The potential of metallic copper as an intrinsically antibacterial material is gaining increasing attention in the face of growing antibiotics resistance of bacteria. However, the mechanism of the so-called "contact killing" of bacteria by copper surfaces is poorly understood and requires further investigation. In particular, the influences of bacteria-metal interaction, media composition, and copper surface chemistry on contact killing are not fully understood. In this study, copper oxide formation on copper during standard antimicrobial testing was measured in situ by spectroscopic ellipsometry. In parallel, contact killing under these conditions was assessed with bacteria in phosphate buffered saline (PBS) or Tris-Cl. For comparison, defined Cu2O and CuO layers were thermally generated and characterized by grazing incidence X-ray diffraction. The antibacterial properties of these copper oxides were tested under the conditions used above. Finally, copper ion release was recorded for both buffer systems by inductively coupled plasma atomic absorption spectroscopy, and exposed copper samples were analyzed for topographical surface alterations. It was found that there was a fairly even growth of CuO under wet plating conditions, reaching 4-10 nm in 300 min, but no measurable Cu2O was formed during this time. CuO was found to significantly inhibit contact killing, compared to pure copper. In contrast, thermally generated Cu2O was essentially as effective in contact killing as pure copper. Copper ion release from the different surfaces roughly correlated with their antibacterial efficacy and was highest for pure copper, followed by Cu2O and CuO. Tris-Cl induced a 10-50-fold faster copper ion release compared to PBS. Since the Cu2O that primarily forms on copper under ambient conditions is as active in contact killing as pure copper, antimicrobial objects will retain their antimicrobial properties even after oxide formation.

bBacteria are rapidly killed on copper surfaces, and copper ions released from the surface have been proposed to play a major role in the killing process. However, it has remained unclear whether contact of the bacteria with the copper surface is also an important factor. Using laser interference lithography, we engineered copper surfaces which were covered with a grid of an inert polymer which prevented contact of the bacteria with the surface. Using Enterococcus hirae as a model organism, we showed that the release of ionic copper from these modified surfaces was not significantly reduced. In contrast, killing of bacteria was strongly attenuated. When E. hirae cells were exposed to a solid iron surface, the loss of cell viability was the same as on glass. However, exposing cells to iron in the presence of 4 mM CuSO 4 led to complete killing in 100 min. These experiments suggest that contact killing proceeds by a mechanism whereby the metal-bacterial contact damages the cell envelope, which, in turn, makes the cells susceptible to further damage by copper ions.

Contact killing is a novel term describing the killing of bacteria when they come in contact with metallic copper or copper-containing alloys. In recent years, the mechanism of contact killing has received much attention and many mechanistic details are available. The authors here review some of these mechanistic aspects with a focus on the critical physicochemical properties of copper which make it antibacterial. Known mechanisms of contact killing are set in context to ionic, corrosive, and physical properties of copper. The analysis reveals that the oxidation behavior of copper, paired with the solubility properties of copper oxides, are the key factors which make metallic copper antibacterial. The concept advanced here explains the unique position of copper as an antibacterial metal. Based on our model, novel design criteria for metallic antibacterial materials may be derived.

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Federation bThe well-established killing of bacteria by copper surfaces, also called contact killing, is currently believed to be a combined effect of bacterial contact with the copper surface and the dissolution of copper, resulting in lethal bacterial damage. Iron can similarly be released in ionic form from iron surfaces and would thus be expected to also exhibit contact killing, although essentially no contact killing is observed by iron surfaces. However, we show here that the exposure of bacteria to iron surfaces in the presence of copper ions results in efficient contact killing. The process involves reduction of Cu 2؉ to Cu ؉ by iron; Cu ؉ has been shown to be considerably more toxic to cells than Cu 2؉ . The specific Cu ؉ chelator, bicinchoninic acid, suppresses contact killing by chelating the Cu ؉ ions. These findings underline the importance of Cu ؉ ions in the contact killing process and infer that ironbased alloys containing copper could provide novel antimicrobial materials.T he killing of bacteria by metallic copper surfaces, so-called "contact killing," is now well established and has explicitly been shown for many species (1). Bacteria are killed within minutes on surfaces of copper or copper alloys containing at least 60% copper. In contrast, cells can survive for days on surfaces of stainless steel, glass, or plastics. Copper and copper alloys have attracted attention as a means of creating self-sanitizing surfaces in the light of increasing nosocomial infections in Western hospitals. In a number of hospital trials, rooms have been fitted with copper alloy table tops, bedrails, door handles, light switches, bathroom fixtures, etc., in an effort to curb nosocomial infections (2-5; K. Laitinen et al., unpublished data). These copper surfaces resulted in a 2-to 3-log reduction of the microbial burden on a continuous basis. However, further data are needed to convincingly demonstrate that these measures also lead to a lasting reduction of nosocomial infections. Nevertheless, it appears clear that copper-containing materials can contribute to hospital hygiene and lower the bacterial burden also in other facilities where clean or aseptic working procedures are required (6).The mechanism of contact killing of bacteria by coppercontaining materials is of interest not only in connection to its use in hospitals but also from a purely scientific point of view. Laboratory studies have shown that bacteria on copper surfaces suffer rapid membrane damage and DNA degradation, in addition to other less well-defined cellular damage (7-12). The importance and the order of the different processes leading to cell death may depend on the type of microorganism (9). One key element required for contact killing is the release of copper ions from the metal surface. Bacterial copper resistance systems appear unable to cope with the released copper (13-15). The second important requirement for contact killing is bacterial contact with the metal surface (16). Recently, we showed that bacteria are also killed effectively ...