The in vitro activity of fifteen antimicrobial agents was tested against 77 Brucella melitensis strains obtained from blood and bone marrow cultures between 1989 and 1992. Doxycycline (MIC90 = 0.125 μg/ml) and tetracycline (MIC90 = 2.0 μg/ml) have excellent in vitro activity; rifampicin (MIC90 = 16.0 μg/ml) and co-trimoxazole (MIC90 = 2.0 μg/ml) are also effective in vitro but with partial response in vivo; and streptomycin (MIC90 = 8.0 μg/ml) has 21.1% of resistant strains. Tetracycline and streptomycin exhibit similar MICs throughout the different periods studied. From the group of fluoroquinolones, ciprofloxacin (MIC90 = 0.25 μg/ml) has the best in vitro activity, and could be a standing alternative for brucellosis treatment. Of the other antimicrobial agents, chloramphenicol (MIC90 = 8.0 μg/ml) and gentamycin (MIC90 = 2.0 μg/ml) have 100% in vitro activity but are not recommended as therapeutic tools for human brucellosis. We noticed no changes in the antibiotic activity pattern of tetracycline against Brucella melitensis in our country during the 25 years of study, which is remarkable since tetracycline continues to be a bactericidal drug with effective therapeutic results.