Overview

Bacterial attachment to surfaces is a common occurrence that can be a problem for humans, for example in tooth decay and infection produced using biomaterial implants, as well as in various natural and engineered systems. Biogeochemical occurrences include: ground and surface water contamination by pathogenic bacteria when these are used as sources for drinking water delivery system; biofouling of membranes and ship hulls; rapid adsorption (via filtration processes) of pollutant-degrading bacteria injected into the ground for soil remediation via bioaugmentation (the addition to soils of laboratory-grown cultures to chemically contaminated soils); bacteria-promoted interactions in mineral cycling (Characklis and Marshall 1990). In industrial processes, cell adhesion typically reduces the efficiency of the devices or processes. In natural systems, a lack of bacterial adhesion can facilitate the spread of pathogenic bacteria in groundwater aquifers. Conversely, during bioaugmentation, rapid cell adhesion can prevent the dissemination of pollutant-degrading bacteria to the location of pollutants in the soil. In general, it is much easier to promote adhesion than it is to reduce adhesion. To successfully transport of bacteria in aquifers, for example, conditions must be developed that produce cells capable of 100 to 1000 unsuccessful collisions in order for bacteria to be transported only 1 m in the groundwater. Unfortunately, extensive results indicate laboratory grown cells typically undergo less than 10 to 50 collisions before attachment (Johnson et al. 1996).

Bacterial adhesion and transport is a special case of a more general area of study in soil-water systems of colloid transport. Colloid mobilization and transport can affect subsurface hydrodynamics and water quality. Changes in water ionic strength can promote or inhibit colloid transport via adsorption and desporption processes. For example, excessive clay colloid desorption and mobilization, and mineral precipitation (via metal oxides), increase the abundance of colloids and can lead to aquifer plugging (Wiesner et al. 1996). Sorption of chemical pollutants onto mobile colloids increase the spread of pollutants otherwise highly retarded dissolved phases (Roy and Dzombak 1997).

Particle transport is primarily controlled by electrostatic forces, and secondarily influenced by hydrophobic interactions (van Loosdrecht et al. 1987, 1990; Camesano and Logan 1998a). Bacteria negatively charged at neutral pH with zero point charges at pHs of 2-4 and negative electrophoretic mobilities (Richmond and Fisher 1973, Glynn et al. 1998). Methods used to date to study bacterial adhesion primarily have been focused on bulk cell properties such as electrophoretic mobility for electrostatic forces(Glynn et al. 1998), and cell partitioning into hydrophobic fluids or contact angle measurements for hydrophobic forces (Doyle and Rosenberg 1990). The extent that adhesion is modified by chemicals can be probed in concert with modification of solution properties such as pH and ionic strength (Gross and Logan 1995; Johnson et al. 1996). Although substantial improvement in our understanding of bacterial adhesion has resulted from these macroscale measurements, these have provided relatively little understanding at the molecular level of forces governing cell attachment to soil particles or engineered structures although it is clear that such information is critical to understanding the surface forces that control cell adhesion to surfaces (Simoni et al. 1998; Jucker et al. 1998; Camesano and Logan 1998b). It is therefore a long term goal of this research to replace relatively crude macroscopic measurements of cell affinities for surfaces, for example due to cell hydrophobicity, with methods that directly measure cell-surface attractive forces such as AFM.

Atomic force microscopy (AFM) provides a unique avenue to study colloid and bacterial attachment at atomic, nanoscale and microscale levels (Bremer et al. 1992; Gad and Ikai 1995; Milling et al. 1996; Bowen et al. 1998). Fundamental research areas include: the role of NOM in cell adhesion; the orientation and topography of adsorbed cells; changes in cell morphology resulting from surface-modifying chemical treatments that promote cell detachment; detailed investigation of the electrostatic and hydrophobic forces affecting attachment; development of electrostatic models of cell adhesion. The abundance and composition of specific molecules responsible for initial bacterial adhesion to a surface can be investigated by techniques such as surface plasmon resonance spectroscopy (SPRS), Raman scattering and nuclear magnetic resonance (NMR) (Yeung et al. 1995; Jorden and Corn 1997). The role of NOM in colloid adhesion and detachment is critical because all water-solid interfaces are coated to some extent with NOM. Using SPRS the extent and mechanism of NOM displacement (or participation) by cell associated adhesion molecules can be measured. When such measurements are coupled with AFM, it is possible to measure changes in localized and overall attractive forces that have occurred.

The topography and charge of surfaces and particles is made possible using two different AFM techniques: direct contact measurements, and tapping measurements. In direct contact mode, the tip is brought to a surface and then pulled across the surface. The changes in the tip height are measured using a laser system. In tapping schemes, surface-tip contact is minimized: the tip is oscillated at its resonant frequency and by monitoring the tip deflection it is possible to map a surface. In both cases, the deflection of the tip can be used to calculate the attractive (or repulsive) forces between the tip and the material being scanned. At Penn State, we are developing methods to translate tip-surface interactions into surface charge measurements using electrostatic (DLVO) theory (Camesano and Logan 1998b).

In order to increase the potential range of the AFM for probing surface forces, it is possible to bond different chemicals to the tips (typically silicon nitride) and even to attach to the tips different types of colloids such as glass beads, single yeast cells and clumps of bacteria (Gad and Ikai 1995; Bowen et al. 1998). Bonding chemicals to the tips permit modification of not only electrostatic, but also hydrophobic forces. By attaching particles to tips, the interactions of chemically coated materials and quartz or quartz-NOM modified surfaces is possible An example of the surface of an individual bacterium taken using an AFM in tapping mode is shown in Figure 1.

Using the AFM to determine bacterial forces of attraction to surfaces in the presence of surface and solution-modifying chemicals should make it possible to design more efficient systems for facilitating bacterial transport during bioaugmentation, but there are many other uses of an AFM. Examples include: monitoring biologically mediated corrosion of metals and mineral surfaces (Grantham and Dove 1996); probing changes in surface potentials and topography resulting from chemical adsorption and reaction (Boronina et al. 1998); mobility and charge of surfactants in soil-aqueous systems; and measuring the topography of membranes used in water treatment (Zhu and Elimelich 1997). For example, using AFM, the improved resistance of a cellulose acetate membrane to fouling can be seen to be result from the smooth surface of the cellulose membrane versus another composite membrane which more easily fouls (Figure 2). The incorporation of an AFM teaching module into training for the students associated with the IGERT activities will provide a wonderful opportunity for students in the BRIE area to interact with students and professors in other areas of engineering and science.