SURFACE AND INTERFACE ANALYSIS Surf. Interface Anal. (In press) Published online in Wiley InterScience (www.interscience.wiley.com) DOI: 10.1002/sia.2570 Short Communication Probing the modifications of polystyrene surface properties after incubation with the Shewanella putrefaciens bacteria at two pH values (4, 10) by atomic force microscopy Fabien Gaboriaud, * Sidney Bailet, Etienne Dague and Fr ´ ed ´ eric Jorand Laboratory of Physical Chemistry and Microbiology for the Environment, Nancy-University, CNRS, 405 rue de Vandœuvre, F-54600 Villers-l ` es-Nancy, France Received 30 November 2006; Revised 16 February 2007; Accepted 9 March 2007 The local surface properties of polystyrene (PS) dishes incubated for 14 h with gram-negative cells (Shewanella putrefaciens) were investigated by atomic force microscopy (AFM) in aqueous solutions at two pH values (4 and 10). The AFM images and force curves revealed the presence of a polymeric layer adsorbed onto the PS surfaces in acid media. In contrast, no evolution was observed in the case of basic media. Copyright  2007 John Wiley & Sons, Ltd. KEYWORDS: AFM; Shewanella putrefaciens; biofilm; adhesion; polystyrene; force spectroscopy INTRODUCTION The colonization of mineral or biological substrates leading to the formation of a biofilm involves bacterial adhesion that is strongly mediated by bacteria – substrate interactions. A bet- ter understanding and control of the processes involved in bioadhesion would surely provide ways to diminish the con- siderable economic loss in biocorrosion of metals or ship hulls and also to anticipate nosocomial diseases due to the contam- ination of surgical tools. In the last decade, many attempts have been made to describe nonspecific bacteria–substrate interactions from the quantification of cell surface properties by using macroscopic measurements. 1 Traditionally, these data are compared with those derived from the colloid inter- action theory (called DLVO theory) in its simplest or extended version. 2,3 This theory was originally developed for hard, impermeable and nondeformable colloids. This statement demonstrates the limitations of these approaches to identify the molecular physicochemical determinants responsible for bacterial adhesion. 4 Among the new experimental techniques that probe soft surfaces at the nanometer scale, the atomic force microscope (AFM) offers unique advantages for quantifying bacterial surface properties in liquid media. 5–7 In particular, remarkable advances have been made in measuring and identifying (i) the mechanical characteristics of cells, 8,9 L Correspondence to: Fabien Gaboriaud, Laboratory of Physical Chemistry and Microbiology for the Environment, Nancy-University, CNRS, 405 rue de Vandœuvre, F-54600 Villers-l` es-Nancy, France. E-mail: gaboriaud@lcpme.cnrs-nancy.fr (ii) the electrical surface charge by using functionalized AFM tips, 10 (iii) the spatial distribution of individual adherents, 11 (iv) the cell–cell and cell–solid interactions by attaching a cell-coated bead to a cantilever. 12 While most of the studies focused on the quantification of he physical properties of bacterial cells, the analysis of the substrate modifications due to cell incubation has received little attention. 13 In fact, cells are expected to modify their local environment with extremely diverse components like proteins, carbohydrates or lipid heteropolymers. 14,15 Briefly, two types of structures can be considered: components anchored to the outer membrane/murein– protein layer and unanchored components. The former typically consists of lipopolysaccharides (LPS) for gram-negative bacteria. The latter type concerns a less well-defined group of polymers regrouped under the term extracellular polymeric substances (EPS). 16 This describes closely (e.g. capsular polymers) or loosely bound EPS, or soluble EPS not displaying any specific contact. This definition includes products of cellular lysis and hydrolysis of macromolecules. The involvement of EPS in biofilm dynamics and structure has been well documented. 16,17 For instance, EPS both strengthen adhesion to the substratum and lead to cell– cell attachment inside this 3D architecture. However, the bacterial production of such EPS may therefore change significantly the surface properties of the colonized substratum to favor bacterial adhesion. 13 In this context, the present work investigates the modi- fications of the surface properties of polystyrene (PS) after incubation with the S. putrefaciens bacteria at two pH values (4, 10). For this purpose, we used both the AFM imaging Copyright  2007 John Wiley & Sons, Ltd.