a Unité des Rickettsies CNRS UPRES-A 6020, Faculté de Médecine, Université de la Méditerranée, 27 Boulevard Jean Moulin, 13385 Marseille Cedex 05, France
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Abstract |
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Introduction |
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These pathogens are Gram-negative bacteria that can be grown in vitro in axenic blood-enriched media. MICs have been determined using methods adapted to their fastidious growth and have shown that many antibiotics are bacteriostatic in vitro against Brucella and Bartonella spp.3,4 However, MICs are poorly correlated with the in vivo efficacy of antibiotics in patients suffering from either brucellosis3 or Bartonella-related infections.4 We hypothesized that bactericidal activity of antibiotics against this group of pathogens may be more critical in predicting their efficacy in humans, especially in chronic or relapsing infections. Thus, we have determined MBCs of several antibiotics for these bacteria and tentatively correlate our results with current clinical experience.
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Materials and methods |
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The antibiotics tested were amoxycillin (BeechamSevigne, Paris, France), gentamicin (Dakota Pharm, Creteil, France), streptomycin (Diamant, Puteaux, France), ciprofloxacin (Bayer Pharma, Sebs, France), erythromycin (Abbott, Rungis, France), rifampicin (Cassenne, Puteaux, France) and doxycycline (Pfizer, Neuilly, France).
MIC determination
A modified version of the antibiotic agar dilution method of the National Committee for Clinical Laboratory Standards (NCCLS) was used for determination of MICs, as described previously.4,5 The optimum time for visualization of bacterial growth was 3 days for Brucella spp., 5 days for B. quintana and B. henselae, and 6 days for B. bacilliformis. The MIC was defined as the lowest concentration of the antibiotic tested giving complete inhibition of bacterial growth as compared with a drug-free control.
MBC determination
The bactericidal activity of antibiotics was determined in a broth assay with Schaedler medium (bioMérieux) supplemented with either 5% or 10% sheep blood, respectively, for Brucella and Bartonella spp. A series of glass tubes were filled with 0.9 mL of a bacterial inoculum of 106 cfu/mL and 0.1 mL of two-fold serial dilutions of each antibiotic tested. One tube receiving 0.1 mL of saline served as a growth control. Tubes were incubated for 24 h, according to NCCLS guidelines, at 30°C for B. bacilliformis or 37°C for other species. After incubation for 24 h, 10-fold serial dilutions of the different bacterial suspensions were then plated on to blood agar (bioMérieux) and reincubated for 36 days before enumeration of colonies. The MBC was defined as the lowest concentration of the antibiotic inducing a 99.9% decrease in bacterial inocula following the 24 h incubation period, as compared with the primary inoculum dose.
Controls
Escherichia coli C.I.P. 53126 and Staphylococcus aureus C.I.P. 103811 (Institut Pasteur, Marnes-la-Coquette, France) were used as controls. The medium used was either MuellerHinton or Columbia agar (bioMérieux) enriched with 10% horse blood. MICs were evaluated after 1, 3, 5 and 6 days of incubation. For determination of MBCs, bacterial suspensions (106 cfu/mL) were incubated for 24 h in liquid Schaedler medium at 37°C, with or without horse blood, and in the presence or absence of CO2. After 1 day of exposure to antibiotics, 10-fold serial dilutions of each bacterial suspension were plated on to agar (MuellerHinton or Columbia) and colonies were counted after various incubation times.
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Results |
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Discussion |
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Current knowledge indicates that brucellosis and Bartonella-related infections share comparable microbiological, pathophysiological and clinical findings. Brucellosis and Bartonella spp. infections are characterized mainly by two clinical forms: an acute and a chronic stage. There is a broad spectrum of chronic infections due to Bartonella or Brucella spp.: endocarditis, cutaneous tumour (verruga peruana due to B. bacilliformis and bacillary angiomatosis due to B. henselae and B. quintana) and osteoarticular damage. Combinations of doxycycline plus streptomycin or rifampicin are recommended for human brucellosis,3 and only streptomycin is considered consistently effective for the treatment of verruga peruana.9 Aminoglycosides, especially gentamicin, are also used to treat patients with Bartonella-related endocarditis. In our experience (unpublished results) as well as in that reported in the literature,1 all patients treated with a combination of tetracycline for 6 weeks and aminoglycoside for 2 weeks recovered fully without any relapse, although valve replacement was required for most of the patients, because of extensive valvular damage.10
In conclusion, because human infections with Brucella and Bartonella spp. share many similarities discussed previously, we propose that our current knowledge in treating human brucellosis may help to define the optimum treatment for infections due to Bartonella spp. In particular, the use of an aminoglycoside seems critical in patients with endocarditis.
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Acknowledgments |
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Notes |
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References |
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2 . Corbel, M. J. (1997). Brucellosis: an overview. Emerging Infectious Diseases 3, 21321.[ISI][Medline]
3 . Hall, W. H. (1990). Modern chemotherapy for brucellosis in humans. Review of Infectious Diseases 12, 106099.[ISI][Medline]
4 . Maurin, M., Gasquet, S., Ducco, C. & Raoult, D. (1995). MICs of 28 antibiotic compounds for 14 Bartonella (formerly Rochalimaea) isolates. Antimicrobial Agents and Chemotherapy 39, 238791.[Abstract]
5 . Sobraques, M., Maurin, M., Birtles, R. J. & Raoult, D. (1999). In vitro susceptibilities of four Bartonella bacilliformis strains to 30 antibiotic compounds. Antimicrobial Agents and Chemotherapy 43, 20902.
6 . Rubinstein, E., Lang, R., Shasha, B., Hagar, B., Diamanstein, L., Joseph, G. et al. (1991). In vitro susceptibility of Brucella melitensis to antibiotics. Antimicrobial Agents and Chemotherapy 35, 19257.[ISI][Medline]
7 . Musso, D., Drancourt, M. & Raoult, D. (1995). Lack of bactericidal effect of antibiotics except aminoglycosides on Bartonella (Rochalimaea) henselae. Journal of Antimicrobial Chemotherapy 36, 1018.[Abstract]
8 . Garcia-Rodriguez, J. A., Garcia Sanchez, J. E. & Trujillano, I. (1991). Lack of effective bactericidal activity of new quinolones against Brucella spp. Antimicrobial Agents and Chemotherapy 35, 7569.[ISI][Medline]
9 . Vargas, C. M. (1998). In Bartonellosis or Carrion Disease. New Aspects of an Old Disease, 1st edn, pp. 11722. A.F.A. Editores Importadores S.A., Lima, Peru.
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Drancourt, M., Mainardi, J. L., Brouqui, P., Vandenesch, F., Carta, A., Lehnert, F. et al. (1995). Bartonella (Rochalimaea) quintana endocarditis in three homeless men. New England Journal of Medicine 332, 41923.
Received 15 February 2000; returned 11 May 2000; revised 20 June 2000; accepted 5 July 2000