The influence of surface topography on bacterial adhesion has been investigated using a range of spatially organized microtopographic surface patterns generated on polydimethylsiloxane (PDMS) and three unrelated bacterial strains. The results presented indicate that bacterial cells actively choose their position to settle, differentiating upper and lower areas in all the surface patterns evaluated. Such selective adhesion depends on the cells' size and shape relative to the dimensions of the surface topographical features and surface hydrophobicity/hydrophilicity. Moreover, it was found that all the topographies investigated provoke a significant reduction in bacterial adhesion (30-45%) relative to the smooth control samples regardless of surface hydrophobicity/hydrophilicity. This remarkable finding constitutes a general phenomenon, occurring in both Gram-positive and Gram-negative cells with spherical or rod shape, dictated by only surface topography. Collectively, the results presented in this study demonstrate that spatially organized microtopographic surface patterns represent a promising approach to controlling/inhibiting bacterial adhesion and biofilm formation.

The cell envelope of gram-negative bacteria is responsible for many important biological functions: it plays a structural role, it accommodates the selective transfer of material across the cell wall, it undergoes changes made necessary by growth and division, and it transfers information about the environment into the cell. Thus, an accurate quantification of cell mechanical properties is required not only to understand physiological processes but also to help elucidate the relationship between cell surface structure and function. We have used a novel, atomic force microscopy (AFM)-based approach to probe the mechanical properties of single bacterial cells by applying a constant compressive force to the cell under fluid conditions while measuring the timedependent displacement (creep) of the AFM tip due to the viscoelastic properties of the cell. For these experiments, we chose a representative gram-negative bacterium, Pseudomonas aeruginosa PAO1, and we used regular V-shaped AFM cantilevers with pyramid-shaped and colloidal tips. We find that the cell response is well described by a three-element mechanical model which describes an effective cell spring constant, k 1 , and an effective time constant, , for the creep deformation. Adding glutaraldehyde, an agent that increases the covalent bonding of the cell surface, produced a significant increase in k 1 together with a significant decrease in . This work represents a new attempt toward the understanding of the nanomechanical properties of single bacteria while they are under fluid conditions, which could be of practical value for elucidating, for instance, the biomechanical effects of drugs (such as antibiotics) on pathogens.

Exopolymers are thought to influence bacterial adhesion to surfaces, but the time-dependent nature of molecular-scale interactions of biopolymers with a surface are poorly understood. In this study, the adhesion forces between two proteins and a polysaccharide [Bovine serum albumin (BSA), lysozyme, or dextran] and colloids (uncoated or BSA-coated carboxylated latex microspheres) were analyzed using colloid probe atomic force microscopy (AFM). Increasing the residence time of an uncoated or BSA-coated microsphere on a surface consistently increased the adhesion force measured during retraction of the colloid from the surface, demonstrating the important contribution of polymer rearrangement to increased adhesion force. Increasing the force applied on the colloid (loading force) also increased the adhesion force. For example, at a lower loading force of approximately 0.6 nN there was little adhesion (less than -0.47 nN) measured between a microsphere and the BSA surface for an exposure time up to 10 s. Increasing the loading force to 5.4 nN increased the adhesion force to -4.1 nN for an uncoated microsphere to a BSA surface and to as much as -7.5 nN for a BSA-coated microsphere to a BSA-coated glass surface for a residence time of 10 s. Adhesion forces between colloids and biopolymer surfaces decreased inversely with pH over a pH range of 4.5-10.6, suggesting that hydrogen bonding and a reduction of electrostatic repulsion were dominant mechanisms of adhesion in lower pH solutions. Larger adhesion forces were observed at low (1 mM) versus high ionic strength (100 mM), consistent with previous AFM findings. These results show the importance of polymers for colloid adhesion to surfaces by demonstrating that adhesion forces increase with applied force and detention time, and that changes in the adhesion forces reflect changes in solution chemistry.

We used a novel atomic force microscopy (AFM)-based technique to compare the local viscoelastic properties of individual gram-negative (Escherichia coli) and gram-positive (Bacillus subtilis) bacterial cells. We found that the viscoelastic properties of the bacterial cells are well described by a three-component mechanical model that combines an instantaneous elastic response and a delayed elastic response. These experiments have allowed us to investigate the relationship between the viscoelastic properties and the structure and composition of the cell envelope. In addition, this is the first report in which the mechanical role of Lpp, the major peptidoglycanassociated lipoprotein and one of the most abundant outer membrane proteins in E. coli cells, has been quantified. We expect that our findings will be helpful in increasing the understanding of the structureproperty relationships of bacterial cell envelopes.The surface layers that isolate the interior of a bacterial cell from its external environment play a crucial mechanical role in the survival of the cell. They must be strong enough to maintain the cellular shape and resist turgor pressure yet, at the same time, be flexible enough to allow cell growth and division. Their elastic response is evident from their ability to recover from transient deformations, such as those induced by the incorporation of additional surface components (e.g., proteins) in response to changes in environmental conditions and the passage of small molecules across the cell boundary. It is therefore clear that understanding many aspects of cell physiology requires knowledge of the mechanical properties of cells.The mechanical properties of the cell originate from the structural organization of the constituent lipids, sugar polymers, and proteins. Lipid molecules are brought together by their hydrophobic domains to form bilayers (membranes) that also incorporate different types of proteins. Polymeric strands of sugar molecules are typically cross-linked by flexible peptide molecules to form the peptidoglycan layer (27). Sometimes, an additional layer of proteins (S layer) is found on the outermost surface of the cell (7,8,40). Depending on the structural organization of the peptidoglycan and lipid bilayers, bacteria can generally be divided into gram-positive and gram-negative bacteria. In gram-positive cells, there is a relatively thick (20-to 35-nm) peptidoglycan layer that, together with the plasma membrane, sandwiches a viscous compartment called the periplasm (31, 32), whereas the envelope of gram-negative cells is made up of two lipid bilayers, the inner and outer membranes, separated by the periplasm, which contains a thin (3-to 8-nm) peptidoglycan layer (5, 33). In gram-negative bacteria, lipoproteins are associated with both the peptidoglycan layer and either the inner or outer membrane. Here, the "lipo" substituent is inserted into the hydrophobic domain of the membrane and the "protein" portion is linked to the peptidoglycan layer by either covalent or electrostatic bonds...