Nanoparticles released into the environment are attracting increasing concern because of their potential toxic effects. Conventional methods for assessing the toxicity of nanoparticles are usually confined to cultivable cells, but not applicable to viable but non-culturable (VBNC) cells. However, it remains unknown whether silver nanoparticles (AgNPs), a typical antimicrobial agent, could induce bacteria into a VBNC state in natural environments. In this work, the viability of E. coli, an indicator bacterium widely used for assessing the antibacterial activity of AgNPs, was examined through coupling plate counting, fluorescence staining and adenosine triphosphate (ATP) production. AgNPs were found to have a considerable antibacterial ability, which resulted in less than 0.0004% of culturable cells on plates. However, more than 80% of the cells still maintained their cell membrane integrity under the stress of 80 mg/L AgNPs. Meanwhile, the residue of ATP production (0.6%) was 1500 times higher than that of the culturable cells (< 0.0004%). These results clearly demonstrate that when exposed to AgNPs, most of cells fell into a VBNC state, instead of dying. Environmental factors, e.g., Cl- and illumination, which could change the dissolution, hydrophilicity and zeta potential of AgNPs, eventually influenced the culturability of E. coli. Inhibition of dissolved Ag+ and reactive oxygen species was found to facilitate the mitigation of the strain into a VBNC state. Our findings suggest the necessity of re-evaluating the environmental effects and antibacterial activities of AgNPs.
Keywords: Antibacterial activity; Culturability; Escherichia coli; Silver nanoparticle; VBNC.
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