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      Rapid in situ assessment of Cu-ion mediated effects and antibacterial efficacy of copper surfaces

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          Abstract

          Release of metal ions from metal-based surfaces has been considered one of the main drivers of their antimicrobial activity. Here we describe a method that enables parallel assessment of metal ion release from solid metallic surfaces and antimicrobial efficacy of these surfaces in a short time period. The protocol involves placement of a small volume of bioluminescent bacteria onto the tested surface and direct measurement of bioluminescence at various time points. In this study, two recombinant Escherichia coli strains, one expressing bioluminescence constitutively and applicable for general antimicrobial testing, and the other induced by Cu ions, were selected. Decrease in bioluminescence of constitutive E. coli on the surfaces showed a good correlation with the decrease in bacterial viability. Response of Cu-inducible E. coli showed a correlation with Cu content in the tested surfaces but not with Cu dissolution suggesting the role of direct bacteria-surface contact in Cu ion-driven antibacterial effects. In summary, the presented protocol enables the analysis of microbial toxicity and bioavailability of surface-released metal ions directly on solid surfaces within 30–60 min. Although optimized for copper and copper alloy surfaces and E. coli, the method can be extended to other types of metallic surfaces and bacterial strains.

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          The survival of Escherichia coli O157 on a range of metal surfaces.

          Escherichia coli O157:H7 is a serious pathogen causing haemorrhagic colitis. It has been responsible for several large-scale outbreaks in recent years. E. coli O157:H7 is able to survive in a range of environments, under various conditions. The risk of infection from contaminated surfaces is recognised, especially due to the low infectious dose required. In this study, a high concentration (10(7) cells) of E. coli O157 was placed onto different metals and survival time measured. Results showed E. coli O157 to survive for over 28 days at both refrigeration and room temperatures on stainless steel. Copper, in contrast, has strong antibacterial properties (no bacteria can be recovered after only 90 min exposure at 20 degrees C, increasing to 270 min at 4 degrees C) but its poor corrosion resistance and durability make it unsuitable for use as a surface material. Other copper-containing alloys, such as copper nickels and copper silvers, have improved durability and anticorrosion properties and greatly reduce bacterial survival times at these two temperatures (after 120 min at 20 degrees C and 360 min at 4 degrees C, no E. coli could be detected on a copper nickel with a 73% copper content). Use of a surface material with antibacterial properties could aid in preventing cross-contamination events in food processing and domestic environments, if standard hygiene measures fail.
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            Antimicrobial applications of copper.

            Copper has long been known to have antimicrobial activity and is used in drinking water treatment and transportation. It has been recognized by the American Environmental Protection Agency as the first metallic antimicrobial agent in 2008. With ongoing waterborne hospital-acquired infections and antibiotic resistance, research on copper as an antimicrobial agent is again very attractive. Many studies have shown that the use of copper surface and copper particles could significantly reduce the environmental bioburden. This review highlights in its first part all the conditions described in the literature to enhance copper antimicrobial activity. Secondly, the different antimicrobial applications of copper in water treatment, hospital care units and public applications are presented. Finally, the future research needs on copper as an antimicrobial agent are discussed.
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              Glutathione and transition-metal homeostasis in Escherichia coli.

              Glutathione (GSH) and its derivative phytochelatin are important binding factors in transition-metal homeostasis in many eukaryotes. Here, we demonstrate that GSH is also involved in chromate, Zn(II), Cd(II), and Cu(II) homeostasis and resistance in Escherichia coli. While the loss of the ability to synthesize GSH influenced metal tolerance in wild-type cells only slightly, GSH was important for residual metal resistance in cells without metal efflux systems. In mutant cells without the P-type ATPase ZntA, the additional deletion of the GSH biosynthesis system led to a strong decrease in resistance to Cd(II) and Zn(II). Likewise, in mutant cells without the P-type ATPase CopA, the removal of GSH led to a strong decrease of Cu(II) resistance. The precursor of GSH, gamma-glutamylcysteine (gammaEC), was not able to compensate for a lack of GSH. On the contrary, gammaEC-containing cells were less copper and cadmium tolerant than cells that contained neither gammaEC nor GSH. Thus, GSH may play an important role in trace-element metabolism not only in higher organisms but also in bacteria.
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                Author and article information

                Contributors
                C.W.Keevil@soton.ac.uk
                angela.ivask@kbfi.ee
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                25 May 2018
                25 May 2018
                2018
                : 8
                : 8172
                Affiliations
                [1 ]ISNI 0000 0004 0410 6208, GRID grid.177284.f, Laboratory of Environmental Toxicology, , National Institute of Chemical Physics and Biophysics, ; Tallinn, Estonia
                [2 ]ISNI 0000000110107715, GRID grid.6988.f, Department of Natural Sciences, , Tallinn University of Technology, ; Tallinn, Estonia
                [3 ]ISNI 0000 0001 0940 4982, GRID grid.418882.f, Estonian Academy of Sciences, Kohtu 6, ; Tallinn, Estonia
                [4 ]ISNI 0000 0004 1936 9297, GRID grid.5491.9, Faculty of Natural and Environmental Sciences, Centre for Biological Sciences, , University of Southampton, ; Southampton, UK
                Author information
                http://orcid.org/0000-0003-1917-7706
                Article
                26391
                10.1038/s41598-018-26391-8
                5970231
                29802355
                9019de06-fa1b-44b1-b0ca-2f4c0a105c79
                © The Author(s) 2018

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 14 November 2017
                : 11 May 2018
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