The potential eco-toxicity of fourteen different nanosized titanium dioxide (TiO2) particles was studied using Gram-positive Bacillus subtilis and Gram-negative Escherichia coli (ATCC K12) as test organisms. These photosensitive nanoparticles (NPs) were found to be harmful to the organisms studied at different degrees; the antibacterial activity increased with primary particle size, reached the maximum level in the range of 16–20 nm, and then decreased as the primary particle size increased. The presence of light played a significant role on the eco-toxicity of the nano-TiO2 particles under most conditions studied, presumably due to the generation of reactive oxygen species (ROS). However, bacterial growth was inhibited also under dark condition, indicating that mechanisms other than photocatalytic ROS were responsible for the toxic effect. Results highlight the need for caution during the use and disposal of manufactured NPs as to prevent unintended environmental impacts, as well as the importance of further research on the mechanisms and factors that control the toxicity of NPs toward aquatic organisms.