Metal nanoparticles, especially silver nanoparticles (AgNPs), have drawn increasing attention for antimicrobial applications. Most studies have emphasized on the correlations between the antibacterial potency of AgNPs and the kinetics of metallic to ionic Ag conversion, while other antimicrobial mechanisms have been underestimated. In this work, we focused on the surface effects of polydopamine (PDA) coating on the antimicrobial activity of AgNPs. A method of fast deposition of PDA was used to synthesize the PDA-AgNPs with controllable coating thickness ranging from 3 to 25 nm. The antimicrobial activities of the PDA-AgNPs were analyzed by fluorescence-based growth curve assays on Escherichia coli. The results indicated that the PDA-AgNPs exhibited significantly higher antibacterial activities than poly­(vinylpyrrolidone)-passivated AgNPs (PVP-AgNPs) and PDA themselves. It was found that the PDA coating synergized with the AgNPs to prominently enhance the potency of the PDA-AgNPs against bacteria. The analysis of X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy elucidated that the synergistic effects could be originated from the interaction/coordination between Ag and catechol group on the PDA coating. The synergistic effects led to increased generation of reactive oxygen species and the consequent bacterial damage. These findings demonstrated the importance of the surface effects on the antimicrobial properties of AgNPs. The underlying molecular mechanisms have shined light on the future development of more potent metal nanoparticle-based antimicrobial agents.

Silver nanoparticles (AgNPs) and ions (Ag+) have recently gained broad attention due to their antimicrobial effects against bacteria and other microbes. In this work, we demonstrate the use of super-resolution fluorescence microscopy for investigating and quantifying the antimicrobial effect of AgNPs at the molecular level. We found that subjecting Escherichia coli (E. coli) bacteria to AgNPs led to nanoscale reorganization of histone-like nucleoid structuring (H-NS) proteins, an essential nucleoid associated protein in bacteria. We observed that H-NS proteins formed denser and larger clusters at the center of the bacteria after exposure to AgNPs. We quantified the spatial reorganizations of H-NS proteins by examining the changes of various spatial parameters, including the inter-molecular distances and molecular densities. Clustering analysis based on Voronoi-tessellation were also performed to characterize the change of H-NS proteins’ clustering behavior. We found that AgNP-treatment led to an increase in the fraction of H-NS proteins forming clusters. Similar effects were observed for bacteria exposed to Ag+ ions, suggesting that the release of Ag+ ions plays an important role in the toxicity of AgNPs. On the other hand, we observed that AgNPs with two surface coatings showed difference in the nanoscale reorganization of H-NS proteins, indicating that particle-specific effects also contribute to the antimicrobial activities of AgNPs. Our results suggested that H-NS proteins were significantly affected by AgNPs and Ag+ ions, which has been overlooked previously. In addition, we examined the dynamic motion of AgNPs that were attached to the surface of bacteria. We expect that the current methodology can be readily applied to broadly and quantitatively study the spatial reorganization of biological macromolecules at the scale of nanometers caused by metal nanoparticles, which are expected to shed new light on the antimicrobial mechanism of metal nanoparticles.

Silver nanoparticles (AgNPs) are well known to exhibit antimicrobial effects through plausible interactions with proteins and/or deoxyribonucleic acid inside the bacteria. Yet a quantitative understanding on the antimicrobial activities of AgNPs remains obscure. Here we conducted in-depth kinetic growth assays and colony-forming unit (CFU) assays on Escherichia coli (E. coli) cultured in AgNPs or Ag ion-containing growth media. Compared to the Ag-absent culture medium, it was found that the growth rate of the bacteria remained unaffected but the lag time of the bacterial growth was extended due to the presence of AgNPs or Ag ions. From the CFU-based time-kill curves, we observed that fractions of E. coli were killed exponentially in the presence of AgNPs or Ag ions. Based on the experimental data, a quantitative model was established to describe the antimicrobial activity of AgNPs. The predictions from this model agree well with the experimental results. We also showed that the parameters in our model as well as their dependence on the concentrations of Ag and bacteria could, in turn, be determined experimentally.It is expected that our quantitative model and the associated parameters provide an alternative means to minimum inhibitory concentration values for characterizing the antimicrobial activities of Ag ions and AgNPs.

Growth curve measurements are commonly used in microbiology, while the use of microplate readers for such measurements provides better temporal resolution and higher throughput. However, evaluating bacterial growth with microplate readers has been hurdled by barriers such as multiple scattering. Here, we report our development of a method based on the time derivatives of the optical density (OD) and/or fluorescence (FL) of bacterial cultures to overcome these barriers. First, we illustrated our method using quantitative models and numerical simulations, which predicted the number of bacteria and the number of fluorescent proteins in time as well as their time derivatives. Then, we systematically investigated how the time derivatives depend on the parameters in the models/simulations, providing a framework for understanding the FL growth curves. In addition, as a demonstration, we applied our method to study the lag time elongation of bacteria subjected to treatment with silver (Ag+) ions and found that the results from our method corroborated well with that from growth curve fitting by the Gompertz model that has been commonly used in the literature. Furthermore, this method was applied to the growth of bacteria in the presence of silver nanoparticles (AgNPs) at various concentrations, where the OD curve measurements failed. We showed that our method allowed us to successfully extract the growth behavior of the bacteria from the FL measurements and understand how the growth was affected by the AgNPs.