Multiplex PCR amplification assay for the detection of blaSHV, blaTEM and blaCTX-M genes in Enterobacteriaceae.

Extended-spectrum beta-lactamases (ESBLs) are often mediated by (bla-)SHV, (bla)TEM and (bla)CTX-M genes in Enterobacteriaceae and other Gram-negative bacteria. Numerous molecular typing methods, including PCR-based assays, have been developed for their identification. To reduce the number of PCR amplifications needed we have developed a multiplex PCR assay which detects and discriminates between (bla-)SHV, (bla)TEM and (bla)CTX-M PCR amplicons of 747, 445 and 593 bp, respectively. This multiplex PCR assay allowed the identification of (bla-)SHV, (bla)TEM and (bla)CTX-M genes in a series of clinical isolates of Enterobacteriaceae with previously characterised ESBL phenotype. The presence of (bla)SHV, (bla)TEM and (bla)CTX-M genes was confirmed by partial DNA sequence analysis. Apparently, the universal well-established CTX-M primer pair used here to reveal plasmid-encoded (bla)CTX-M genes would also amplify the chromosomally located K-1 enzyme gene in all Klebsiella oxytoca strains included in the study.

Patterns of mutations in target genes in septicemia isolates of Escherichia coli and Klebsiella pneumoniae with resistance or reduced susceptibility to ciprofloxacin.

Thirty-two Escherichia coli and 21 Klebsiella pneumoniae septicemia isolates with varying degrees of resistance to ciprofloxacin were analyzed for the presence of point mutations within the quinolone-resistance target genes. The number of mutations observed in the resistant isolates agreed with the level of ciprofloxacin resistance in both species. Such isolates were also resistant to nalidixic acid. Isolates with borderline susceptibility to ciprofloxacin, on the other hand, behaved differently in the two species. In E. coli all the isolates harbored at least one mutation and these isolates were also resistant to nalidixic acid, while no mutations were detected in the K. pneumoniae isolates, and susceptibility to nalidixic acid was unpredictable. Therefore, nalidixic acid cannot be used as a class representative. Time-kill curve studies on an isolate with borderline susceptibility from each species showed higher degrees of resistance to ciprofloxacin in comparison to that of the wild-type E. coli. A previously unreported parC mutation, S57-->T, was detected in a resistant E. coli isolate and might expand the QRDR of this gene. Normalized resistance interpretations of histograms confirmed the setting of microbiological zone breakpoints for ciprofloxacin testing.

Cefepime as empirical monotherapy in febrile patients with hematological malignancies and neutropenia: a randomized, single-center phase II trial.

The purpose of this phase II trial was to evaluate the efficacy and safety of cefepime monotherapy in patients with neutropenia expected to last more than 7 days. Sixty-nine patients with neutropenia (<0.5 x 10(9)/1) were randomized during 94 episodes of fever to receive either cefepime monotherapy (n=76) or combination therapy with trimethoprim/sulfamethoxazole plus amikacin (TMP/SMZ plus AMI, n=18). A successful response to cefepime was seen in 31/76 (41%) episodes, with 10/36 (28%) in microbiologically documented infections, 3/4 (75%) in clinically documented infections and 18/36 (50%) in fever of unknown origin. No patient in either treatment group died due to the presenting infection. One patient in the cefepime group discontinued treatment due to a rash. Susceptibility testing of blood isolates by E-test strip showed low MIC values to cefepime for most isolates. It is concluded that cefepime monotherapy appeared both safe and effective as empirical therapy in patients with febrile neutropenia.

Genetic characterization of resistance to extended-spectrum beta-lactams in Klebsiella oxytoca isolates recovered from patients with septicemia at hospitals in the Stockholm area.

Two beta-lactamase gene regions were characterized by DNA sequencing in eight clinical isolates of Klebsiella oxytoca. The blaOXY-2a region encoded a beta-lactamase nearly identical to OXY-2 (one amino acid residue substituted) and conferred aztreonam and cefuroxime resistance on the K. oxytoca isolates. Overproduction of OXY-2a was caused by a G-to-A substitution of the fifth nucleotide in the -10 consensus sequence of blaOXY-2a. The blaOXY-1a was identified in a susceptible strain, and the OXY-1a enzyme differed from OXY-1 by two amino acid residues.

Characterization and nucleotide sequence of a Klebsiella oxytoca cryptic plasmid encoding a CMY-type beta-lactamase: confirmation that the plasmid-mediated cephamycinase originated from the Citrobacter freundii AmpC beta-lactamase.

Plasmid pTKH11, originally obtained by electroporation of a Klebsiella oxytoca plasmid preparation into Escherichia coli XAC, expressed a high level of an AmpC-like beta-lactamase. The enzyme, designated CMY-5, conferred resistance to extended-spectrum beta-lactams in E. coli; nevertheless, the phenotype was cryptic in the K. oxytoca donor. Determination of the complete nucleotide sequence of pTKH11 revealed that the 8,193-bp plasmid encoded seven open reading frames, including that for the CMY-5 beta-lactamase (blaCMY-5). The blaCMY-5 product was similar to the plasmidic CMY-2 beta-lactamase of K. pneumoniae and the chromosomal AmpC of Citrobacter freundii, with 99.7 and 97.0% identities, respectively; there was a substitution of phenylalanine in CMY-5 for isoleucine 105 in CMY-2. blaCMY-5 was followed by the Blc and SugE genes of C. freundii, and this cluster exhibited a genetic organization identical to that of the ampC region on the chromosome of C. freundii; these results confirmed that C. freundii AmpC was the evolutionary origin of the plasmidic cephamycinases. In the K. oxytoca host, the copy number of pTKH11 was very low and the plasmid coexisted with plasmid pNBL63. Analysis of the replication regions of the two plasmids revealed 97% sequence similarity in the RNA I transcripts; this result implied that the two plasmids might be incompatible. Incompatibility of the two plasmids might explain the cryptic phenotype of blaCMY-5 in K. oxytoca through an exclusion effect on pTKH11 by resident plasmid pNBL63.

Postantibiotic effect and postantibiotic sub-mic effect of dirithromycin and erythromycin against respiratory tract pathogenic bacteria.

The postantibiotic effect (PAE) of dirithromycin and erythromycin against strains Streptococcus pyogenes group A M12, NCTC P1800, Streptococcus pneumoniae 23, Staphylococcus aureus Oxford strain 209, Moraxella catarrhalis 15616 and Haemophilus influenzae 5590 was investigated in vitro and in vivo by use of the tissue cage model in rabbits. By exposing strains to 2.5-5 x MIC levels for 6 h or 12 h, both compounds induced in vitro PAEs of 1-9 h, and in two cases >20 h. Cultures in the PAE-phase were then re-exposed to subinhibitory concentrations (0.25 x MIC and 0.5 x MIC) of antibiotic and prolonged suppression of regrowth was obtained for 2->20 h. Following i.v. antibiotic treatment of rabbits (10 mg/kg or 20 mg/kg dirithromycin and 20 mg/kg or 40 mg/kg erythromycin) and bacterial infection of the implanted tissue cages in the same rabbit, the tissue cage fluid (TCF) was sampled 6 h after infection and regrowth was monitored by sampling from new tissue cages in untreated rabbits. These i.v. single doses of both antibiotics induced in vivo PAEs of >6 h, but <20 h against S. pyogenes. Suppression of regrowth in TCF was also obtained for > or = 20 h on infection with exposed S. pyogenes in the PAE-phase in newly implanted tissue cages in rabbits that had been treated with low doses of antibiotic to produce subinhibitory concentrations in the TCE Dirithromycin was in general as active as erythromycin in inducing PAE and in prolonging suppression of bacterial regrowth in the PAE phase.

Incidence of antibiotic resistance in blood and urine isolates from hospitalized patients. Report from a European collaborative study. European Study Group on Antibiotic Resistance (ESGAR).

During 1992-93, 2544 isolates from blood cultures, comprising 52% gram-negative bacilli, 24% Staphylococcus aureus, 15% other staphylococci, 7% Enterococcus faecalis and 2% E. faecium, were consecutively collected and identified in 30 laboratories in 21 European countries. In addition 2512 urine isolates, comprising 82% gram-negative bacilli, 3% S. aureus, 4% other staphylococci and 11% enterococci were collected. The bacteria were sent to 3 laboratories for susceptibility testing by the microdilution method in Mueller-Hinton broth. The MICs of penicillins and aztreonam for all susceptible gram-negative bacilli were 0.25-8 mg/l, penems 0.032-2 mg/l, cefotaxime, ceftazidime and cefpirome or cefepime 0.032-0.25 mg/l, gentamicin, tobramycin and netilmicin 0.125-2 mg/l, amikacin 0.5-4 mg/l, ciprofloxacin 0.016-1 mg/l, trimethoprim 0.25-1 mg/l and tetracycline 1-2 mg/l. For susceptible staphylococci the MICs of erythromycin were 0.25-0.5 mg/l, clindamycin 0.125-0.25 mg/l, methicillin 2-8 mg/l, vancomycin and trimethoprim 1-4 mg/l, ciprofloxacin 0.25-1 mg/l, gentamicin and tobramycin 0.25-1 mg/l. For the enterococci the MICs of ampicillin and vancomycin were 2-4 mg/l and of imipenem, teicoplanin and trimethoprim 0.5-1 mg/l. The antibiotic resistance rates varied between laboratories, being lower in northern Europe, except for the penems, cefpirome and cefepime, which showed uniformly lower resistance rates. Compared to the earlier European studies the resistance rates to beta-lactam antibiotics among the gram-negatives have not changed except with an increase to cefotaxime and ceftazidime in central Europe. Resistance to aminoglycosides had also increased in central Europe from 7-8% to 20-21%, but decreased in southern Europe from 22-24% to 13-14% among the blood isolates and from 12-28% to 6-7% among the urine isolates. There was an increase in resistance to ciprofloxacin and gentamicin in staphylococci from southern Europe. The prevalence of MRSA was significant in central and southern Europe. It is of importance that collaborative national and international studies on the incidence of antibiotic resistance are being performed on a repetitive basis.

Susceptibility testing of Klebsiella spp.--an international collaborative study in quality assessment.

In order to compare the prevalence of antibiotic resistance in different geographical areas, it is necessary to ensure agreement between laboratories on the assignment of strains to 'susceptible' and 'resistant' categories. An international quality assessment was performed to investigate the performance of susceptibility testing of Klebsiella spp. Ninety-five strains of klebsiellae were selected from clinical isolates at the London Hospital Medical College (LHMC). These included strains with a diversity of susceptibility profiles to amoxycillin/clavulanate, piperacillin, ceftazidime, cefuroxime, ciprofloxacin, gentamicin and trimethoprim. The strains were sent to 13 participating laboratories in Europe and the USA and laboratories were asked to test the susceptibility of these strains to these antibiotics by their usual methods. They were also asked to provide details of the method used to test susceptibility. Several different standard recommended testing methods were used. Reporting of susceptibilities was generally accurate, but a number of anomalies were noted. Discrepancies of reporting between the LHMC and the participating laboratories was more marked for resistant strains, particularly in the detection of resistance to cefuroxime and ciprofloxacin, as well as the assignment of susceptibility and resistance to piperacillin and amoxycillin/clavulanate. Some discrepancies could be attributed to the use of different breakpoints, leading to differing assignment of susceptibility. Methodological variations including disc content, inoculum and failure to measure and interpret zone sizes consistently also led to anomalies. This quality assessment programme has helped to identify problems in susceptibility testing which should be investigated further.

Susceptibility testing of Haemophilus influenzae--an international collaborative study in quality assessment.

In order to compare the prevalence of antibiotic resistance in different geographical areas, it is necessary to ensure that agreement is achieved between laboratories on the assignment of strains to 'susceptible' and 'resistant' categories. An international quality assessment study, involving 15 laboratories in eight countries, was performed to investigate the standard of performance of the susceptibility testing of Haemophilus influenzae. One hundred and fifty strains of H. influenzae were distributed from the London Hospital Medical College (LHMC) to all laboratories who were asked to test the susceptibility of the strains to ampicillin, chloramphenicol, tetracycline, trimethoprim, cephalosporins and ciprofloxacin. Laboratories were also asked to provide the details of methodology to test the susceptibility. Significant discrepancy between the LHMC and the participating laboratories appeared in the detection of resistance to ampicillin (especially beta-lactamase-negative strains resistant to ampicillin) as well as the assignment of susceptibility and resistance to chloramphenicol, tetracycline and trimethoprim. Often these reflected the use of inappropriate breakpoints which led to erroneous assignment of susceptibility. Other variations including disc content, medium and supplement, inoculum as well as failure to measure zone sizes properly also led to some repeating anomalies.

Production of a plasmid mediated AmpC-like beta-lactamase by a Klebsiella pneumoniae septicaemia isolate.

A septicaemia Klebsiella pneumoniae isolate from Greece (Ath1) was shown to be resistant to third generation cephalosporins, aztreonam and cefoxitin and this resistance was not decreased in the presence of clavulanic acid. The gene coding for the resistance phenotype, associated with a beta-lactamase showing cephalosporinase activity and a pI of 8.6, could be transferred into Escherichia coli K-12 by conjugation and transformation. DNA-hybridisation showed that this gene was located on two different plasmids, 7.8 and 80 kb respectively. The larger, conjugative plasmid also carried genes coding for another beta-lactamase (pI 6.6) and resistance to aminoglycosides, tetracycline, chloramphenicol and trimethoprim. Under highly stringent conditions the gene coding for the pI 8.6 beta-lactamase hybridised with chromosomal DNA from Citrobacter freundii OS60 but not from E. coli or Enterobacter cloacae. Furthermore, the restriction map of this beta-lactamase gene corresponded to that of ampC from C. freundii OS60. The regulatory ampR gene could not be detected in the plasmid DNA from the Ath1 K. pneumoniae by DNA hybridisation. The relationship between a plasmid-mediated extended spectrum beta-lactamase in a K. pneumoniae septicaemia isolate and chromosomal AmpC beta-lactamase from C. freundii was demonstrated.

Extended-spectrum beta-lactamases in Escherichia coli and Klebsiella spp. in European septicaemia isolates.

Five per cent of Escherichia coli and klebsiella septicaemia isolates from the European Study Group on Antibiotic Resistance (ESGAR) study in 1987 to 1988 showed reduced susceptibility or resistance to cefotaxime, ceftazidime and/oraztreonam. Six of 15 isolates studied were susceptible to cefoxitin and MICs of cefuroxime, cefotaxime, ceftazidime and aztreonam were reduced by clavulanic acid. The isoelectric points of their beta-lactamases were in the range of 5.3-7.6. DNA hybridization showed that four of these beta-lactamases belonged to the TEM or SHV family. Transfer of cefotaxime resistance by conjugation was seen in two of the strains. Nine strains were resistant to cefoxitin (MIC > 16 mg/L) and MICs of cefuroxime, cefotaxime, ceftazidime and aztreonam were only slightly reduced in the presence of clavulanic acid. All nine strains produced at least one beta-lactamase of chromosomal origin with pI > 8.4, and four of these strains also harboured beta-lactamases with a pI range of 6.6-8.2. Cefoxitin resistance could be transferred by conjugation in one strain. Thus E. coli and Klebsiella spp. from the ESGAR septicaemia isolates were found to produce extended-spectrum beta-lactamases of both chromosomal and plasmid origin.

Extended spectrum beta-lactamase from Klebsiella oxytoca, not belonging to the TEM or SHV family.

In clinical isolates of Klebsiella oxytoca resistance to cefuroxime and aztreonam was mediated by a beta-lactamase, designated KH, (pI 5.25) which could be transferred into Escherichia coli by electroporation, but not by conjugation. The transformants produced two enzymes with pIs 5.25 and 8.4 respectively, and showed resistance to cefuroxime, aztreonam, cefotaxime and ceftazidime. Substrate and inhibition profiles indicated that KH beta-lactamase was different from TEM- or SHV-like enzymes, but similar to chromosomal K1 beta-lactamase. The enzyme profile with pI 8.4 was similar to the enzyme from the recipient and showed elevated activity in transformants. The plasmid profiles of the transformants were different from those of their donors. However, a plasmid fragment of the K. oxytoca isolate KH11 hybridized with a plasmid ranging in size from 4.8 to 7.8 kilobases in all the transformants and most of the donors. Gene probes encoding TEM-1 or SHV-1 did not hybridize with plasmid DNA from the K. oxytoca isolates. Furthermore, a probe of the ampC gene did not hybridize with the plasmid but to DNA fragments of the same size in whole cell DNA preparations from the E. coli XAC recipient and the TKH11 transformants. This indicates that no gross rearrangements in the chromosomal beta-lactamase gene region had occurred in the transformants which could explain the increased expression of the pI 8.4 beta-lactamase.

Synergy and cumulated killing effect of the penems FCE 22101 and FCE 25199 in combination with gentamicin against bacteria isolated from septicaemia.

Blood isolates of Enterococcus faecalis, Streptococcus sanguis, Staphylococcus aureus, E. coli and Klebsiella oxytoca were tested for their synergistic and cumulated killing effect (CKE) with the new penems FCE 22101 or FCE 25199 in combination with gentamicin. The tissue cage model in rabbits was used to study the CKE in vivo after antibiotic treatment of the bacteria in vitro. Synergy was observed within two to seven h with all isolates in early logarithmic phase, except with S. aureus, which was rapidly killed by the penems alone. After one h treatment with the antibiotic combinations in vitro, a CKE was demonstrated for up to six h both in vitro and in vivo. The magnitude of the CKE differed between strains and in vitro vs. in vivo.

Characterization of Klebsiella oxytoca septicaemia isolates resistant to aztreonam and cefuroxime.

Eleven clinical isolates of Klebsiella oxytoca from Stockholm hospitals were found to be resistant to aztreonam and cefuroxime, but susceptible to cefotaxime, ceftazidime and imipenem. Resistance could be overcome by combining the beta-lactams with the inhibitor clavulanic acid. Crude beta-lactamase preparations from the isolates inactivated aztreonam and cefuroxime rapidly. By isoelectric focusing, a single common beta-lactamase of pI 5.25 was detected. The K. oxytoca isolates belonged to three subgroups, based on their plasmid profiles and Bg/II restriction endonuclease digestion of plasmid DNA. It was concluded that resistance to aztreonam and cefuroxime in these isolates was conferred by a beta-lactamase distinct from TEM-1, TEM-2 and SHV-1, but possibly derived from TEM-like enzymes.

Characterization of beta-lactam-resistant Klebsiella oxytoca isolated in a neonatal intensive care unit.

The occurrence of Klebsiella oxytoca resistant to ampicillin, piperacillin, aztreonam and cefuroxime in a neonatal intensive care unit, including two cases of septicemia, was shown to consist of a spread on three consecutive occasions caused by three different biochemical Klebsiella oxytoca phenotypes. All isolates, except six surface isolates from one infant belonging to phenotype 1, were sensitive to cefotaxime (MIC 0.5-4 mg/l) and ceftazidime (MIC 0.25-1 mg/l). Isolates of phenotypes 1 and 2 produced a beta-lactamase with an isoelectric point of 5.5 and isolates of phenotype 3, a beta-lactamase with an isoelectric point of 7.9. The beta-lactamases of all three phenotypes hydrolysed benzylpenicillin and more slowly cephalothin. All phenotype 1 isolates carried a 2.9 Md plasmid and most isolates also a 36 Md plasmid. All phenotype 2 isolates carried a 4.8 Md plasmid and one isolate also a 30 Md plasmid. The phenotype 3 isolates carried only one 85 Md plasmid.

Resistance to beta-lactam antibiotics and ciprofloxacin in gram-negative bacilli and staphylococci isolated from blood: a European collaborative study. European Study Group on Antibiotic Resistance.

In 1987 and 1988 members of the European Study Group on Antibiotic Resistance collected 3440 consecutive isolates of Gram-negative bacilli (63%) and staphylococci (37%) from blood cultures and performed susceptibility testing by the microdilution method. The MICs of ampicillin and cefazolin for susceptible Gram-negative bacteria were 1-8 mg/l, of piperacillin less than or equal to 0.5-4 mg/l, of aztreonam, imipenem, cefotaxime and ceftazidime less than or equal to 0.125-1 mg/l and of ciprofloxacin less than or equal to 0.125-0.5 mg/l. For susceptible staphylococci the MICs of cefazolin were less than or equal to 0.5-8 mg/l, of cefotaxime 1-4 mg/l, of ceftazidime 4-16 mg/l, of imipenem and ciprofloxacin less than or equal to 0.125-1 mg/l. The antibiotic resistance rates varied between laboratories, being generally lower in northern Europe, except for imipenem, which showed uniform, low resistance rates. In Escherichia coli resistance to ampicillin, piperacillin and cefazolin could be seen to have increased since a previous survey.

Resistance to aminoglycoside antibiotics in gram-negative bacilli and staphylococci isolated from blood. Report from a European collaborative study. The ESGAR Study Group (European Study Group on Antibiotic Resistance).

The incidence of resistance to gentamicin, tobramycin, amikacin and netilmicin was determined by the microdilution method in Mueller-Hinton broth among blood culture isolates consecutively collected in 37 laboratories in 14 European countries. The distribution of bacteria was similar in each laboratory, Escherichia coli and staphylococci predominating. Resistance levels varied between laboratories but they were higher to all four antibiotics in Southern Europe than in Central and Northern Europe. Aminoglycoside resistance was usually associated with production of aminoglycoside-modifying enzymes, ANT(2"), AAC (3)-V, AAC (6')-I predominating in Gram-negative bacilli and APH (2") + AAC (6') and ANT (4')-I in staphylococci.

Antibiotic susceptibility and beta-lactamase production in clinical isolates of Enterobacter spp.

The in vitro susceptibility of 237 clinical isolates of Enterobacter spp. (E. aerogenes, E. agglomerans and E. cloacae; 41, 64 and 132 respectively) to 16 different antibiotics is described. Four quinolones (ciprofloxacin, lomefloxacin, norfloxacin and ofloxacin), two new cephalosporins (cefpirome and cefepime) and imipenem, all showed high activity against the three Enterobacter species tested (MIC50 less than or equal to 0.125 mg/l, MIC90 less than or equal to 0.5 mg/l). Also the aminoglycosides gentamicin and tobramycin were highly active antibiotics (MIC50 less than or equal to 0.5 mg/l, MIC90 less than or equal to 1.0 mg/l). The susceptibility of beta-lactam-antibiotics to beta-lactamase produced by Enterobacter spp. was evaluated, and imipenem and cefepime were found to be most stable. Different methods for detection of inducible beta-lactamases were used, the agar dilution method being more sensitive than the double-disc diffusion test. Elevated beta-lactamase production was detected, via induction, in 83% of E. aerogenes strains and 70% of E. cloacae strains, with cefamandole used as the substrate and cefoxitin as the inducer. Constitutive, high level enzyme production was detected in 7 and 13% respectively of the E. aerogenes and the E. cloacae strains. In all the strains of E. agglomerans, 10% of E. aerogenes and 13% of E. cloacae, no beta-lactamases could be detected with the methods studied.

Pharmacokinetics in healthy subjects of FCE 22101 and its acetoxymethyl ester, FCE 22891: effect of co-administration of imipenem/cilastatin on the renal metabolism of FCE 22101.

The pharmacokinetics of FCE 22101 were studied in eight healthy male subjects who received FCE 22101 intravenously alone or together with imipenem/cilastatin which was given to inhibit dehydropeptidase-I, a renal enzyme metabolizing penem and carbapenem antibiotics. The kinetics of FCE 22101 were also studied following oral administration of its acetoxymethyl ester, FCE 22891. For comparative purposes, the kinetics of imipenem and cilastatin, given alone or together with FCE 22101, were calculated. Intravenously administered FCE 22101 at a dose of 250 mg gave peak plasma concentrations of about 12 mg/l and the plasma half-life was about 60 min. Co-administration of FCE 22101 with imipenem/cilastatin did not affect the plasma kinetics of FCE 22101, nor did FCE 22101 influence the kinetics of imipenem or cilastatin. Cilastatin increased the urinary recovery of FCE 22101 from 17.5% to 53.0% with FCE 22101 alone to 73.2% to 91.8% when it was given with cilastatin. There was a high correlation between the urinary recovery of FCE 22101 in this study and that of imipenem given alone to the same subjects in previous studies; subjects who were high metabolizers of imipenem were also high metabolizers of FCE 22101. When FCE 22891 was given orally at a dose of 500 mg (corresponding to 400 mg of FCE 22101 free acid), peak concentrations of 2.2 to 6.1 mg/l were found. The absorption was rapid with peak concentrations achieved 20 to 80 min after administration. In comparison with imipenem, FCE 22101 seems to undergo less non-renal metabolism.

Postantibiotic effect of the penem FCE 22101 against selected gram-positive and gram-negative bacteria in vitro and in vivo by the use of a tissue cage model in rabbits.

Isolates of Staphylococcus aureus, Streptococcus pneumoniae, Haemophilus influenzae and Klebsiella pneumoniae were tested for their bactericidal activity and postantibiotic effect (PAE) with the new penem FCE 22101. The tissue cage model in rabbits was used to study PAE in vivo. The bactericidal activity against all four species was shown to be in the range of 0.05-4.0 mg/l. A 99.9% killing effect at MBC concentrations was reached within 2 hours with S. pneumoniae and K. pneumoniae and within 6-8 hours with S. aureus and H. influenzae. After in vitro exposure by FCE 22101 a PAE in vitro and in vivo was obtained against S. aureus, S. pneumoniae and H. influenzae strains but no PAE could be demonstrated against K. pneumoniae. FCE 22101 showed a good bactericidal activity and PAE against the strains investigated, except for K. pneumoniae.