Comparison of screening methods for TEM- and SHV-derived extended-spectrum β-lactamase detection

Objective   To compare common extended-spectrum β -lactamase (ESBL) screening methods and β -lactams for their ability to detect TEM- and SHV-related ESBL enzymes.
Methods   This study compared disk diffusion testing by NCCLS methodology, the Jarlier double disk test, a disk-on-disk test, a modified three-dimensional test and the E test method for their sensitivity and specificity in detecting TEM- and SHV-related ESBL producers. Three negative and 22 positive controls were studied. These were two Klebsiella pneumoniae and 23 Escherichia coli transconjugants. Seventeen β -lactam antibiotics were tested: cefamandole, cefotetan, cefoxitin, cefuroxime, cefixime, cefoperazone, cefotaxime, cefpodoxime, cefsulodin, ceftazidime, ceftibuten, ceftizoxime, ceftriaxone, moxalactam, cefepime, cefpirome and aztreonam.
Results   NCCLS disk diffusion was 14% sensitive with ceftriaxone, 36% with cefotaxime, 64% with aztreonam, 68% with cefpodoxime, and 73% with ceftazidime. Cefoperazone, cefamandole, cefpodoxime and cefpirome showed 91% sensitivity using the Jarlier test. Using the disk-on-disk test, cefsulodin showed 95% sensitivity, and cefoperazone, cefepime and cefamandole showed 91% sensitivity. With the modified three-dimensional test, cefoperazone, cefpodoxime and cefpirome showed 91% sensitivity.
Conclusions   For practical reasons, we would recommend use of either the Jarlier test or the commercial cephalosporin disks containing clavulanic acid to screen for ESBL producers. Cefoperazone, cefamandole, cefpodoxime and cefpirome showed good sensitivity across the methods tested.     

Effect of β-lactam antibiotics on the in vitro development of resistance in Pseudomonas aeruginosa

Objective   To investigate whether stepwise selection of resistance mutations may mirror the continued bacterial exposure to antibiotics that occurs in the clinical setting.
Methods   We examined the in vitro development of resistance to a number of commonly used antibiotics (cefepime, cefpirome, ceftazidime, cefotaxime, piperacillin and imipenem) in Pseudomonas aeruginosa , a significant nosocomial pathogen. Stepwise resistance was assessed by serial passage of colonies located nearest to the inhibition zone on antibiotic-containing gradient plates.
Results   The lowest frequencies of spontaneous resistance mutations were found with cefepime and imipenem; these drugs also resulted in the slowest appearance of resistance of spontaneous resistance mutations. In five wild-type P. aeruginosa strains, cefepime-selected isolates required a mean of 30 passages to reach resistance; resistance occurred more rapidly in strains selected with other cephalosporins. P. aeruginosa strains that produced β -lactamase or non-enzymatic resistance generally developed resistance more rapidly than wild-type strains. For most strains, resistance to all antibiotics except imipenem correlated with increased levels of β -lactamase activity. Cross-resistance of cephalosporin-selected resistant mutants to other cephalosporins was common. Cephalosporin-resistant strains retained susceptibility to imipenem and ciprofloxacin.
Conclusions   From our in vitro study, we can conclude that the rate of development of resistance of P. aeruginosa is lower with cefepime compared with other cephalosporines.     

A simple and reliable method to screen isolates of Escherichia coli and Klebsiella pneumoniae for the production of TEM- and SHV-derived extended-spectrum β-lactamases

Objective: To evaluate which of 24 β-lactams used in susceptibility tests best discriminated between strains of Klebsiella pneumoniae and Escherichia coli that produce extended spectrum β-lactamases (ESBLs) from strains that produce older, more familiar, plasmid-mediated β-lactamases such as TEM-1 and SHV-1.
Methods: Susceptibility to the 24 β-lactam agents was determined by agar dilution and disk diffusion methodologies, using 27 strains of K. pneumoniae and E. coli that produced 22 different older plasmid-mediated β-lactamases and 28 strains that produced 17 different ESBLs.
Results: In general, strains that produced ESBLs were intermediate or resistant to cefpodoxime, whereas those that produced other β-lactamases were susceptible to this agent. The agar dilution test exhibited 96% sensitivity and 100% specificity in discriminating these two groups of organisms. The disk diffusion test exhibited 100% sensitivity and 96% specificity. All other β-lactam agents tested were inferior discriminators between the two groups of organisms.
Conclusions: Agar dilution and disk diffusion tests with cefpodoxime can be used to discriminate strains of K. pneumoniae and E. coli that produce ESBLs from those that produce older, plasmid-mediated β-lactamases.     

Amoxycillin/clavulanate resistance as related to β-lactamase content in Escherichia coli strains from community-acquired urinary tract infections in Greece

Objective: To determine the resistance rate to amoxycillin/clavulanate (AMC) in 100 Escherichia coli strains isolated from outpatients with urinary tract infection (UTI) in four Greek hospitals and assess the relationship between β-lactamase content and resistance to AMC.
Methods: Susceptibility to β-lactams was determined with the E-test. Sonic cell extracts were used as β-lactamase preparations. Conjugal transfer of resistance was performed in broth cultures. β-Lactamase quantities were evaluated by measuring nitrocefin hydrolysis. Isoelectric points (pls) of β-lactamases were determined by electrofocusing. The substrate specificity of the enzymes and the inhibitory activity of clavulanate were studied spectrophotometrically.
Results: Thirty-two isolates were resistant to ampicillin. Eight were resistant (MIC ≥ 32 mg/L) and 11 showed decreased susceptibility (MIC 4–16 mg/L) to AMC. The latter expressed at least four-fold higher amounts of TEM-1 β-lactamase compared with the TEM-1-producing AMC-susceptible isolates. Seven AMC-resistant isolates produced at least 16-fold higher amounts of TEM-1; in one isolate, resistance was attributed to an OXA-type β-lactamase. None of the AMC-resistant isolates was able to transfer resistance to AMC by conjugation. Clavulanate-resistant TEM variants were not detected.
Conclusions: Amoxycillin/clavulanate-resistant E. coli strains have become established in the Greek community. Resistance is mainly due to the production of large amounts of TEM-1 β-lactamase which is encoded from non-self-transmissible plasmids.     

Evidence for a true post-β-lactamase-inhibitor effect of clavulanic acid against Klebsiella pneumoniae and Haemophilus influenzae

Objective   The purpose of this study was to investigate and characterize in vitro the post- β -lactamase inhibitor effect (PLIE) of clavulanic acid against two β -lactamase-producing species of bacteria.
Methods   The PLIE was investigated against one strain of Klebsiella pneumoniae and one strain of Haemophilus influenzae . A stationary-phase inoculum of about 10⁷ colony-forming units per mL of each bacterium was pre-exposed for 2 h to clavulanic acid, either alone or in combination with amoxicillin at various concentrations. After pre-exposure, the dilution required to remove the β -lactamase inhibitor was 1:100 or 1:1000 according to the bacterial species and their susceptibilities to clavulanic acid. Bacteria were counted hourly after drug removal, on solid agar medium.
Results   Control cultures exposed to amoxicillin alone after dilution, showed a delay in growth, which may be inherent to the time required to synthesize sufficient β -lactamase after the dilution steps. Control experiments clearly distinguished the post-antibiotic effect and the growth delay from the PLIE.
Conclusion   The PLIE could be one of several factors explaining why β -lactam/ β -lactamase inhibitor combinations remain effective throughout the dosing interval, even if a few hours after in vivo administration, serum concentrations of β -lactamase inhibitor fall below levels that are active in vitro.     

Recommendation for treatment of severe infections caused by Enterobacteriaceae producing extended-spectrum β-lactamases (ESBLs)

Extended-spectrum β-lactamase (ESBL)-producing organisms are a global problem. No randomized controlled trials have ever been performed to guide optimal treatment. However, in vitro studies and observational studies strongly suggest that carbapenems (imipenem or meropenem) should be regarded as drugs of choice for serious infections due to ESBL-producing organisms. Other β-lactam antibiotics (cefepime, β-lactam/β-lactamase inhibitor combinations) are not suitable as first-line therapy. The increasing frequency of the association between quinolone resistance and ESBL production have greatly limited the role of this class of antibiotic against ESBL producers.     

Susceptibility of Enterobacteriaceae to β-lactam agents and fluoroquinolones: a 3-year survey in France

Objective   To assess trends in the susceptibility to β -lactam agents and to fluoroquinolones of clinically relevant Enterobacteriaceae isolated over a 3-year period in 14 French hospital laboratories.
Methods   During the second quarter of 1996, 1997 and 1998, 180 consecutive non-duplicate isolates of Enterobacteriaceae were collected in each center. Sixteen β -lactams and four quinolones were tested by the disk diffusion method. In addition, the double-disk synergy test was used to screen for the production of extended-spectrum β -lactamase (ESBL).
Results   Totals of 2507, 2312 and 2506 clinical isolates were obtained in each period, respectively. The distribution of Enterobacteriaceae species according to clinical specimens and wards was similar in each study period. No significant variation in the susceptibility rates to β -lactams and fluoroquinolones was observed, except in Klebsiella pneumoniae and Enterobacter aerogenes. The prevalence of ESBL-producing isolates decreased from 18% to 9% in the former, while it increased from 32% to 54% in the latter. At the same time, the susceptibility to ofloxacin and pefloxacin increased for K. pneumoniae ( P  < 0.003) and cephalosporinase-producing species ( P  < 0.05), except Enterobacter spp.
Conclusion   Over the 3-year study period β -lactams and fluoroquinolones remained highly active against Enterobacteriaceae clinical isolates, with the exception of E. aerogenes , probably as a result of the dissemination of multiresistant clones in French hospitals.     

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.     

Molecular diversity of Proteus mirabilis isolates producing extended-spectrum β-lactamases in a French university hospital

Between February 1997 and December 2002, 3340 hospitalised patients yielded samples positive for Proteus mirabilis, of whom 45 (1.3%) were colonised/infected by P. mirabilis producing extended-spectrum beta-lactamases (ESBLs). The gross incidence of patients colonised/infected by ESBL-producing P. mirabilis was 1.61/10(5) days of hospitalisation, with 20% of isolates being collected from patients in urology wards, most frequently (53.3%) from urine samples. Seventeen (37.7%) of the 43 isolates were obtained from samples collected within 48 h of hospitalisation, indicating that they were community-acquired. Isoelectric focusing assays and sequencing identified the TEM-24, TEM-92 and TEM-52 ESBLs. Pulsed-field gel electrophoresis revealed eight pulsotypes (I-VIII), with the two most common pulsotypes, IV and VI, comprising ten (23.3%) and 12 (26.6%) isolates, respectively. These pulsotypes were considered to represent epidemic strains and spread in various wards of the hospital.     

In vitro selective concentrations of cefepime and ceftazidime for AmpC β-lactamase hyperproducer Enterobacter cloacae variants

Methods: A mixed culture of a wild-type ceftazidime/cefepime-susceptible (4×10 ⁷ CFU/mL) strain and an ampC derepressed Enterobacter cloacae (10 ⁵ CFU/mL) strain (relative proportions 99.75% and 0.25%) was challenged for 4 h with different antibiotic concentrations of ceftazidime and cefepime (0.03–4096 mg/L), and then transferred to drug-free medium. The proportion of wild-type versus derepressed population was evaluated after 24 h.
Results: Ceftazidime and cefepime selected the derepressed variant at concentrations ranging from 1 to 4096 and from 0.12 to 16 mg/L respectively.
Conclusions: These results suggest that serum concentrations attainable with a 2 g/8 h cefepime dosage may be able to suppress the emergence of derepressed ampC mutants.     

Resistance to β-lactams among blood isolates of Salmonella spp. in European hospitals: results from the SENTRY Antimicrobial Surveillance Program 1997–98

The susceptibility to β -lactams and the β -lactamase content of 110 Salmonella spp. blood isolates collected during 1997–98 in 19 European centers participating in the SENTRY Surveillance Program were studied. Thirty-one isolates (28%) were resistant to penicillins, due to production of TEM-1 (27 isolates), OXA-1 (three isolates) or TEM-1 + OXA-1 (one isolate). All OXA-1 producers and 10 TEM-1-producing isolates were also resistant to penicillin–clavulanic acid combinations. In the latter isolates, this phenotype was associated with increased production of TEM-1. Sixteen TEM-1-producing Salmonella Enteritidis isolates and one OXA-1-producing S. Typhimurium isolate were able to transfer β -lactam resistance by conjugation.     

Inducible β-lactamase-mediated resistance to third-generation cephalosporins

The emergence of multiple resistance to β-lactam antimicrobial agents is a major problem in the treatment of patients infected with Enterobacteriaceae that characteristically produce inducible β-lactamases. Inducible and 'derepressed' AmpC β-lactamases are produced by Enterobacter spp., Citrobacter freundii, Serratia marcescens, Morganella morganii and Providencia spp. Resistance to broad-spectrum β-lactams has emerged in 16-44% of these strains from infections treated with one of the newer cephalosporins, even in combination with other antimicrobials. Multiply resistant organisms have spread widely both locally, within hospitals, and nationally. This trend has been shown to correlate closely with the extent of usage of some third-generation cephalosporins. These resistant strains, especially Enterobacter spp., are more regularly isolated from seriously ill patients (especially from respiratory sources), or in intensive care units and pose one of the greatest challenges to contemporary chemotherapy of infections in hospitalized patients. Zwitterionic fourth-generation cephalosporins combine the properties of rapid bacterial outer membrane penetration with high stability to AmpC β-lactamase with good affinity for the penicillin-binding proteins to achieve in vitro activity against AmpC-producing organisms, including the majority of strains highly resistant to ceftazidime and other earlier generation cephalosporins. These features have contributed to their clinical success in the therapy of infections caused by Enterobacter spp. with and without resistance to third-generation compounds. Other alternative agents for chemotherapy of infections due to AmpC β-lactamase-producing strains (inducible or derepressed expression) should also be considered e.g. carbapenems, aminoglycosides and fluoroquinolones.     

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.     

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.     

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.     

Prevalence and diversity of extended-spectrum β-lactamases in faecal Escherichia coli isolates from healthy humans in Spain

Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates were detected in seven of 105 faecal samples from healthy humans, from two Spanish cities, during 2007. In these isolates, five ESBLs were detected, CTX-M-14 ( n  = 2), CTX-M-1 ( n  = 2), CTX-M-32 ( n  = 1), CTX-M-8 ( n  = 1) and TEM-52 ( n  = 1). Both bla _CTX-M-14a (surrounded by IS Ecp1 -IS 903 ) and bla _CTX-M-14b variants (included in an integron structure) were identified in this study. This is the first time that the bla _CTX-M-8 gene and ESBLs of the CTX-M-8 group have been found in Europe and Spain, respectively. Faecal E. coli of healthy humans therefore constitute a reservoir of bla _CTX-M genes with different surrounding genetic elements.     

Transferable plasmid mediating resistance to multiple antimicrobial agents in Klebsiella pneumoniae isolates in Greece

Objective   To investigate the underlying resistance mechanisms in 10 Klebsiella pneumoniae isolates.
Methods   Ten K. pneumoniae strains according to distinct bacteriocin typing and REP-PCR, were examined for their plasmid content, their ability to transfer their resistance to aminoglycosides and third-generation cephalosporins, and their production of aminoglycoside-modifying enzymes and β -lactamases.
Results   Transfer of resistance to the above-mentioned antibiotics as well as to co-trimoxazole and tetracycline in Escherichia coli strain RC 85 at a frequency of 5–10⁶ was achieved for all strains by conjugation. Similar strains harbor a self-transferable multiresistant plasmid (80 kb) with similar Eco RI and Hind III restriction patterns. This plasmid encodes an extended-spectrum β -lactamase which confers high-level resistance to third-generation cephalosporins and aztreonam. It produces SHV-5 β -lactamase, as demonstrated by isoelectric focusing and DNA sequencing. Aminoglycoside resistance was co-transferred, and AAC(6')-I, mediating resistance to gentamicin, tobramycin, netilmicin and amikacin, and AAC(3)-I, mediating resistance to gentamicin and sisomycin, were encoded in all isolates and their transconjugants, while APH(3')-I, mediating resistance to kanamycin and neomycin, was encoded in seven strains.
Conclusions   It appears that a multiresistant transferable plasmid encoding the SHV-5 β -lactamase, causing unusually high resistance to ceftazidime and aztreonam, and the combination AAC(6')-I + AAC(3)-I of acetylating enzymes causing, also resistance to all clinically available aminoglycosides, is established in K. pneumoniae in Greece.     

Susceptibility of extended-spectrum-beta-lactamase-producing Enterobacteriaceae according to the new CLSI breakpoints

In 2010 the Clinical and Laboratory Standards Institute (CLSI) lowered the susceptibility breakpoints of some cephalosporins and aztreonam for Enterobacteriaceae and eliminated the need to perform screening for extended-spectrum β-lactamases (ESBLs) and confirmatory tests. The aim of this study was to determine how many ESBL-producing strains of three common species of Enterobacteriaceae test susceptible using the new breakpoints. As determined with the CLSI screening and confirmatory tests, 382 consecutive ESBL-producing strains were collected at Huashan Hospital between 2007 and 2008, including 158 strains of Escherichia coli, 164 of Klebsiella pneumoniae, and 60 of Proteus mirabilis. Susceptibility was determined by the CLSI agar dilution method. CTX-M-, TEM-, and SHV-specific genes were determined by PCR amplification and sequencing. bla(CTX-M) genes alone or in combination with bla(SHV) were present in 92.7% (354/382) of these ESBL-producing strains. Forty-two (25.6%) strains of K. pneumoniae harbored SHV-type ESBLs alone or in combination. No TEM ESBLs were found. Utilizing the new breakpoints, all 382 strains were resistant to cefazolin, cefotaxime, and ceftriaxone, while 85.0 to 96.7% of P. mirabilis strains tested susceptible to ceftazidime, cefepime, and aztreonam, 41.8 to 45.6% of E. coli strains appeared to be susceptible to ceftazidime and cefepime, and 20.1% of K. pneumoniae were susceptible to cefepime. In conclusion, all ESBL-producing strains of Enterobacteriaceae would be reported to be resistant to cefazolin, cefotaxime, and ceftriaxone by using the new CLSI breakpoints, but a substantial number of ESBL-containing P. mirabilis and E. coli strains would be reported to be susceptible to ceftazidime, cefepime, and aztreonam, which is likely due to the high prevalence of CTX-M type ESBLs.     

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 .