An assay utilizing luminescent bacteria,Photobacteriumphosphoreum, was adapted to assess the antibacterial effects of acoustic energy. Acoustic pressures up to 67 kPa in the 100‐ to 800‐Hz frequency range were applied to bacteria freely suspended in a liquid medium. Bacterial luminescence decreased after sonication, thus showing sensitivity to the effects of acoustic energy. This decreased luminescence was linearly related to exposure duration, appeared independent of acoustic frequency in this range, and was significantly heightened by the presence of cavitation. High‐frequency components of the acoustic emission were recorded from the sonicated fluid, and it was found that the decrease in luminescence due to sonication was directly related to the logarithm of the acoustic emission. Viability studies on exposed bacteria indicated a diminution of luminescence without bacterial death. The potential use of luminescent bacteria in assessing the biological effects of acoustic energy‐generating systems is discussed.