Department of Medical Microbiology, Royal Free and University Medical School, Rowland Hill St, London NW3 2PF, UK
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Abstract |
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Introduction |
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Many in-vitro studies have been performed to test these compounds against mycobacteria; several have useful activity against M. tuberculosisalthough their activity against non-tuberculosis mycobacteria is more variable. These studies have been reviewed elsewhere.2,3
Moxifloxacin is an 8-methoxyquinolone which has been shown to have activity against a wide range of bacterial pathogens.4 This agent has been studied in clinical trials for the management of respiratory infections. We tested the activity of moxifloxacin, four other quinolones and isoniazid against a battery of M. tuberculosis and non-tuberculosis mycobacteria to assess its potential usefulness in the management of these infections.
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Materials and methods |
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All bacteria were isolates cultured from specimens submitted to the Academic Department of Microbiology of the Royal Free Hospital, London, UK for investigation of mycobacterial disease. The specimens were cultivated on LowensteinJensen (LJ) medium incorporating pyruvate or glycerol. M. tuberculosis was identified on the basis of microscopic morphology, failure to grow on medium containing para-nitroaminobenzoic acid, and by nitrate and Tween reaction. The Mycobacterium Reference Laboratory of the PHLS at Dulwich, UK performed species identification of non-tuberculosis isolates by conventional biochemical techniques.5 The isolates tested were M. tuberculosis (19), Mycobacterium avium-intracellulare (12), Mycobacterium kansasii (10), Mycobacterium fortuitum (7) and Mycobacterium chelonae (1).
Source of drugs
Antibiotic preparations used in this study were obtained as pure substance from their manufacturers: ciprofloxacin, moxifloxacin (Bayer, Newbury, UK), ofloxacin, levofloxacin (Hoechst Marion Roussel, Denham, UK), sparfloxacin (Rhone Poulenc Rorer, West Malling, UK) and isoniazid (Sigma, Poole, UK).
Minimal inhibitory concentration (MIC) determination
The antibiotic concentration ranges tested were as follows: for M. tuberculosis, ciprofloxacin 0.251 mg/L, levofloxacin 0.120.5 mg/L, sparfloxacin 0.030.12 mg/L and moxifloxacin 0.120.5 mg/L; for M. kansasii similar ranges were used except for moxifloxacin (0.060.5 mg/L); for M. avium-intracellulare, ciprofloxacin and levofloxacin 18 mg/L, sparfloxacin and moxifloxacin 0.252 mg/L. For the rapid growers the range tested for all drugs was 0.0616 mg/L.
BACTEC 460-TB method. This method was used for MIC determinations for M. kansasii and M. avium-intracellulare. Bacteria grown previously on LJ media were sub-cultured on to Middlebrook 7H9 broth until a growth of 0.5 to 1 MacFarland standard was obtained. The broth was then diluted with 0.02% Tween, 0.1% albumin diluent 1:20 for M. kansasii to provide an inoculum of c. 5 x 104 cfu and 1:100 for M. avium to produce an inoculum of c. 104 cfu. These inocula were added to the drug-containing vials. The drug free control was inoculated with a 1:100 dilution of the inoculum used to inoculate the antibiotic-containing vials.6 The MIC was defined as the lowest concentration showing a daily change in growth index through the 1% control.7
The proportion method. This method was used for MIC determinations for M. tuberculosis. Bacteria from LJ slants were subcultured into Middlebrook 7H9 broth (Difco, Oxford, UK) to provide an inoculum. Once growth had reached 0.5 to 1 MacFarland, the broth was diluted 1:100 to provide an inoculum of c. 104 cfu for inoculation on to drug-containing solid Middlebrook 7H10 agar media (Difco). It was diluted 1:10,000 in 0.02% Tween 0.1% albumin diluent for inoculation on to antibiotic free media. Plates were incubated for 23 weeks. The MIC was defined as the lowest concentration showing growth <1% of that of the initial inoculum on the antibiotic free plate.
Agar incorporation MIC. This method was used for MIC determination for M. fortuitum and M. chelonae. Agar incorporation MICs were determined on MüllerHinton agar (Unipath, Basingstoke, UK) supplemented with 10% OADC (Difco). An inoculum of 104 cfu/spot was diluted 1:10 from a 0.5 MacFarland culture and delivered by a Denley multi-point inoculator. Plates were incubated at 30°C for 48 h. The MIC was defined as the lowest concentration of drug to inhibit macroscopically visible colonies.
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Results |
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Against M. tuberculosis, moxifloxacin was more active than ciprofloxacin, levofloxacin and slightly superior to sparfloxacin (Table). The MICs of ciprofloxacin and levofloxacin for M. avium-intracellulare were variable: both sparfloxacin and moxifloxacin showed consistently good activity against this organism. Moxifloxacin was the most active of the quinolones tested against isolates of M. kansasii with an MIC90 of 0.25. Of the rapidly growing mycobacteria tested, a single strain of M. chelonae was highly resistant to all of the fluoroquinolones tested. In contrast only ofloxacin was poorly active against M. fortuitum. Moxifloxacin exhibited the greatest activity against these isolates.
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Discussion |
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Moxifloxacin was the most active of the fluoroquinolones tested against M. tuberculosis, having slightly better activity than sparfloxacin. Previous clinical studies have indicated that a ciprofloxacin-containing regimen lacked sterilizing activity in comparison with a standard protocol,8 but the MICs of moxifloxacin were at least four-fold lower than those of ciprofloxacin. Ciprofloxacin has been shown to have early bactericidal activity against M. tuberculosis (a signifcant fall in cfu in the first 48 h of therapy),9 a characteristic shared only with isoniazid.2,10 The activity demonstrated in the present study suggests that moxifloxacin may also have this characteristic. The evidence of enhanced activity compared with other quinolones and favourable pharmacokinetics of moxifloxacin suggest that it may have a role in anti-tuberculosis therapy in the future.11
The superior activity of moxifloxacin in comparison with other quinolones was also found for the other slow-growing pathogens, M. avium-intracellulare and M. kansasii, as well as for the rapidly growing M. fortuitum. A single strain of M. chelonae was resistant to all of the agents tested. Other studies of moxifloxacin have shown similar activity against mycobacterial species.12
Of particular note is the activity against M. avium-intracellulare. This organism is often difficult to treat and moxifloxacin has markedly better in-vitro activity than the other available agents, ciprofloxacin and levofloxacin. These data encourage further in-vitro and clinical studies to investigate the usefulness of moxifloxacin in the treatment of mycobacterial infections.
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Acknowledgments |
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Notes |
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References |
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2 . Gillespie, S. H. & Kennedy, N. (1998). Fluoroquinolones; a new treatment for tuberculosis? International Journal of Tuberculosis and Lung Diseases 2, 26571.
3 . Yew, W. W., Piddock, L. J. V., Li, M. S. K., Lyon, D., Chan, C. Y. & Cheng, A. F. B. (1994). In-vitro activity of quinolones and macrolides against mycobacteria. Journal of Antimicrobial Chemotherapy34 , 34351.[Abstract]
4 . Woodcock, J. M., Andrews, J. M., Boswell, F. J., Brenwald, N. P. & Wise, R. (1997). In vitro activity of BAY 12-8039, a new fluoroquinolone. Antimicrobial Agents and Chemotherapy 41, 1016.[Abstract]
5 . Collins, C. G., Grange, J. M. & Yates, M. D. (1997). Tuberculosis Bacteriology Organization and Practice, 2nd edn. Butterworth Heinemann, Oxford, UK.
6 . Rastogi, N., Goh, K. S., Bryskier, A. & Devallois, A. (1996). Spectrum of activity of levofloxacin against non-tuberculosis mycobacteria and its activity against the M. avium complex in combination with ethambutol, rifampicin, roxithromycin, amikacin and clofazimine. Antimicrobial Agents and Chemotherapy 40, 24837.[Abstract]
7 . Hawkins, J. E., Wallace, R. J. & Brown, B. A. (1992). Antibacterial susceptibility tests: Mycobacteria. In Manual of Clinical Microbiology, 5th edn (Balows, A., Hansler, W. J., Herrmann, K., Isenberg, H. D. & Shadomy, H. J., Eds), pp. 113852. American Society for Microbiology, Washington, DC.
8 . Kennedy, N., Berger, L., Curram, J., Fox, R., Gutmann, J., Kisyombe, G. M. et al. (1996). Randomized controlled trial of a drug regimen that includes ciprofloxacin for the treatment of pulmonary tuberculosis. Clinical Infectious Diseases 22, 82733.[ISI][Medline]
9 . Jindani, A., Aber, V. R., Edwards, D. A. & Mitchison, D. A. (1980). The early bactericidal activity of drugs in patients with pulmonary tuberculosis. American Review of Respiratory Disease 121, 93949.[ISI][Medline]
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11 . Stass, H. H., Kubitza, D. & Schükly, U. (1997). BAY 12-8039 a new methoxyquinolone: pharmacokinetics, safety, and tolerability of single ascending intraveneous doses in healthy male volunteers. In Program and Abstracts of the Thirty-Seventh Interscience Congress on Antimicrobial Agents and Chemotherapy, Toronto, 1997. Abstract F-153. p. 172. American Society for Microbiology, Washington, DC.
12 . Gross, W. M., Vadney, F. S., Ladutko, L., Bonato, D. A., Campbell, S. I. (1997). In vitro activity of BAY 12-8039, a new 8-methoxyquinolone, against mycobacteria. In Program and Abstracts of the Thirty-Seventh Interscience Congress on Antimicrobial Agents and Chemotherapy, Toronto, 1997. Abstract F-144. p. 170. American Society for Microbiology, Washington, DC.
Received 5 December 1997; returned 21 January 1998; revised 11 February 1999; accepted 26 April 1999