1887

Abstract

Applying self-sanitizing copper surfaces to commonly touched places within hospital facilities is an emerging strategy to prevent healthcare-associated infections. This is due to the fact that bacterial pathogens are rapidly killed on copper, a process termed contact killing. However, the mechanisms of contact killing are not fully understood, and the potential of bacterial pathogens to develop resistance has rarely been explored. Here, we hypothesize that bacteria are predominantly killed by a burst release of toxic copper ions, resulting from chemical reactions between bacterial cell surface components and metallic copper. To test this, we isolated and characterized small colony variants (SCVs) derived from and . SCVs overproduce extracellular polymeric substances (EPS), which will enhance copper ion release, causing more rapid death on copper. Indeed, all 13 SCVs tested were more rapidly killed than wild-types on the surfaces of both pure copper and brass (63.5 % Cu). Next, using the non-pathogenic SBW25 as a model, we examined the roles of specific cell surface components in contact killing, including EPS, LPS, capsule, flagella and pili. We also subjected SBW25 to daily serial passage of sub-lethal conditions on brass. After 100 transfers, there was a slight increase of survival rate, but ~97 % of cells can still be killed within 60 min on brass. Together, our data implicate that the rate of contact killing on copper is largely determined by the cell surface components, and bacteria have limited ability to evolve resistance to metallic copper.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.000348
2016-10-18
2020-02-29
Loading full text...

Full text loading...

/deliver/fulltext/jmm/65/10/1143.html?itemId=/content/journal/jmm/10.1099/jmm.0.000348&mimeType=html&fmt=ahah

References

  1. Alvarez E., Uslan D. Z., Malloy T., Sinsheimer P., Godwin H.. 2016; It is time to revise our approach to registering antimicrobial agents for health care settings. Am J Infect Control44:228–232 [CrossRef][PubMed]
    [Google Scholar]
  2. Baker-Austin C., Wright M. S., Stepanauskas R., McArthur J. V.. 2006; Co-selection of antibiotic and metal resistance. Trends Microbiol14:176–182 [CrossRef][PubMed]
    [Google Scholar]
  3. Berg J., Tom-Petersen A., Nybroe O.. 2005; Copper amendment of agricultural soil selects for bacterial antibiotic resistance in the field. Lett Appl Microbiol40:146–151 [CrossRef][PubMed]
    [Google Scholar]
  4. Biswas L., Biswas R., Schlag M., Bertram R., Gotz F.. 2009; Small-colony variant selection as a survival strategy for Staphylococcus aureus in the presence of Pseudomonas aeruginosa. Appl Environ Microb 75:6910–6912 [CrossRef]
    [Google Scholar]
  5. Chen G., Chen X., Yang Y., Hay A. G., Yu X., Chen Y.. 2011; Sorption and distribution of copper in unsaturated Pseudomonas putida CZ1 biofilms as determined by X-ray fluorescence microscopy. Appl Environ Microb 77:4719–4727 [CrossRef]
    [Google Scholar]
  6. Collins T. J.. 2007; ImageJ for microscopy. Biotechniques43:25 [CrossRef][PubMed]
    [Google Scholar]
  7. de Carvalho C. C., Caramujo M. J.. 2014; Bacterial diversity assessed by cultivation-based techniques shows predominance of Staphylococccus species on coins collected in Lisbon and Casablanca. FEMS Microbiol Ecol88:26–37 [CrossRef][PubMed]
    [Google Scholar]
  8. Elguindi J., Wagner J., Rensing C.. 2009; Genes involved in copper resistance influence survival of Pseudomonas aeruginosa on copper surfaces. J Appl Microbiol106:1448–1455 [CrossRef][PubMed]
    [Google Scholar]
  9. Ferguson G. C., Bertels F., Rainey P. B.. 2013; Adaptive divergence in experimental populations of Pseudomonas fluorescens. V. Insight into the niche specialist fuzzy spreader compels revision of the model Pseudomonas radiation. Genetics195:1319–1335 [CrossRef][PubMed]
    [Google Scholar]
  10. Ghafoor A., Hay I. D., Rehm B. H. A.. 2011; Role of exopolysaccharides in Pseudomonas aeruginosa biofilm formation and architecture. Appl Environ Microb77:5238–5246 [CrossRef]
    [Google Scholar]
  11. Grass G., Rensing C., Solioz M.. 2011; Metallic copper as an antimicrobial surface. Appl Environ Microb77:1541–1547 [CrossRef]
    [Google Scholar]
  12. Graves N., Nicholls T. M., Morris A. J.. 2003; Modeling the costs of hospital-acquired infections in New Zealand. Infect Control Hosp Epidemiol24:214–223 [CrossRef][PubMed]
    [Google Scholar]
  13. Hans M., Mathews S., Mücklich F., Solioz M.. 2016; Physicochemical properties of copper important for its antibacterial activity and development of a unified model. Biointerphases11:018902 [CrossRef]
    [Google Scholar]
  14. Hao Z., Lou H., Zhu R., Zhu J., Zhang D., Zhao B. S., Zeng S., Chen X., Chan J. et al. 2014; The multiple antibiotic resistance regulator MarR is a copper sensor in Escherichia coli. Nat Chem Biol10:21–28 [CrossRef][PubMed]
    [Google Scholar]
  15. Ji X., Shen Q., Liu F., Ma J., Xu G., Wang Y., Wu M.. 2012; Antibiotic resistance gene abundances associated with antibiotics and heavy metals in animal manures and agricultural soils adjacent to feedlots in Shanghai; China. J Hazard Mater235-236:178–185 [CrossRef][PubMed]
    [Google Scholar]
  16. Johns B. E., Purdy K. J., Tucker N. P., Maddocks S. E.. 2015; Phenotypic and genotypic characteristics of small colony variants and their role in chronic infection. Microbiol Insights8:15–23 [CrossRef][PubMed]
    [Google Scholar]
  17. Karpanen T. J., Casey A. L., Lambert P. A., Cookson B. D., Nightingale P., Miruszenko L., Elliott T. S.. 2012; The antimicrobial efficacy of copper alloy furnishing in the clinical environment: a crossover study. Infect Control Hosp Epidemiol33:3–9 [CrossRef][PubMed]
    [Google Scholar]
  18. Kleanthous C., Armitage J. P.. 2015; The bacterial cell envelope. Philos Trans R Soc Lond B Biol Sci370:20150019 [CrossRef][PubMed]
    [Google Scholar]
  19. Latimer J., Forbes S., McBain A. J.. 2012; Attenuated virulence and biofilm formation in Staphylococcus aureus following sublethal exposure to triclosan. Antimicrob Agents Chemother56:3092–3100 [CrossRef][PubMed]
    [Google Scholar]
  20. Liu Y., Rainey P. B., Zhang X. X.. 2015; Molecular mechanisms of xylose utilization by Pseudomonas fluorescens: overlapping genetic responses to xylose, xylulose, ribose and mannitol. Mol Microbiol98:553–570 [CrossRef][PubMed]
    [Google Scholar]
  21. Malone J. G.. 2015; Role of small colony variants in persistence of Pseudomonas aeruginosa infections in cystic fibrosis lungs. Infect Drug Resist8:237–247 [CrossRef][PubMed]
    [Google Scholar]
  22. Marais F., Mehtar S., Chalkley L.. 2010; Antimicrobial efficacy of copper touch surfaces in reducing environmental bioburden in a South African community healthcare facility. J Hosp Infect74:80–82 [CrossRef][PubMed]
    [Google Scholar]
  23. Mathews S., Hans M., Mucklich F., Solioz M.. 2013; Contact killing of bacteria on copper is suppressed if bacterial-metal contact is prevented and is induced on iron by copper ions. Appl Environ Microb79:2605–2611 [CrossRef]
    [Google Scholar]
  24. McDonald M. J., Gehrig S. M., Meintjes P. L., Zhang X. X., Rainey P. B.. 2009; Adaptive divergence in experimental populations of Pseudomonas fluorescens. IV. genetic constraints guide evolutionary trajectories in a parallel adaptive radiation. Genetics183:1041–1053 [CrossRef][PubMed]
    [Google Scholar]
  25. Michels H. T., Keevil C. W., Salgado C. D., Schmidt M. G.. 2015; From laboratory research to a clinical trial: copper alloy surfaces kill bacteria and reduce hospital-acquired infections. Herd-Health Env Res9:64–79 [CrossRef]
    [Google Scholar]
  26. Mikolay A., Huggett S., Tikana L., Grass G., Braun J., Nies D. H.. 2010; Survival of bacteria on metallic copper surfaces in a hospital trial. Appl Microbiol Biot87:1875–1879 [CrossRef]
    [Google Scholar]
  27. Molteni C., Abicht H. K., Solioz M.. 2010; Killing of bacteria by copper surfaces involves dissolved copper. Appl Environ Microb76:4099–4101 [CrossRef]
    [Google Scholar]
  28. Muller M. P., MacDougall C., Lim M.. Ontario Agency for Health Protection and Promotion Public Health Ontario Provincial Infectious Diseases Advisory Committee on Infection Prevention and Control 2016; Antimicrobial surfaces to prevent healthcare-associated infections: a systematic review. J Hosp Infect92:7–13 [CrossRef][PubMed]
    [Google Scholar]
  29. O'Gorman J., Humphreys H.. 2012; Application of copper to prevent and control infection. where are we now?. J Hosp Infect81:217–223 [CrossRef][PubMed]
    [Google Scholar]
  30. O'Toole G. A.. 2011; Microtiter dish biofilm formation assay. J Vis Exp47:2437 [CrossRef][PubMed]
    [Google Scholar]
  31. Pachter B., Kozer L., Pachter S., Weiner M.. 1997; Dirty money? a bacteriologic investigation of US currency. Infect Med14:574
    [Google Scholar]
  32. Rainey P. B., Desprat N., Driscoll W. W., Zhang X. X.. 2014; Microbes are not bound by sociobiology: response to Kümmerli and Ross-Gillespie (2013). Evolution68:3344–3355 [CrossRef][PubMed]
    [Google Scholar]
  33. Rensing C., Grass G.. 2003; Escherichia coli mechanisms of copper homeostasis in a changing environment. FEMS Microbiol Rev27:197–213 [CrossRef][PubMed]
    [Google Scholar]
  34. Salgado C. D., Sepkowitz K. A., John J. F., Cantey J. R., Attaway H. H., Freeman K. D., Sharpe P. A., Michels H. T., Schmidt M. G.. 2013; Copper surfaces reduce the rate of healthcare-acquired infections in the intensive care unit. Infect Control Hosp Epidemiol34:479–486 [CrossRef][PubMed]
    [Google Scholar]
  35. Santo C. E., Taudte N., Nies D. H., Grass G.. 2008; Contribution of copper ion resistance to survival of Escherichia coli on metallic copper surfaces. Appl Environ Microbiol74:977–986 [CrossRef][PubMed]
    [Google Scholar]
  36. Santo C. E., Morais P. V., Grass G.. 2010; Isolation and characterization of bacteria resistant to metallic copper surfaces. Appl Environ and Microb76:1341–1348[CrossRef]
    [Google Scholar]
  37. Schmidt M. G., Attaway H. H., Sharpe P. A., John J., Sepkowitz K. A., Morgan A., Fairey S. E., Singh S., Steed L. L. et al. 2012; Sustained reduction of microbial burden on common hospital surfaces through introduction of copper. J Clin Microbiol50:2217–2223 [CrossRef][PubMed]
    [Google Scholar]
  38. Schmidt M. G., von Dessauer B., Benavente C., Benadof D., Cifuentes P., Elgueta A., Duran C., Navarrete M. S.. 2016; Copper surfaces are associated with significantly lower concentrations of bacteria on selected surfaces within a pediatric intensive care unit. Am J Infect Control44:203–209 [CrossRef][PubMed]
    [Google Scholar]
  39. Schmitz F. J., von Eiff C., Gondolf M., Fluit A. C., Verhoef J., Peters G., Hadding U., Heinz H. P., Jones M. E.. 1999; Staphylococcus aureus small colony variants: rate of selection and MIC values compared to wild-type strains, using ciprofloxacin, ofloxacin, levofloxacin, sparfloxacin and moxifloxacin. Clin Microbiol Infect5:376–378 [CrossRef][PubMed]
    [Google Scholar]
  40. Silby M. W., Cerdeño-Tárraga A. M., Vernikos G. S., Giddens S. R., Jackson R. W., Preston G. M., Zhang X. X., Moon C. D., Gehrig S. M. et al. 2009; Genomic and genetic analyses of diversity and plant interactions of Pseudomonas fluorescens. Genome Biol10:R51 [CrossRef][PubMed]
    [Google Scholar]
  41. Simões A. S., Couto I., Toscano C., Gonçalves E., Póvoa P., Viveiros M., Lapão L. V.. 2016; Prevention and control of antimicrobial resistant healthcare-associated infections: the microbiology laboratory rocks!. Front Microbiol7:855 [CrossRef][PubMed]
    [Google Scholar]
  42. Stover C. K., Pham X. Q., Erwin A. L., Mizoguchi S. D., Warrener P., Hickey M. J., Brinkman F. S., Hufnagle W. O., Kowalik D. J. et al. 2000; Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic pathogen. Nature406:959–964 [CrossRef][PubMed]
    [Google Scholar]
  43. Vu B., Chen M., Crawford R. J., Ivanova E. P.. 2009; Bacterial extracellular polysaccharides involved in biofilm formation. Molecules14:2535–2554 [CrossRef][PubMed]
    [Google Scholar]
  44. Warnes S. L., Caves V., Keevil C. W.. 2012; Mechanism of copper surface toxicity in Escherichia coli O157:H7 and Salmonella involves immediate membrane depolarization followed by slower rate of DNA destruction which differs from that observed for Gram-positive bacteria. Environ Microbiol14:1730–1743 [CrossRef][PubMed]
    [Google Scholar]
  45. Zhang X., Rainey P. B.. 2007a; Construction and validation of a neutrally-marked strain of Pseudomonas fluorescens SBW25. J Microbiol Methods71:78–81 [CrossRef]
    [Google Scholar]
  46. Zhang X. X., Rainey P. B.. 2007b; The role of a P1-type ATPase from Pseudomonas fluorescens SBW25 in copper homeostasis and plant colonization. Mol Plant-Microbe Interact20:581–588[CrossRef]
    [Google Scholar]
  47. Zhang X. X., Rainey P. B.. 2008; Regulation of copper homeostasis in Pseudomonas fluorescens SBW25. Environ Microbiol10:3284–3294[CrossRef]
    [Google Scholar]
  48. Zhang X. X., Rainey P. B.. 2013; Exploring the sociobiology of pyoverdin-producing Pseudomonas. Evolution67:3161–3174 [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.000348
Loading
/content/journal/jmm/10.1099/jmm.0.000348
Loading

Data & Media loading...

Supplements

Supplementary File 1

PDF
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error