## 3 Biological Mechanisms in Bacterial Adhesion as Potential Subjects of Label-Free Monitoring
### Role of Bacteria's Glycocalyx in Their Adhesion  
Bacterial glycocalyx is a highly hydrated polymeric layer that extends outwards from bacterial surface; 99% of its volume is water [19]. Branching sugar molecules or a mass of tangled fibers of substituted polysaccharides form the glycocalyx that surrounds an individual cell or colony [18; 317]. Bacteria must expend energy to generate and maintain a glycocalyx. Thus, in the environment of a pure and unthreatened culture, it is an unnecessary material conferring no selective advantage; cells that create these coatings are usually eliminated from pure cultures by uncoated ones that can allocate more energy to proliferation. In natural, competitive circumstances, selection prefer cells that are protected and able to adhere to a proper surface by their glycocalyx. Thus, having a glycocalyx is required for most bacteria to succeed in their different natural environments [18]. It can be visualized by ruthenium red staining, which is taken up by any polysaccharide present in the glycocalyx fibers [318]. The constituents of the glycocalyx may be either homopolymers or complex heteropolymers of monosaccharides (mainly neutral hexoses, 6-deoxyhexoses, polyols, uronic acids, and amino sugars), with substituents formate, pyruvate, phosphate, and succinate. Glyocalyxes are divided into two types based on how tightly they are attached to the bacterial cell. Loosely attached, diffuse fibers form the amorphous slime layer, while in the capsule, the bacteria is tightly covered with a gelatinous, distinct fibrous matrix, in which the forming polysaccharide is covalently attached [319]. Physiological factors regulate the synthesis of the glycocalyxes [247; 320]. Mostly, capsule formation is favored in case of high C:N ratio of the environment, and ion concentrations have been shown to affect capsule production as well [19; 321]. When the glycocalyx is lost (in laboratory circumstances), other polymers are presented (for example, the LPS of the Gram-negative bacterial cell wall) [19]. However, during infections, bacteria may also change their expression of antigenic determinants, including polysaccharides, in a process called phase variation, which may well help them evade the host immune response and thus affect their virulence [322]. The highly variable nature of bacterial polysaccharide expression is also reflected in the relatively high number of serotypes identified in certain bacteria [319; 323]. Importantly, some bacteria even express polysaccharides mimicking host molecules to avoid recognition by the immune system [324]. Bacterial adhesion to an inert surface must overcome the electrostatic repulsive forces [325]. Long hydrophilic extensions of the bacterial cell can bridge the space between surface and bacterium and allow H-bonding or ion pair formation (attractive forces) to start attachment [19; 326]. After its adhesion to the surface, the cell surrounds itself with further glycocalyx fibers and multiplies itself within this matrix to form a microcolony [18; 19]. Other morphologically distinct bacteria then adhere by their glycocalyx to create a layered bacterial consortium, forming a biofilm [16; 19; 317; 327; 328]. The negatively charged glycocalyx can form a polar bond with polysaccharides via divalent positive ions in the medium, and lectins can also form a bridge between them. The nature of the host/eukaryote glycocalyx changes as the cell ages. Furthermore, the glycocalyx alters in the cells that have been infected by a virus, which may explain the enhancement in susceptibility to bacterial illness that is often observed after viral infection [18]; viral infection changes the tissue cell surface and allows the colonization by opportunistic bacteria [19]. Adhesion to a certain tissue provides bacteria a constantly renewed supply of organic nutrients with physical conditions favoring growth. Because of adhesion, infecting bacteria can be resistant to removal, especially in a system that is sterile among normal conditions.