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Role of bacterial motility in differential resistance mechanisms of silver nanoparticles and silver ions

Nature Nanotechnology volume 16, pages 996–1003 (2021)Cite this article

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Abstract

Unlike conventional antimicrobials, the study of bacterial resistance to silver nanoparticles (AgNPs) remains in its infancy and the mechanism(s) through which it evolves are limited and inconclusive. The central question remains whether bacterial resistance is driven by the AgNPs, released Ag(I) ions or a combination of these and other factors. Here, we show a specific resistance in an Escherichia coli K-12 MG1655 strain to subinhibitory concentrations of AgNPs, and not Ag(I) ions, as indicated by a statistically significant greater-than-twofold increase in the minimum inhibitory concentration occurring after eight repeated passages that was maintained after the AgNPs were removed and reintroduced. Whole-population genome sequencing identified a cusS mutation associated with the heritable resistance that possibly increased silver ion efflux. Finally, we rule out the effect of particle aggregation on resistance and suggest that the mechanism of resistance may be enhanced or mediated by flagellum-based motility.

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Fig. 1: Resistance profiles of hypermotile (+IS1) E. coli.
Fig. 2: Resistance profiles of non-motile (ΔflhD::kan) E. coli.
Fig. 3: AgNP aggregation by UV–vis and DLS.

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Data availability

Figures 1–3, Tables 1–3 and all compiled supplementary figures and tables were composed from raw data that are available from the corresponding author. Whole-population genome sequencing data is available through NCBI SRA BioProject ID PRJNA729974.

References

  1. Mijnendonckx, K., Leys, N., Mahillon, J., Silver, S. & Van Houdt, R. Antimicrobial silver: uses, toxicity and potential for resistance. BioMetals 26, 609–621 (2013).

    Article  CAS  Google Scholar 

  2. Molling, J. W., Seezink, J. W., Teunissen, B. E., Muijrers-Chen, I. & Borm, P. J. Comparative performance of a panel of commercially available antimicrobial nanocoatings in Europe. Nanotechnol. Sci. Appl. 7, 97–104 (2014).

    Article  Google Scholar 

  3. Silver nanoparticles market by application (electronics & electrical, healthcare, food & beverages, textiles) and segment forecasts to 2022. GrandView Research (2015); https://www.grandviewresearch.com/industry-analysis/silver-nanoparticles-market

  4. Panacek, A. et al. Silver colloid nanoparticles: synthesis, characterization, and their antibacterial activity. J. Phys. Chem. B 110, 16248–16253 (2006).

    Article  CAS  Google Scholar 

  5. Franci, G. et al. Silver nanoparticles as potential antibacterial agents. Molecules 20, 8856–8874 (2015).

    Article  CAS  Google Scholar 

  6. Duran, N. et al. Silver nanoparticles: a new view on mechanistic aspects on antimicrobial activity. Nanomedicine 12, 789–799 (2016).

    Article  CAS  Google Scholar 

  7. Lara, H. H., Ayala-Nunez, N. V., Turrent, L. & Padilla, C. R. Bactericidal effect of silver nanoparticles against multidrug-resistant bacteria. World J. Microbiol. Biotechnol. 26, 615–621 (2010).

    Article  CAS  Google Scholar 

  8. Graves, J. L. et al. Rapid evolution of silver nanoparticle resistance in Escherichia coli. Front. Genet. 6, 42 (2015).

    Article  CAS  Google Scholar 

  9. Gunawan, C., Teoh, W. Y., Marquis, C. P. & Amal, R. Induced adaptation of Bacillus sp. to antimicrobial nanosilver. Small 9, 3554–3560 (2013).

    Article  CAS  Google Scholar 

  10. Panacek, A. et al. Bacterial resistance to silver nanoparticles and how to overcome it. Nat. Nano. 13, 65–71 (2018).

    Article  CAS  Google Scholar 

  11. Losasso, C. et al. Antibacterial activity of silver nanoparticles: sensitivity of different Salmonella serovars. Front. Microbiol. 5, 227 (2014).

    Article  Google Scholar 

  12. Valentin, E. et al. Heritable nanosilver resistance in priority pathogen: a unique genetic adaptation and comparison with ionic silver and antibiotics. Nanoscale 12, 2384–2392 (2020).

    Article  CAS  Google Scholar 

  13. Gunawan, C. et al. Widespread and indiscriminate nanosilver use: genuine potential for microbial resistance. ACS Nano 11, 3438–3445 (2017).

    Article  CAS  Google Scholar 

  14. Haefeli, C., Franklin, C. & Hardy, K. Plasmid-determined silver resistance in Pseudomonas stutzeri isolated from a silver mine. J. Bacteriol. 158, 389–392 (1984).

    Article  CAS  Google Scholar 

  15. Li, X. Z., Nikaido, H. & Williams, K. E. Silver-resistant mutants of Escherichia coli display active efflux of Ag+ and are deficient in porins. J. Bacteriol. 179, 6127–6132 (1997).

    Article  CAS  Google Scholar 

  16. Gupta, A., Matsui, K., Lo, J. F. & Silver, S. Molecular basis for resistance to silver cations in Salmonella. Nat. Med. 5, 183–188 (1999).

    Article  CAS  Google Scholar 

  17. Silver, S. Bacterial silver resistance: molecular biology and uses and misuses of silver compounds. FEMS Microbiol. Rev. 27, 341–353 (2003).

    Article  CAS  Google Scholar 

  18. Pelgrift, R. Y. & Friedman, A. J. Nanotechnology as a therapeutic tool to combat microbial resistance. Adv. Drug Deliv. Rev. 65, 1803–1815 (2013).

    Article  CAS  Google Scholar 

  19. Wang, L., Hu, C. & Shao, L. The antimicrobial activity of nanoparticles: present situation and prospects for the future. Int J. Nanomed. 12, 1227–1249 (2017).

    Article  CAS  Google Scholar 

  20. Johnston, K. A. et al. Impacts of broth chemistry on silver ion release, surface chemistry composition, and bacterial cytotoxicity of silver nanoparticles. ES: Nano 5, 304–312 (2018).

    CAS  Google Scholar 

  21. Johnston, K. A., Smith, A. M., Marbella, L. E. & Millstone, J. E. Impact of as-synthesized ligands and low-oxygen conditions on silver nanoparticle surface functionalized. Langmuir 32, 3820–3826 (2016).

    Article  CAS  Google Scholar 

  22. Bastus, N. G., Merkoci, F., Piella, J. & Puntes, V. Synthesis of highly monodisperse citrate-stabilized silver nanoparticles of up to 200 nm: kinetic control and catalytic properties. Chem. Mater. 26, 2836–2846 (2014).

    Article  CAS  Google Scholar 

  23. Ivanova, E. P., Bazaka, K. & Crawford, R. J. in New Functional Biomaterials for Medicine and Healthcare Ch. 3, 71–99 (Woodhead, 2014).

  24. Olofsson, S. K. & Cars, O. Optimizing drug exposure to minimize selection of antibiotic resistance. Clin. Infect. Dis. 45, S129–S136 (2007).

    Article  CAS  Google Scholar 

  25. Gullberg, E. et al. Selection of resistant bacteria at very low antibiotic concentrations. PLoS Pathog. 7, e1002158 (2011).

    Article  CAS  Google Scholar 

  26. Adam, M., Murali, B., Glenn, N. O. & Potter, S. S. Epigenetic inheritance based evolution of antibiotic resistance in bacteria. BMC Evol. Biol. 8, 52 (2008).

    Article  CAS  Google Scholar 

  27. George, A. M. & Levy, S. B. Amplifiable resistance to tetracycline, chloramphenicol, and other antibiotics in Escherichia coli: involvement of a non-plasmid-determined efflux of tetracycline. J. Bacteriol. 155, 531–540 (1983).

    Article  CAS  Google Scholar 

  28. Zhao, Y. et al. Small molecule-capped gold nanoparticles as potent antibacterial agents that target Gram-negative bacteria. J. Am. Chem. Soc. 132, 12349–12356 (2010).

    Article  CAS  Google Scholar 

  29. Ji, F. et al. Tetrabromobisphenol A (TBBPA) exhibits specific antimicrobial activity against Gram-positive bacteria without detectable resistance. Chem. Commun. 53, 3512–3515 (2017).

    Article  CAS  Google Scholar 

  30. Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Twentieth Informational Supplement Vol. 30(1) (Approved Standard M100-S20, Clinical and Laboratory Standards Institute, 2010).

  31. Silver, S. in Molecular Biology, Pathogenicity, and Ecology of Bacterial Plasmids (eds Levy, S. B. et al.) 179–189 (Plenum, 1981).

  32. Flynn, K. M., Cooper, T. F., Moore, F. B. G. & Cooper, V. S. The environment affects epistatic interactions to alter the topology of an empirical fitness landscape. PLoS Genet. 9, e1003426 (2013).

    Article  CAS  Google Scholar 

  33. Hall, A. E. et al. Environment changes epistasis to alter trade-offs along alternative evolutionary paths. Evolution 73, 2094–2105 (2019).

    Article  Google Scholar 

  34. Stabryla, L. M., Johnston, K. A., Millstone, J. E. & Gilbertson, L. M. Emerging investigator series: it’s not all about the ion: support for particle-specific contributions to silver nanoparticle antimicrobial activity. ES: Nano 5, 2047–2068 (2018).

    CAS  Google Scholar 

  35. Sandoval-Motta, S. & Aldana, M. Adaptive resistance to antibiotics in bacteria: a systems biology perspective. WIREs Syst. Biol. Med 8, 253–267 (2016).

    Article  Google Scholar 

  36. Normark, B. H. & Normark, S. Evolution and spread of antibiotic resistance. J. Intern. Med. 252, 91–106 (2002).

    Article  CAS  Google Scholar 

  37. Lok, C.-N. et al. Proteomic identification of the cus system as a major determinant of constitutive Escherichia coli silver resistance of chromosomal origin. J. Proteome Res. 7, 2351–2356 (2008).

    Article  Google Scholar 

  38. Knetsch, M. L. & Koole, L. H. New strategies in the development of antimicrobial coatings: the example of increasing usage of silver and silver nanoparticles. Polymers 3, 340–366 (2011).

    Article  CAS  Google Scholar 

  39. Chopra, I. The increasing use of silver-based products as antimicrobial agents: a useful development or a cause for concern? J. Antimicrob. Chemother. 59, 587–590 (2007).

    Article  CAS  Google Scholar 

  40. Bridges, K., Kidson, A., Lowbury, E. J. L. & Wilkins, M. D. Gentamicin- and silver-resistant Pseudomonas in a burns unit. Brit. Med. J. 1, 446–449 (1979).

    Article  CAS  Google Scholar 

  41. Gudipaty, S. A. & McEvoy, M. M. The histidine kinase CusS senses silver ions through direct binding by its sensor domain. Chem. Biochem. 1844, 1656–1661 (2014).

    CAS  Google Scholar 

  42. Affandi, T. & McEvoy, M. M. Mechanism of metal ion-induced activation of a two-component sensor kinase. Biochem. J. 476, 115–135 (2019).

    Article  CAS  Google Scholar 

  43. Cooper, V. S. Experimental evolution as a high-throughput screen for genetic adaptations. mSphere 3, e00121–18 (2018).

    Article  Google Scholar 

  44. Tajkarimi, M. et al. Selection for ionic- confers silver nanoparticle resistance in Escherichia coli. JSM Nanotechnol. Nanomed. 5, 1047 (2017).

    Google Scholar 

  45. Randall, C. P., Gupta, A., Jackson, N., Busse, D. & O’Neill, A. J. Silver resistance in Gram-negative bacteria: a dissection of endogenous and exogenous mechanisms. J. Antimicrob. Chemother. 70, 1037–1046 (2015).

    Article  CAS  Google Scholar 

  46. Koskella, B., Taylor, T. B., Bates, J. & Buckling, A. Using experimental evolution to explore natural patterns between bacterial motility and resistance to bacteriophages. ISME J. 5, 1809–1817 (2011).

    Article  Google Scholar 

  47. Samad, T. et al. Swimming bacteria promote dispersal of non-motile Staphylococcal species. ISME J. 11, 1933–1937 (2017).

    Article  Google Scholar 

  48. Gauger, E. J. et al. Role of motility and the flhDC operon in Escherichia coli MG1655 colonization of the mouse intestine. Infect. Immun. 75, 3315–3324 (2007).

    Article  CAS  Google Scholar 

  49. Barker, C. S., Prub, B. M. & Matsumura, P. Increased motility of Escherichia coli by insertion sequence element integration into the regulatory region of the flhD operon. J. Bacteriol. 186, 7529–7537 (2004).

    Article  CAS  Google Scholar 

  50. Sanchez-Torres, V., Hu, H. & Wood, T. K. GGDEF proteins YeaI, YedQ, and YfiN reduce early biofilm formation and swimming motility in Escherichia coli. Appl. Microbiol. Biotechnol. 90, 651–658 (2011).

    Article  CAS  Google Scholar 

  51. Butler, M. T., Wang, Q. & Harshey, R. M. Cell density and mobility protect swarming bacteria against antibiotics. Proc. Natl Acad. Sci. USA 107, 3776–3781 (2010).

    Article  CAS  Google Scholar 

  52. Lai, S., Tremblay, J. & Deziel, E. Swarming motility: a multicellular behaviour conferring antimicrobial resistance. Environ. Microbiol. 11, 126–136 (2009).

    Article  CAS  Google Scholar 

  53. Sun, E. et al. Broad-spectrum adaptive antibiotic resistance associated with Pseudomonas aeruginosa mucin-dependent surfing motility. Antimicrob. Agents Chemother. 62, e00848–18 (2018).

    Article  CAS  Google Scholar 

  54. Zhang, H. et al. Stress resistance, motility and biofilm formation mediated by a 25kb plasmid pLMSZ08 in Listeria monocytogenes. Food Control 94, 345–352 (2018).

    Article  CAS  Google Scholar 

  55. Asadishad, B., Hidalgo, G. & Tufenkji, N. Pomegranate materials inhibit flagellin gene expression and flagellar-propelled motility of uropathogenic Escherichia coli strain CFT073. FEMS Microbiol. Lett. 334, 87–94 (2012).

    Article  CAS  Google Scholar 

  56. Paramelle, D. et al. A rapid method to estimate the concentration of citrate capped silver nanoparticles from UV-visible light spectra. Analyst 139, 4855–4861 (2014).

    Article  CAS  Google Scholar 

  57. Zhang, W., Crittenden, J., Li, K. & Chen, Y. Attachment efficiency of nanoparticle aggregation in aqueous dispersions: modeling and experimental validation. Environ. Sci. Technol. 46, 7054–7062 (2012).

    Article  CAS  Google Scholar 

  58. Li, X., Lenhart, J. J. & Walker, H. W. Dissolution-accompanied aggregation kinetics of silver nanoparticles. Langmuir 26, 16690–16698 (2010).

    Article  CAS  Google Scholar 

  59. Ma, R. et al. Size-controlled dissolution of organic-coated silver nanoparticles. Environ. Sci. Technol. 46, 752–759 (2012).

    Article  CAS  Google Scholar 

  60. Biggest Threats and Data (Centers for Disease Control and Prevention, 2020); https://www.cdc.gov/drugresistance/biggest-threats.html

  61. Demirdjian, S. et al. Phosphatidylinositol-(3,4,5)-trisphosphate induces phagocytosis of nonmotile Pseudomonas aeruginosa. Infect. Immun. 86, 215–218 (2018).

    Article  Google Scholar 

  62. Deatherage, D. E. & Barrick, J. E. Identification of mutations in laboratory-evolved microbes from next-generation sequencing data using breseq. Methods Mol. Biol. 1151, 165–188 (2014).

    Article  CAS  Google Scholar 

  63. Narayanaswamy, V. P. et al. In vitro activity of novel glycopolymer against clinical isolates of multidrug-resistant Staphylococcus aureus. PLoS One 13, e0191522 (2018).

    Article  CAS  Google Scholar 

  64. Li, X. et al. Functional gold nanoparticles as potent antimicrobial agents against multi-drug-resistant bacteria. ACS Nano 8, 10682–10686 (2014).

    Article  CAS  Google Scholar 

  65. Landman, D., Salamera, J. & Quale, J. Irreproducible and uninterpretable polymyxin B MICs for Enterobacter cloacae and Enterobacter aerogenes. J. Clin. Microbiol. 51, 4106–4111 (2013).

    Article  CAS  Google Scholar 

  66. El-Halfawy, O. M. & Valvano, M. A. Antimicrobial heteroresistance: an emerging field in need of clarity. Clin. Microbiol. Rev. 28, 191–207 (2015).

    Article  CAS  Google Scholar 

  67. Baalousha, M. et al. The concentration-dependent behavior of nanoparticles. Environ. Chem. 13, 1–3 (2015).

    Article  CAS  Google Scholar 

  68. Lecture 11: Antimicrobial Susceptibility Testing—Broth Dilution. Online video clip. Technical Univ. Denmark (n.d); https://www.coursera.org/lecture/antimicrobial-resistance/lecture-11-antimicrobial-susceptibility-testing-broth-dilution-VeNw0

  69. Clinical and Laboratory Standards Institute. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically 9th edn, Vol. 32(2) (Approved Standard M7-A9, Clinical and Laboratory Standards Institute, 2012).

  70. Riley, M. et al. Escherichia coli K-12: a cooperatively developed annotation snapshot—2005. Nucleic Acids Res. 34, 1–9 (2006).

    Article  CAS  Google Scholar 

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Acknowledgements

L.M.S. and L.M.G. acknowledge the generous support from the Department of Defense (DoD) through the National Defense Science and Engineering Graduate Fellowship (NDSEG) Program, the 3M Non-Tenured Faculty Award and the Department of Civil and Environmental Engineering at the University of Pittsburgh. Additionally, K.A.J., N.A.D. and J.E.M. thank the Research Corporation for Science Advancement and the Department of Chemistry at the University of Pittsburgh for their support in carrying out this research. We thank M. J. Fritz for her assistance with the DNA extractions prior to whole-population genome sequencing.

Author information

Authors and Affiliations

  1. Department of Civil and Environmental Engineering, University of Pittsburgh, Pittsburgh, PA, USA

    Lisa M. Stabryla, Sarah-Jane Haig & Leanne M. Gilbertson

  2. Department of Chemistry, University of Pittsburgh, Pittsburgh, PA, USA

    Kathryn A. Johnston, Nathan A. Diemler & Jill E. Millstone

  3. Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, PA, USA

    Vaughn S. Cooper

  4. Department of Chemical and Petroleum Engineering, University of Pittsburgh, Pittsburgh, PA, USA

    Jill E. Millstone & Leanne M. Gilbertson

  5. Department of Mechanical Engineering and Materials Science, University of Pittsburgh, Pittsburgh, PA, USA

    Jill E. Millstone

  6. Department of Environmental and Occupational Health, Graduate School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA

    Sarah-Jane Haig

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  1. Lisa M. Stabryla
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Contributions

L.M.S. and L.M.G. conceived the project and designed the evolutionary experiments dealing with bacteria resistance to AgNPs and aggregation. L.M.S. carried out the microbiological and evolutionary experiments, the synthesis and long-term stability study of AgNPs, the aggregation experiments, the microbial growth kinetics analyses and statistical analyses. K.A.J. and N.A.D. were responsible for the characterizations of the AgNPs (that is, ICP-MS for Ag atom concentration and NMR for ligand density and TEM for particle size, respectively). V.S.C. conducted the genomic variant analysis. S.-J.H. contributed guidance on the microbiological and evolutionary experiments, the analysis of microbial growth kinetics and the statistical analyses. J.E.M. aided in discussion and data interpretation of the AgNP characterization and stability experiments as well as making edits to the manuscript. L.M.S. and L.M.G. wrote the paper with input from all co-authors.

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Correspondence to Lisa M. Stabryla or Leanne M. Gilbertson.

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Stabryla, L.M., Johnston, K.A., Diemler, N.A. et al. Role of bacterial motility in differential resistance mechanisms of silver nanoparticles and silver ions. Nat. Nanotechnol. 16, 996–1003 (2021). https://doi.org/10.1038/s41565-021-00929-w

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