In the present study, we conducted a 2 week microcosm experiment with a natural freshwater bacterial community to assess the effects of titanium dioxide nanoparticles (TiO2-NPs) at various concentrations (0, 1, 10 and 100 mg/L) on planktonic and sessile bacteria under dark conditions. Results showed an increase of planktonic bacterial abundance at the highest TiO2-NP concentration, concomitant with a decrease from that of sessile bacteria. Bacterial assemblages were most affected by the 100 mg/L TiO2-NP exposure and overall diversity was found to be lower for planktonic bacteria and higher for sessile bacteria at this concentration. In both compartments, a 100 mg/L TiO2-NPs exposure induced a decrease in the ratio between the Betaproteobacteria and Bacteroidetes. For planktonic communities, a decrease of Comamonadaceae was observed concomitant with an increase of Oxalobacteraceae and Cytophagaceae (especially Emticicia). For sessile communities, results showed a strong decrease of Betaproteobacteria and particularly of Comamonadaceae.