Metal ions, not metal-catalyzed oxidative stress, cause clay leachate antibacterial activity

• Published in: PloS one 2014-12, Vol.9 (12), p.e115172
• Main Authors: Otto, Caitlin C, Koehl, Jennifer L, Solanky, Dipesh, Haydel, Shelley E
• Format: Article
• Language: English
• Subjects: Acids; Aluminum Silicates - administration & dosage; Aluminum Silicates - chemistry; Anti-Bacterial Agents - administration & dosage; Anti-Bacterial Agents - chemistry; Antibacterial activity; Antibacterial agents; Antibiotics; Bacteria; Binding sites; Biology and Life Sciences; Cadmium; Catalysis; Cell death; Clay; Deoxyribonucleic acid; DNA; DNA, Bacterial - drug effects; E coli; Earth Sciences; Escherichia coli; Escherichia coli O157 - drug effects; Ethylenediaminetetraacetic acids; Gene expression; Hydrogen peroxide; Hydrogen Peroxide - chemistry; Hydrogen-Ion Concentration; Ions - toxicity; Laboratories; Leachates; Lesions; Life sciences; Lipid peroxidation; Lipid Peroxidation - drug effects; Macromolecules; Medicine and Health Sciences; Metal ions; Metals - toxicity; Oxidation; Oxidation-reduction potential; Oxidative stress; Oxidative Stress - drug effects; Oxygen; Peroxidation; pH effects; Physical Sciences; Proteins; Reactive oxygen species; Reactive Oxygen Species - metabolism; Redox potential; Redox properties; Silver; Staphylococcus aureus; Staphylococcus infections; Stress response; Supplements; Toxicity; Viability
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0115172

Aqueous leachates prepared from natural antibacterial clays, arbitrarily designated CB-L, release metal ions into suspension, have a low pH (3.4-5), generate reactive oxygen species (ROS) and H2O2, and have a high oxidation-reduction potential. To isolate the role of pH in the antibacterial activity of CB clay mixtures, we exposed three different strains of Escherichia coli O157:H7 to 10% clay suspensions. The clay suspension completely killed acid-sensitive and acid-tolerant E. coli O157:H7 strains, whereas incubation in a low-pH buffer resulted in a minimal decrease in viability, demonstrating that low pH alone does not mediate antibacterial activity. The prevailing hypothesis is that metal ions participate in redox cycling and produce ROS, leading to oxidative damage to macromolecules and resulting in cellular death. However, E. coli cells showed no increase in DNA or protein oxidative lesions and a slight increase in lipid peroxidation following exposure to the antibacterial leachate. Further, supplementation with numerous ROS scavengers eliminated lipid peroxidation, but did not rescue the cells from CB-L-mediated killing. In contrast, supplementing CB-L with EDTA, a broad-spectrum metal chelator, reduced killing. Finally, CB-L was equally lethal to cells in an anoxic environment as compared to the aerobic environment. Thus, ROS were not required for lethal activity and did not contribute to toxicity of CB-L. We conclude that clay-mediated killing was not due to oxidative damage, but rather, was due to toxicity associated directly with released metal ions.