The ethanol extracts obtained from Ballota nigra subsp. anatolica P.H. Davis (Lamiaceae) were investigated for their antimicrobial activities. The antimicrobial activity of the extract was tested against Bacillus subtilis ATCC 6633, Bacillus cereus ATCC 7064, Staphylococcus aureus ATCC 6538P, Escherichia coli ATCC 10538, Proteus vulgaris ATCC 6899, Salmonella typhimurium CCM 5445 and Pseudomonas aeruginosa ATCC 27853, Candida albicans ATCC 10239, Debaryomyces hansenii DSM 70238, Kluyveromyces fragilis ATCC 8608 and Rhodotorula rubra DSM 70403 by disk diffusion method and microdilution method. Some antibacterial and antifungal antibiotics were used as a positive reference standard to determine the sensitivity of the strains. The extracts showed strong antibacterial activity against Escherichia coli, with inhibition zones of 13.0 mm and with minimum inhibitory concentrations (MIC's) and minimum bactericidal concentrations (MBC's) of 250 (500) mu g/mL, respectively. The extracts also exhibited moderate activity against the other test microorganisms. The results demonstrate that the ethanol extract of B. nigra subsp. anatolica has significant activity and suggest that it may be useful in the treatment of infections.