Molecular determinants of bacterial adhesion monitored by atomic force microscopy

Abstract

Bacterial adhesion and the subsequent formation of biofilm are major concerns in biotechnology and medicine. The initial step in bacterial adhesion is the interaction of cells with a surface, a process governed by long-range forces, primarily van der Waals and electrostatic interactions. The precise manner in which the force of interaction is affected by cell surface components and by the physiochemical properties of materials is not well understood. Here, we show that atomic force microscopy can be used to analyze the initial events in bacterial adhesion with unprecedented resolution. Interactions between the cantilever tip and conf luent monolayers of isogenic strains of Escherichia coli mutants exhibiting subtle differences in cell surface composition were measured. It was shown that the adhesion force is affected by the length of core lipopolysaccharide molecules on the E. coli cell surface and by the production of the capsular polysaccharide, colanic acid. Furthermore, by modifying the atomic force microscope tip we developed a method for determining whether bacteria are attracted or repelled by virtually any biomaterial of interest. This information will be critical for the design of materials that are resistant to bacterial adhesion.Bacterial adhesion onto inanimate surfaces is a critical issue in processes ranging from the biofouling of industrial equipment to dental decay to infections of biomaterials for medical use. Bacterial infections associated with the formation of biofilms refractile to antibiotic therapy is one of the main reasons for the failure of devices such as catheters, vascular grafts, joint prostheses, and heart valves (1-3). The first step in bacterial adhesion is the immediate attachment of bacteria onto a substratum which is a reversible, nonspecific process (3-5). This initial interaction between bacteria and artificial surfaces is a key determinant in biofilm formation. If the approach of bacteria to a surface is unfavorable, cells must overcome an energy barrier to establish direct contact with the surface. Only when bacteria are in close proximity to the surface do shortrange interactions become significant. Thereafter, proteinligand-binding events mediated by a plethora of microbial adhesins and in some cases the production of extracellular polymers render the binding process practically irreversible (6).Initial bacterial attraction or repulsion to a particular surface can be described in terms of colloidal interactions. Consequently, the force of interaction depends on physiochemical parameters such as surface-free energy and charge density (7-11). The propensity of bacteria to adhere onto surfaces has been estimated by counting the number of bacteria that remain attached to surfaces following incubation for a specified length of time (5, 12). This approach is qualitative, time consuming, and has low sensitivity. Moreover, the resulting number of adherent bacteria is determined by multiple factors in addition to long-range attractive/repulsive interactions. A direct and ...

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Force measurements with the atomic force microscope: Technique, interpretation and applications

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