a Dipartimento di Scienze Microbiologiche, Università di Catania, via Androne 81, 95124 Catania and b Direzione Medica GlaxoWellcome S.p.A., Verona, Italy
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Abstract |
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Introduction |
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Treatment of S. maltophilia infection poses some therapeutic challenges owing to its resistance to antibiotics, which has been attributed to decreased outer membrane permeability4 and simultaneous production of two inducible chromosomally encoded ß-lactamases, namely L1 and L2.4,5
Levofloxacin (the l-isomer of the racemic ofloxacin) is a recently developed oral and injectable fluoroquinolone characterized by an expanded spectrum of antibacterial activity, favourable pharmacokinetic properties, clinical efficacy and safety.6
We report here the in vitro activity of levofloxacin on clinical isolates of S. maltophilia.
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Materials and methods |
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Sixty S. maltophilia isolates were isolated from hospitalized patients and identified by the API 20NE system (bioMérieux, France). S. maltophilia NCTC10257 and Psuedomonas aeruginosa ATCC27853 were used as control strains.
Susceptibility tests
Since the susceptibility of S. maltophilia to ß-lactam compounds varies according to the test medium used,7 MICs were determined by the agar dilution method using IsoSensitest (IST) agar (Unipath, Milan, Italy). The following antibiotics and breakpoints were used: levofloxacin (2 mg/L), ceftazidime (
8 mg/L) (GlaxoWellcome S.p.A., Verona, Italy), ciprofloxacin (
1 mg/L), ofloxacin (
2 mg/L), trimethoprim/sulfamethoxazole (co-trimoxazole) (
2/38 mg/L), amikacin (
16 mg/L) (Sigma, Milan, Italy), imipenem (
4 mg/L) (MerckSharp and Dohme, Rome, Italy), ticarcillin/clavulanate (
16 mg/L) (SmithKline Beecham, Milan, Italy) and piperacillin (
16 mg/L) (Wyeth Lederle, Catania, Italy).
Two different inocula, 104 and 106 cfu/spot, were delivered on to the surface of agar plates using a multipoint inoculator. MICs were defined as the lowest concentration of antibiotic that completely suppressed bacterial growth after 18 h incubation at 37°C.
The minimum bactericidal concentrations (MBCs) of levofloxacin, ciprofloxacin and ofloxacin for three S. maltophilia clinical isolates and the NCTC10257 control strain were determined by subculturing broth microdilutions. MBC was defined as the lowest concentration of antimicrobial agent achieving a 99.9% reduction of the original inoculum after 24 h.
Timekill determination
Three S. maltophilia clinical isolates and the control strain NCTC10257 were grown to reach 108 cfu/mL on Mueller Hinton (MH) broth and then diluted to 106 cfu/mL in the same prewarmed broth containing levofloxacin (2 mg/L), ofloxacin (2 mg/L) or ciprofloxacin (1 mg/L). These concentrations were selected according to the susceptibility breakpoint values. An antibiotic-free control was similarly inoculated. At 0, 1, 2, 4, 8 and 24 h after drug exposure, 0.1 mL aliquots were collected, diluted in phosphate-buffered saline, inoculated on to MH agar plates and then incubated at 37°C for 24 h, to determine viable cfu/mL. All experiments were performed in duplicate. Killing curves were constructed by plotting the log10 of cfu/mL versus time. Bactericidal activity was defined as a >3 log10 decrease of the initial inoculum size.
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Results |
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The MBC values of levofloxacin were generally equal to or one dilution higher than the MIC, indicating a consistent bactericidal activity. Ciprofloxacin and ofloxacin showed higher MBC/MIC ratios indicating lower bactericidal power (data not shown).
The Figure shows the results of the timekill experiments with levofloxacin, ciprofloxacin and ofloxacin against the NCTC10257 strain and one clinical isolate (all three clinical isolates gave the same results). Only levofloxacin showed a rapid bactericidal activity against the control and clinical strains and produced a >3 log10 reduction of the inoculum after 24 h, while ciprofloxacin and ofloxacin had a bacteriostatic effect which allowed regrowth after 8 h.
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Discussion |
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ß-Lactams are virtually inactive, mainly because of the production of L1 and L2 ß-lactamases which are rapidly derepressed when S. maltophilia is grown in the presence of ß-lactams.5
In our study, few S. maltophilia clinical isolates were susceptible to amikacin. Mechanisms of resistance to aminoglycosides in this microorganism are complex and poorly delineated. Plasmid-mediated resistance has been described, and might represent a hospital source of aminoglycoside resistance that may be transferable to other Gram-negative bacteria.9
Resistance to fluoroquinolones appears to be supported by different mechanisms. The poor activity of ciprofloxacin against S. maltophilia may be due to alteration of the outer membrane proteins (OMPs).10
This is the first report on the in vitro activity of levofloxacin against S. maltophilia carried out on a significant number of clinical isolates. Levofloxacin proved to exhibit good in vitro activity even with a high inoculum size and/or against ciprofloxacin- and ofloxacin resistant-strains. The more lipophilic nature of levofloxacin and ofloxacin in comparison with ciprofloxacin may favour their permeation across the bacterial outer membrane. Once in the cytoplasm, the double concentration of levofloxacin alone, with respect to ofloxacin (which contains 50% of the bacteriologically inactive d-isomer) contributes to its higher activity. Further, with ofloxacin, the d-isomer (which has a low but significant affinity for topoisomerases) might interfere by competition with the binding of levofloxacin to the DNAtopoisomerase complex.
In conclusion, our in vitro results suggest that levofloxacin may be useful in the treatment of S. maltophilia infections, although evidence from clinical trials is awaited.
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Acknowledgments |
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Notes |
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References |
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2
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5 . Akova, M., Bonfiglio, G. & Livermore, D. M. (1991). Susceptibility to ß-lactam antibiotics of mutant strains of Xanthomonas maltophilia with high- and low-level constitutive expression of L1 and L2 ß-lactamases. Journal of Medical Microbiology 35, 20813.[Abstract]
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7 . Bonfiglio, G. & Livermore, D. M. (1991). Effect of media composition on the susceptibility of Xanthomonas maltophilia to ß-lactam antibiotics. Journal of Antimicrobial Chemotherapy 28, 83742.[Abstract]
8 . Vartivarian, S., Anaissie, E., Bodey, G., Sprigg, H. & Rolston, K. (1994). A changing pattern of susceptibility of Xanthomonas maltophilia to antimicrobial agents: implications for therapy. Antimicrobial Agents and Chemotherapy 38, 6247.[Abstract]
9 . Krcméry, V., Antal, M., Langsàdl, L. & Knothe, H. (1985). Transferable amikacin resistance in Pseudomonas maltophilia and Acinetobacter calcoaceticus. Infection 13, 8990.[ISI][Medline]
10 . Lecso-Bornet, M., Pierre, J., Sarkis-Karam, D., Lubera, S. & Bergogne-Berezin, E. (1992). Susceptibility of Xanthomonas maltophilia to six quinolones and study of outer membrane proteins in resistant mutants selected in vitro. Antimicrobial Agents and Chemotherapy 36, 66971.[Abstract]
Received 27 May 1999; returned 16 August 1999; revised 10 September 1999; accepted 22 September 1999