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.     

Natural antibiotic susceptibility of Listeria species: L. grayi, L. innocua, L. ivanovii, L. monocytogenes, L. seeligeri and L. welshimeri strains

Objective   To investigate the natural susceptibility to 71 antimicrobial agents of 103 Listeria strains belonging to all known Listeria species ( L. monocytogenes ( N  = 21), L. innocua ( N  = 21), L. seeligeri ( N  = 21), L. ivanovii ( N  = 19), L. welshimeri ( N  = 11), and L. grayi ( N  = 10)).
Methods   MICs were determined using a microdilution procedure in H-Medium.
Results   All listeriae were naturally sensitive or intermediate to tetracyclines, aminoglycosides, penicillins (except oxacillin), loracarbef, cefazoline, cefaclor, cefotiam, cefoperazone, carbapenems, macrolides, lincosamides, glycopeptides, dalfopristin/quinupristin, chloramphenicol and rifampicin (probably except L. grayi ). Listeria spp. were naturally resistant or intermediate to most 'modern' cephalosporins (cefetamet, cefixime, ceftibuten, ceftazidime, cefdinir, cefpodoxime, cefotaxime, ceftriaxone, cefuroxime), aztreonam, pipemidic acid, dalfopristin quinupristin and sulfamethoxazole. Significant differences in natural susceptibility among the species were seen with the quinolones, trimethoprim, co-trimoxazole, rifampicin, fosfomycin and fusidic acid. It seems likely that L. grayi is naturally resistant to all antifolates; the species was least susceptible to rifampicin and most susceptible to quinolones, whereas L. ivanovii was naturally resistant to most quinolones. L. ivanovii was naturally sensitive to fosfomycin, whereas L. innocua and L. monocytogenes were naturally resistant. L. ivanovii was also the most susceptible species to fusidic acid.
Conclusions   The present study describes a database on the natural susceptibility of Listeria spp. to a wide range of antibiotics, which can be used to validate susceptibility testing results of these microorganisms.     

Cross-reactivity and tolerability of imipenem in patients with delayed-type, cell-mediated hypersensitivity to β-lactams

Background:  Administration of imipenem-cilastatin to patients with IgE-mediated hypersensitivity to β-lactams has always been considered potentially harmful. Recent studies have demonstrated the tolerability of carbapenems (imipenem-cilastatin and meropenem) in patients with IgE-mediated hypersensitivity to β-lactams; there are no studies on this topic regarding patients with cell-mediated allergy to β-lactams. The aim of this study is to assess cross-reactivity and tolerability of imipenem in patients with cell-mediated allergy to β-lactams.
Methods:  From our database we selected 73 patients with cell-mediated allergy to β-lactams, diagnosed by means of immediate-type skin tests, delayed reading intradermal tests, patch tests and detection of specific IgE. Patients with negative patch tests with imipenem-cilastatin underwent an intramuscular test dosing.
Results:  Our patients had a total of 94 nonimmediate reactions to penicillins. All patients had positive patch tests and/or delayed reading intradermal tests for at least one of the penicillin reagent tested and negative immediate-type skin tests and specific IgE. Four patients out of 73 had a positive patch tests to at least one penicillin reagent and imipenem-cilastatin showing cross-reactivity. Sixty-four patients underwent the imipenem-cilastatin intramuscular test dosing and none of them had a clinical reaction.
Conclusions:  Our rate of cross-reactivity between imipenem-cilastatin and other β-lactams was 5.5%. This result is different from previous findings and this may be explained by the fact that we investigated patients with cell-mediated allergy to β-lactams. Patients with cell-mediated allergy to β-lactams should undergo patch tests and a tolerance challenge test before treatment with imipenem-cilastatin.     

Natural antibiotic susceptibility and biochemical profiles of Yersinia enterocolitica-like strains: Y. bercovieri, Y. mollaretii, Y. aldovae and 'Y. ruckeri'.

The natural susceptibility of 54 Yersinia enterocolitica-like strains of Y. bercovieri (formerly Y. enterocolitica biovar 3B, n = 17), Y. mollaretii (formerly Y enterocolitica biovar 3A, n = 12), Y. aldovae (formerly Y. enterocolitica-like group chi2, n = 10) and 'Y ruckeri' (n = 15) was tested to 69 antibiotics. MIC values were determined with a microdilution procedure in IsoSensitest broth for all strains and in cation-adjusted Mueller Hinton broth for some strains. All yersiniae tested showed uniform MIC distributions to most antibiotics and were naturally sensitive or intermediate to aminoglycosides, several cephalosporins, and penicillins, carbapenems, aztreonam, quinolones, tetracyclines, antifolates, chloramphenicol and nitrofurantoin, and naturally resistant to benzylpenicillin, oxacillin, all macrolides except azithromycin, lincosamides, streptogramins, glycopeptides, rifampicin and fusidic acid. Significant differences in susceptibility affecting clinical assessment criteria were seen with aminopenicillins (in the presence and absence of beta-lactamase inhibitors), some cephalosporins (e.g., cefoxitin) and fosfomycin. Whereas strains of Y. aldovae and 'Y. ruckeri' were naturally sensitive or intermediate to amoxicillin and amoxicillin/clavulanate, strains of Y. bercovieri and Y. mollaretii were naturally resistant or naturally resistant or intermediate, respectively. Strains of the two latter species were also highly susceptible to fosfomycin. These data can be valuable for the validation of routine susceptibility test results. beta-Lactam MICs suggest that Y bercovieri and Y. mollaretii strains express chromosomally encoded AmpC beta-lactamases and that most Y. aldovae and 'Y. ruckeri' strains express no, or only small amounts, of enzyme. An evaluation of 30 biochemical tests that determined phenotypic identification to the Yersinia species level is presented.     

Enterobacter cancerogenus (Urosevi?, 1966) Dickey and Zumoff 1988, a senior subjective synonym of Enterobacter taylorae Farmer et al. (1985)

Strains labelled Enterobacter cancerogenus (Erwinia cancerogena) and strains labelled Enterobacter taylorae were found to constitute a single DNA-relatedness group (S1 nuclease hybridization method). Furthermore, no phenotypic test among the conventional and nutritional tests performed could differentiate Enterobacter cancerogenus from Enterobacter taylorae. Therefore, Enterobacter cancerogenus (Urosevi?, 1966) Dickey and Zumoff, 1968, is a senior subjective synonym for Enterobacter taylorae Farmer et al., 1985.     

Antimicrobial susceptibilities of clinical isolates of Haemophilus influenzae and Moraxella catarrhalis collected during 1999–2000 from 13 countries

Objective   To determine antimicrobial activity against Haemophilus influenzae and Moraxella catarrhalis .
Methods   A central laboratory performed NCCLS susceptibility testing for all isolates and β -lactamase and capsular serotype determinations for H. influenzae .
Results   A total of 2712 H. influenzae and 1079 M. catarrhalis were collected . H. influenzae susceptibilities were >90% for amoxicillin/clavulanate, cefaclor, loracarbef, cefprozil, cefuroxime, ciprofloxacin, azithromycin and clarithromycin and were <80% for trimethoprim/sulfamethoxazole and ampicillin. 19.3% were β -lactamase positive. The most common serotype was type-b (5.6%); 86.1% were nontypeable. M. catarrhalis had MIC₉₀ within therapeutic range for all antimicrobials except ampicillin.
Conclusion   The conclusion of the study is that antimicrobials, except ampicillin and trimethoprim/sulfamethoxazole, remain good empiric choices against H. influenzae and M. catarrhalis .     

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 .     

阪崎杆菌侵袭人脑微血管内皮细胞对细胞骨架的影响

目的观察阪崎杆菌侵袭人脑血管内皮细胞后,细胞骨架中微丝和微管的变化。方法用荧光染色技术观察阪崎杆菌侵袭人脑微血管内皮细胞后,细胞骨架中的微丝和微管的变化,观察微丝和微管的抑制剂对阪崎杆菌侵袭人脑微血管内皮细胞的影响。结果阪崎杆菌侵袭人脑微血管内皮细胞后,细胞骨架结构发生明显的改变,微丝和微管部分断裂,模糊,散在分布,微丝抑制剂(细胞松弛素D)和微管抑制剂(秋水仙素)可以抑制阪崎杆菌侵袭人脑微血管内皮细胞。结论阪崎杆菌侵袭人脑血管内皮细胞破坏细胞骨架结构,微丝和微管抑制剂抑制阪崎杆菌侵袭。     

Antibiotic susceptibilities of Haemophilus influenzae in central Scotland

Objective: To ascertain the incidence of antibiotic resistance in Haemophilus influenzae in central Scotland and the β-lactamases produced by these isolates.
Methods: A total of 213 H. influenzae isolates from four medical centers in Scotland [Aberdeen ( n = 58), Edinburgh ( n = 55), Glasgow ( n = 64) and Dundee ( n = 36)] were tested for susceptibility to a range of antimicrobials including β-lactams, β-lactam/β-lactamase-inhibitor combinations, and a representative 4-quinolone, antifolate and macrolide. Susceptibility testing of the β-lactam/β-lactamase-inhibitor combination amoxicillin plus clavulanic acid was conducted at both 2:1 and 4:1 ratios and with clavulanic acid fixed at a concentration of 2 mg/L. Each strain was further investigated for the presence of β-lactamase activity.
Results: Although the incidence of resistance to amoxicillin was 15%, in the presence of clavulanic acid, this resistance was reduced to 4.2%, 5.6% and 4.2% with the 2:1 ratio, 4:1 ratio and 2 mg/L fixed concentration, respectively. Sixteen percent of the isolates demonstrated immediate β-lactamase production. Isoelectric focusing showed that 77.4%, 16.1% and 6.5% of the β-lactamase-positive strains were found to contain TEM-1, VAT-1 and both TEM-1 and VAT-1 β-lactamases, respectively. A further 29% of the strains were recognized as being β-lactamase-positive after prolonged incubation with nitrocephin.
Conclusions: This study suggests that current testing for β-lactamases may underestimate the prevalence of β-lactamase production in H. influenzae.     

Mechanisms of quinolone resistance in clinical isolates of Enterobacter cloacae

Objective: To study and evaluate changes in the gyrA gene and the outer-membrane protein patterns in relation to evolution of resistance against the quinolones in Enterobacter cloacae.
Methods: Strains expressing gyrA -mediated quinolone resistance become susceptible to quinolones upon insertion of the plasmid pNJR3–2. This plasmid (containing wild-type Escherichia coli quinolone-susceptible DNA gyrase A subunits) and pLA2917 (the vector) were introduced into 10 resistant or moderately susceptible clinical isolates of Enterobacter cloacae by conjugation. The transconjugants, the original isolates, the plasmid and the vector control were screened for susceptibility to ofloxacin, ciprofloxacin and sparfloxacin. Additionally, examinations of the outer-membrane proteins were performed.
Results: A reduction of MICs by a factor of 8–32 was found for the transconjugants of five Enterobacter cloacae isolates in the presence of the gene probe, suggesting that these isolates harbored mutations in gyrA. No discernible difference in the patterns of outer-membrane proteins of sensitive and resistant strains could be detected.
Conclusions: It seems that changes in the target site such as alterations in gyrA are important factors leading to a change in the susceptibility of bacteria to the quinolones, whereas there were no evident changes in the outer-membrane proteins to account for evolution of resistance.     

Enzymatic profiles of Enterobacter sakazakii and related species with special reference to the alpha-glucosidase reaction and reproducibility of the test system

The enzymatic profiles of Enterobacter sakazakii, Enterobacter cloacae, Enterobacter aerogenes, and Enterobacter agglomerans were determined with the API ZYM system (API System S.A., La Balme Les Grottes, France). Each assay was performed three times. A simple formula was derived and applied to assess the reproducibility of the API ZYM tests. In addition, a separate alpha-glucosidase test was performed. All E. sakazakii isolates produced alpha-glucosidase, in contrast to the other Enterobacter isolates. No phosphoamidase activity was detected in any of the E. sakazakii isolates, whereas it was present in 72% of E. cloacae, 89% of E. agglomerans, and 100% of E. aerogenes isolates. It was concluded that detection of alpha-glucosidase permits rapid and reliable differentiation between E. sakazakii and other Enterobacter species. The reproducibilities of alpha-glucosidase and phosphoamidase reactions were estimated to be 89 and 81%, respectively.     

Epidemiology of extended-spectrum beta-lactamase-producing Enterobacter isolates in a Spanish hospital during a 12-year period

Fifteen Enterobacter clinical isolates (11 Enterobacter cloacae isolates, 3 Enterobacter aerogenes isolates, and 1 Enterobacter gergoviae isolate), representing 0.4% of all Enterobacter isolates recovered in our hospital from 1989 to 2000, were suspected of harboring an extended-spectrum beta-lactamase (ESBL). These isolates were recovered from 14 different patients. ESBLs were transferred by conjugation into an Escherichia coli recipient strain. Pulsed-field gel electrophoresis (PFGE) revealed a single clone of E. aerogenes and six different clones of E. cloacae. Four of these E. cloacae clonal types were represented by only one isolate each, but the other two were represented by three and four isolates, respectively. Isoelectric focusing, susceptibility phenotyping, PCR analysis, and sequencing demonstrated the presence of three different ESBLs. The most frequent was the recently characterized CTX-M-10 ESBL, which was found in the E. gergoviae isolate and in all but one of the E. cloacae isolates. The remaining E. cloacae isolate harbored a TEM-27 ESBL, and the three E. aerogenes isolates harbored a TEM-24 ESBL. PFGE revealed that our E. aerogenes strain was indistinguishable from the French TEM-24-producing E. aerogenes endemic clone. Although a low prevalence of ESBL-producing Enterobacter isolates was found in our institution over a 12-year period, a diversity of nonepidemic E. cloacae clones was detected, as was the persistence of the CTX-M-10 beta-lactamase. The presence of the TEM-24-producing E. aerogenes French clone in our institution also demonstrates the intercountry dissemination of ESBL-producing isolates.     

Bacteremia associated with Enterobacter sakazakii (yellow, pigmented Enterobacter cloacae).

A case report of bacteremia due to Enterobacter sakazakii, listed previously as yellow-pigmented Enterobacter cloacae (R. Sakazaki, in R. E. Buchanan and N. E. Gibbons, ed., Bergey's Manual of Determinative Bacteriology, 8th ed., p. 325, 1974), occurred in a 7-day-old, Caucasian male who responded successfully to ampicillin therapy. The source of the infection was not known; however, because of the time lapse between birth and the onset of symptoms, the infection was thought to have occurred postnatally.     

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.     

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.     

Enterobacter sakazakii in infants: novel phenomenon in India

E. sakazakii has been implicated in necrotizing enterocolitis, bloodstream and central nervous system infections, with mortality rates of 40-80%. Two cases of E. sakazakii infections; one preterm very low birth weight neonate with meningitis and a two month infant with bacteraemia, are described for the first time in India. The first baby succumbed to the infection while the other responded to appropriate therapy. Powdered infant milk formulae have been implicated in causing neonatal infections and the first baby was on formula feed with classic signs of sepsis and meningitis. The second infant was on breast feed and probably developed nosocomial E. sakazakii bacteraemia.     

Identification of "Cronobacter" spp. (Enterobacter sakazakii)

A taxonomic reclassification of the neonatal pathogen Enterobacter sakazakii to consist of five species within a new genus, "Cronobacter," has recently been proposed. The correct identification of these organisms is important to clinicians. Therefore, using 312 Enterobacteriaceae, including 210 "Cronobacter" strains, the reliabilities of biochemical and genetic confirmation tests were investigated. All "Cronobacter" isolates were positive using dnaG and gluA PCR protocols, and all expressed alpha-glucosidase activity. ID32E v3.0 identified 99.5% of "Cronobacter" isolates (as the nearest match to E. sakazakii).     

Influence of total serum IgE levels on the in vitro detection of β-lactams-specific IgE antibodies

Background Allergic reactions to β-lactams are a frequent cause of adverse drug reactions; the diagnosis is based on history, clinical examination, skin testing (prick and intradermal) and demonstration of serum-specific IgE antibodies (Abs).
Objective We compared the diagnostic performance of the Phadia CAP system for the detection of IgE to β-lactams carried out using the new test with cut-off limits of 0.10 kUA/L and the old test with cut-off limits of 0.35 kUA/L for positive results; subsequently, we analysed the effect of total serum IgE values and of atopic phenotype on the diagnostic performance of the tests.
Methods The study comprised a total of 34 patients with a history of immediate adverse reactions to β-lactams, which were confirmed by positive skin testing, and 115 control subjects with tolerance to β-lactams over the last year. The Phadia CAP System was used for the determination of serum total and specific IgE Abs towards penicilloyl G (c1), penicilloyl V (c2), ampicilloyl (c5) and amoxicilloyl (c6). The overall diagnostic performance was assessed as a diagnostic odds ratio (DOR).
Results The new test showed a higher sensitivity (85% vs. 44%) than the old test and a lower specificity (54% vs. 80%) but the overall diagnostic performance was poor (DOR 6.78 vs. 3.16, P =0.333) in both tests. The total IgE value influences the DOR of both tests; DOR was better for values under 200 kU/L [DOR=66; 95% confidence interval (CI): 11.3–384.1] or 500 kU/L (DOR=45.7; 95% CI: 5.3–394.4) for the new and old tests, respectively.
Conclusions The reduction in the positive cut off value has not significantly improved the overall diagnostic performance of the β-lactams-specific IgE assay. Because of the influence of serum total IgE on the detection of β-lactam-specific IgE Abs, the combination of both tests is mandatory in the in vitro diagnostic approach of β-lactam allergy.     

Comparison of the phenotyping methods ID 32E and VITEK 2 compact GN with 16S rRNA gene sequencing for the identification of Enterobacter sakazakii

A total of 34 isolates (28 Enterobacter sakazakii and 6 Enterobacteriaceae) from infant formulae, milk powder, and the production environment of milk powder factories were identified using ID 32E and VITEK 2 compact GN systems (bioMérieux, France). The ID 32E version 3.0 and VITEK 2 compact GN version 01.01b correctly identified 100% (28) of the Enterobacter sakazakii isolates tested, whereas the previous software version 2.0 for ID 32E showed only 71.4% correct results. None of the non-E. sakazakii isolates tested were misidentified as E. sakazakii with either of the identification systems used.     

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.