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It has been shown that treatment with titanium dioxide nanoparticles (TiO2 NPs) combined with near-ultraviolet (UV-A) irradiation or in certain dark conditions reduced the numbers of various microorganisms, but the mechanism of this effect remains unclear. In this study to further clarify the mechanism of the antibacterial effect of TiO2 NPs the physiological state of E. coli K12 cells was estimated after incubation with the NPs (0.2 g/L) for different periods of time, with or without UV-A irradiation. Cell incubation with TiO2 NPs, combined or not combined with UV-A irradiation, showed that inactive cells were located only within cell aggregates formed after incubation with TiO2 NPs and that the larger the aggregate, the greater the number of such cells. When the formation of large aggregates was prevented, exposure to NPs under UV-A irradiation failed to result in cell inactivation. A comparative analysis of fluorescence and optical microscopic images of the same aggregates showed that the location of inactivated cells coincided with the zone of increased optical density within the aggregate. After treatment with TiO2 NPs under UV-A for 30, 60, or 120 min cells within the aggregates were the first to be inactivated. Cells on which NPs irradiated more strongly (at the periphery of large aggregates and single) remained active for a longer time than cells within the aggregates. As the time of treatment increased, so did the degree of cell compaction, with some zones of the aggregates eventually transforming into an acellular mass. After UV-A irradiation the cell aggregates spontaneously moved toward each other and gradually fused into larger structures, indicating that such exposure enhanced mutual attraction of cells treated with the NPs. Present study provides evidence for hypothesis that bacterial cells covered with TiO2 NPs are inactivated due to their mutual attraction and consequent compression.

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The antimicrobial effect of TiO2 nanoparticles (TiO2 NPs) has been reliably established. However, the mechanism of this action remains unclear. Currently, according to the dominant hypothesis, it is assumed that the key destructive mechanism is the photocatalytic oxidation of cell components, and the key antimicrobial factor is the reactive oxygen species (ROS) formed on the surface of TiO2 NPs as a result of photocatalysis. In this communication, for the first time, the influence of the electric field created by cationic TiO2 NPs is suggested as a key killing factor, and the key antimicrobial mechanism - the electrical breakdown of the cell membranes and the destruction of the DNA molecule under the action of this field. Here, it was shown by mathematical calculation that TiO2 NPs are able to generate an electric field with a strength sufficient for an electrical breakdown of bilayer lipid membranes, which can lead to a violation of the structure and vital functions of the cytoplasmic membrane (CPM) and to collapse of DNA molecule.GRAPHICAL ABSTRACT