The disc diffusion, Etest and agar dilution techniques were compared to evaluate the antimicrobial susceptibility profile of 70 Stenotrophomonas maltophilia isolates to seven antimicrobial agents. The S. maltophilia isolates were consecutively collected from May 2000 to May 2002 from individual patients, who were hospitalized in a private Brazilian hospital. The antimicrobial susceptibility tests were carried out and interpreted according to the National Committee for Clinical Laboratory Standards (NCCLS) recommendations. The Etest was carried out according to the manufacturer's instructions. There was good agreement among the distinct susceptibility testing results for chloramphenicol, doxycycline, gatifloxacin, trimethoprim-sulfamethoxazole and ticarcillin-clavulanate, suggesting that the disc diffusion and Etest methods are reliable for testing this group of antimicrobials against S. maltophilia. In contrast, a weak correlation was found between the disc diffusion and agar dilution techniques for testing polymyxin B and colistin with unacceptable very major error rates (18.1% and 22.7% for polymyxin B and colistin, respectively). Trimethoprim- sulfamethoxazole (MIC50, 0.06 mg/L; 98.5% susceptible) and gatifloxacin (MIC50, 0.12 mg/L; 98.5% susceptible) were the most potent antimicrobial agents tested against S. maltophilia isolates. In contrast, the worst in vitro activity was found for ticarcillin-clavulanate (MIC50, 16 mg/L; 59.1% susceptible). Although our results confirm that trimethoprim-sulfamethoxazole, gatifloxacin and doxycycline have an excellent in vitro activity against S. maltophilia, further clinical studies are necessary to evaluate the clinical efficacy of these compounds for the treatment of S. maltophilia infections, since no randomized controlled trials have been carried out and no correlation between the clinical response and susceptibility testing results has been reported.