Methanolic extracts prepared from 8-week-old Maytenus senegalensis in vitro grown root cultures were screened for antibacterial activity against Bacillus subtilis, Staphylococcus aureus, Escherichia coli and Klebsiella pneumoniae. Antibacterial activity was only exhibited against Gram-positive bacterial strains. Root cultures grown on different MS media strengths (1 / 4, 1 / 2, 3 / 4 or full) exhibited antibacterial activity to the same extent. Extracts prepared from roots cultured in the presence of high sucrose concentrations (100 mg l − 1 ) had no antibacterial activity. Roots cultured in the dark exhibited a slightly higher antibacterial activity against S. aureus compared to roots cultured in the light.