESBL in our study. Interestingly, 12 clonally related E. cloacae isolates possessed a novel bla(VEB)-type beta-lactamase, bla(VEB-3). Bla(VEB-3) was encoded by the chromosome and was located in an integron. Nine of the 12 isolates harbored both the bla(VEB-3) and the bla(CTX-M-3)-like ESBLs. This is the first report of a VEB-1-like ESBL in China and the first report of the simultaneous presence of VEB-1 and CTX-M-3-like ESBLs in an isolate.     

Detection of extended-spectrum-beta-lactamase-producing members of the family Enterobacteriaceae with Vitek ESBL test.

A three-phase analysis of the Vitek ESBL test and a double-disk (2 disk) test was performed to assess their ability to detect extended-spectrum beta-lactamases (ESBLs) in members of the family Enterobacteriaceae. In the first two phases involving detection of ESBLs in 157 stains processing well-characterized beta-lactamases, sensitivity and specificity were found to be 99.5 and 100%, respectively, for the Vitek ESBl test and 98.1 and 99.4%, respectively, for the 2-disk test. In the third phase, in which the ability of each test to detect ESBLs in 295 clinical isolates was assessed, there was only one false positive (Vitek ESBL test). Across all three phases, the Vitek ESBL test was found to be much easier to perform than the 2-disk test. The latter also involved subjective interpretation of results. There were a total of 176 Escherichia coli and 157 Klebsiella pneumoniae isolates and less than 40 isolates of each of 14 other species evaluated. In a supplemental study of Klebsiella oxytoca, an organism possessing a chromosomal beta-lactamase similar to an ESBL, the Vitek ESBL test was found to be capable of detecting hyperproduction of this enzyme in strains of this species as well. These data indicate that the Vitek ESBL test is reliable for the detection of ESBLs in E. coli and K. pneumoniae, the two species in which ESBLs are most common, and of hyperproduction of the K. oxytoca beta-lactamase, a situation which engenders a level of resistance to this species similar to that seen with ESBLs.     

Community-wide outbreaks of clonally related CTX-M-14 beta-lactamase-producing Escherichia coli strains in the Calgary health region

Enterobacteriaceae producing extended-spectrum beta-lactamases (ESBLs) typically cause nosocomial infections. Previous surveillance in the Calgary Health Region showed that Escherichia coli strains producing ESBLs were common among community patients. During the period (2000 to 2002): 23 of 157 (15%) of the strains were positive for blaCTX-M genes from the CTX-M-I group (CTX-M-1-like) and 87 of 157 (55%) of the strains were positive for blaCTX-M genes from the CTX-M-III group (CTX-M-14-like). The objective of this study was to investigate the molecular epidemiology of these strains. The beta-lactamases were characterized, and the genetic relatedness of the isolates was analyzed by digesting genomic DNA with the restriction endonuclease XbaI and by performing pulse-field gel electrophoresis (PFGE). PFGE revealed two closely related restriction patterns (clusters CTXM14A and CTXM14AR) among 67 (77%) CTX-M-14 producers. These strains from CTXM14A had nearly identical susceptibility patterns and were isolated most often from urine samples obtained at community sites during 2000 and 2001. Strains from the CTX-M-1-like and CTX-M-negative groups were unrelated to clusters CTXM14, CTXM14AR, and CTXM14NR. We conclude that clonally related strains of E. coli producing CTX-M-14 beta-lactamases were responsible for a predominantly community-wide outbreak. Further studies are warranted to investigate whether community-onset diseases caused by ESBL-producing E. coli are related to a point source or transmission within the community.