a Department of Internal Medicine, Inselspital, 3010 Berne; b Department of Internal Medicine, Zieglerspital, 3007 Berne; c Institute of Medical Microbiology, University of Berne, Berne, Switzerland
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Abstract |
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Introduction |
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ß-Lactam antibiotics remain the drugs of choice for pneumococcal diseases,6 except when penetration to the site of infection is severely restricted, as is the case in meningitis. Because of treatment failures with cephalosporin monotherapy,7,8 a combination of vancomycin and a cephalosporin is often used for meningitis caused by resistant strains.9,10,11 Reliably active monotherapy would represent a significant advantage to the empirical therapy of meningitis. Cefepime is a broad-spectrum cephalosporin with good activity against the majority of human bacterial pathogens, including penicillin-resistant pneumococci, and good penetration into the CSF.12,13 In the present study we tested cefepime alone and in combination with vancomycin in the rabbit meningitis model and in vitro. Ceftriaxone, alone and combined with vancomycin served as comparison regimens.
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Material and methods |
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The meningitis model, originally described by Dacey and Sande,14 was slightly modified. Briefly, young New Zealand white rabbits weighing 22.5 kg were anaesthetized by im injections of ketamine (30 mg/kg) and xylazine (15 mg/kg), and were immobilized in stereotactic frames for induction of meningitis and CSF samplings. An inoculum containing c. 1 x 105 cfu of penicillin-resistant pneumococci serotype 6 was injected directly into the cisterna magna. The MICs of the antibiotics for the infecting strain were as follows: penicillin G, 4; ceftriaxone, 0.5; vancomycin, 0.120.25; and cefepime, 0.5 mg/L, respectively.
A long-acting anaesthetic (ethylcarbamate = urethane; 3.5 g/rabbit) was injected sc and animals were returned to their cages. Fourteen hours later the cisterna magna was punctured again for periodic CSF sampling before and 0.75, 2.5, 4, 6 and 8 h after initiation of therapy. Antibiotics were administered through a peripheral ear vein as bolus injections at the following concentrations: cefepime, 100 mg/kg; ceftriaxone, 125 mg/kg; and vancomycin, 20 mg/kg. Ceftriaxone was injected once at 0 h and cefepime and vancomycin at 0 and 4 h according to Friedland et al.15 Untreated controls received saline. During the whole experimental period rabbits were kept anesthetized by repeated iv injections of nembutal. At the end of the experiment (8 h) euthanasia was induced by a lethal iv dose of nembutal.
Numbers of bacteria (cfu) were measured by 10-fold serial dilutions of CSF samples, plated on blood agar plates containing 5% sheep blood and incubated overnight at 37°C. In parallel, 20 µL of undiluted CSF were plated (limit of detectability, 50 cfu/mL). Comparisons between different dilutions of CSF were used to exclude significant carryover effects during therapy. The antimicrobial activity of the regimens during the 8 h treatment was calculated by linear regression analysis and expressed as decrease of log10 cfu/mL/h (log10 cfu/mL/h). A value of 1.7 (log10 of the limit of detectability) was assigned to the first sterile CSF sample and a value of 0 to any following sterile sample. The results are expressed as means ± S.D. Statistical significance of differences was determined by the Newman Keuls test.
Measurement of antibiotic levels in the CSF
Antibiotic concentrations in the CSF were determined by the agar diffusion method. Standard curves were performed in saline with 5% rabbit serum in order to mimic CSF protein concentration during meningitis.16 Escherichia coli (ATCC 29522) was used as test strain for ceftriaxone and cefepime13 and Bacillus subtilis (ATCC 6633) for vancomycin.17 The intra- and interday variability of this method was less than 10%. The limit of detection was 0.5, 1 and 1.5 mg/L for vancomycin, ceftriaxone and cefepime, respectively.
In vitro assays
The pneumococcal strain was grown in C + Y medium18 to optical density 0.3 at 590 nm and then diluted 40-fold to 106 cfu/mL, corresponding to the CSF bacterial titre in rabbits before initiation of therapy. Antibiotics were added in concentrations ranging from 0.5 to 5 mg/L corresponding to 1x, 2x, 5x and 10 x MIC of cefepime and ceftriaxone (MIC 0.5 mg/L). Combinations of vancomycin (0.12 mg/L) with cefepime (0.5 mg/L) or ceftriaxone (0.5 mg/L) were also tested. Bacterial numbers were determined at 0, 2, 4 and 6 h by serial dilution of samples, plated on agar plates containing 5% sheep blood and incubated at 37°C for 24 h. Experiments were performed in triplicate and results were expressed as means ± S.D. Synergy was defined as bactericidal effect of a drug combination significantly exceeding the sum of the bactericidal effects of each agent alone.
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Results |
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Discussion |
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Little is known about the effectiveness of cefepime in meningitis caused by resistant pneumococci. In the rabbit model of experimental meningitis, we compared cefepime alone and in combination with vancomycin with the standard therapy consisting of ceftriaxone combined with vancomycin.
The cefepime dose (100 mg/kg) was deliberately chosen in order to mimic the pharmacokinetics of high doses of cefepime in humans (2 g iv).21 Forty-five minutes after intravenous injection, blood levels in rabbits (110 mg/L) were comparable to those reported in humans (109 mg/L). Three hours after injection, the serum level in rabbits (30 mg/L) correlated closely with trough levels measured in humans (33 mg/L).21 This provided the rationale to administer a second dose of cefepime to rabbits at this time. With this dosing regimen, CSF concentrations of cefepime ranged from 10 to 16 mg/L, with ratios of CSF concentration/MIC above 20 during the entire treatment period (Figure 1). These results confirm previous reports of the excellent penetration of cefepime into the subarachnoid space during meningitis (20% versus 9% for ceftriaxone).22,23
The doses of vancomycin (2 x 20 mg/kg) and ceftriaxone (1 x 125 mg/kg) were standard doses that have been used in previous studies in the same model15 and produced serum and CSF concentrations corresponding to high-dose regimens in humans.24,25 In the present study, ceftriaxone CSF concentrations (46 mg/L) corresponded to concentrations described in previous studies.15 Similarly, mean peak CSF concentrations were 4.0 ± 1.7 mg/L after 2 x 20 mg of vancomycin, corresponding closely to the CSF concentration measured in humans.25,26
The reason for the superior killing effect of cefepime compared with ceftriaxone in vivo is not immediately clear, as they had similar antibacterial activity in vitro. The higher ratio of CSF concentration/MIC might conceivably have favoured cefepime over ceftriaxone, since we have previously shown that this pharmacodynamic parameter is a major determinant of bactericidal activity in this model.13
The addition of vancomycin improved the antimicrobial efficacy of both cephalosporins, however, without reaching an extent that qualified as synergy. A synergy between ceftriaxone and vancomycin in vivo and in vitro against resistant pneumococci has been described in previous studies using similar experimental systems to those of the present study.15
In an attempt to confirm in vitro the additive effect of cefepime and vancomycin observed in vivo, we selected concentrations that led to a marginal decrease of bacterial numbers with monotherapies (e.g. concentrations around the MIC). In this setting, the addition of vancomycin to either ceftriaxone or cefepime produced a synergic effect (Figures 5 and 6). To our knowledge, a synergy between cefepime and vancomycin has not been described previously.
The good penetration of cefepime into the inflamed meninges (c. 20%)23 and the excellent bactericidal activity against penicillin-resistant pneumococci qualify cefepime alone or in combination with vancomycin as a potential candidate for the treatment of pneumococcal meningitis caused by strains highly resistant to penicillin. This new combination could be particularly useful in cases where broad antibacterial activity is required in the initial empirical treatment of meningitis. Although our data are preliminary, cefepime with or without vancomycin deserves further clinical evaluation for the therapy of pneumococcal meningitis.
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Acknowledgments |
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Notes |
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References |
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Received 2 March 1999; returned 14 June 1999; revised 4 August 1999; accepted 31 August 1999