Surface-active substances synthesized by Rhodococcus erythropolis ІMВ Ас-5017 during co-cultivation with inducing bacteria either Bacillus subtilis BT-2 or Escherichia coli ІЕM-1 (SASI) had the higher antimicrobial and antiadhesive activities in comparison with surface-active substances synthesized in the medium without cells of inducing bacteria (SAS). Minimum inhibitory concentrations of SASI ranged from 3 to12 μg/mL and were in 4–32 times lower than the same parameter for SAS. Treatment of abiotic surfaces (ceramic, steel, and glass) with SASI decreased adhesion of bacteria Staphylococcus aureus BMC-1 or yeasts Candida albicans D-6 to the level of 10–32 % in comparison with 32–87 % after treatment of surfaces with SAS. Destruction of bacterial and yeast biofilms treated with SASI was in the range from 40 to 94 %, while it was 32–65 % in the case when SAS was used. Increasing of biological activity of surface-active substances synthesized by R. erythropolis in co-culture with inducing bacteria E. coli or B. subtilis was shown for the first time.