UNRAVELING THE MODE OF ACTION FOR A NATURAL ANTIBACTERIAL CLAY OF LACUSTRINE ORIGIN

It has been shown that certain natural clays are able to destroy bacteria or significantly inhibit their growth. While the antibacterial mode of action is not totally understood, researchers have linked their toxicity with metal species that can produce deleterious effects to cells via production of reactive oxygen species. This study focuses on recently discovered antibacterial clay from the Colombian Amazon. The evidence shows that this clay contains transition metals not soluble in water but available when bacteria are in contact with the clay.

We determined the mineralogy using X-ray diffraction of bulk powders and oriented clays (<2µm). Chemical analyses of the clay were performed using X-ray Fluorescence and trace elements were determined by ICP-MS and ion chromatography on dissolved clays and their aqueous leachate (50mg clay/ml deionized water, equilibrated 24 hrs).

Standard microbiological methods were used to assess the viability of two model bacteria (Escherichia coli and Bacillus subtilis) after incubation with clay at 37˚C for 24 hrs. The antibacterial effectiveness of different clay size fractions (2.0–1.0μm, 0.5–1.0μm, 0.2–0.5μm and <0.2μm) was tested and the bulk clay was again tested after cation exchange. Scanning and transmission electron microscopy were used to observe clay-bacteria interactions and morphological changes of bacteria as a result of clay treatments.

XRD results show that the clay is composed of quartz (15%), kaolinite (47%), and mixed-layer illite-smectite (28%) and kaolinite-smectite (10%). Bioimaging (TEM, SEM) of the clay-bacteria contacts show attraction of the clay to bacterial cell walls and formation of electron dense areas inside the cells. All size fractions of the clay are bacteriostatic or bactericidal, but after cation exchange there is no inhibition of bacterial growth. ICP-MS analysis of the digested clay fraction shows concentrations of Cr, Cu, Zn above the Mimimum Inhibitory Concentration (MIC) of E.coli at neutral pH, in addition to high Fe and Ti compared to controls. We conclude that these elements are adsorbed to the clay surfaces, and impede bacterial growth by direct contact. Future testing will evaluate this antibacterial process by examining metabolic responses to the presence of the clay using gene expression studies.