The Cariogenic Challenge
Dental plaque is an example of a biofilm, a film of micro-organisms adhering to a solid surface. Biofilms exist in a wide variety of types adapted to different habitats. Life in a biofilm requires physiological adaptations on the part of the constituent microorganisms and also provides several advantages, for instance, protection against antimicrobial agents.35 The structure of biofilms varies widely, but in the case of dental plaque the constituent bacteria are closely packed together, occupying ca. 75% of the volume.36 The remaining volume is made up of a matrix comprising proteins, carbohydrate polymers, and other substances (Fig. 2.10). Many matrix components, such as extracellular polysaccharides, are largely of bacterial origin, but others, including several proteins, originate from saliva and gingival crevicular fluid.
Because of the dense structure of dental plaque, movement of nutrients and metabolic end products between the oral cavity and plaque, and within plaque, is mediated by diffusion, which is a relatively slow process (Fig. 2.11a). One consequence is that availability of nutrients or antibacterial substances will not be uniform but will vary with depth. For instance, when a nutrient is ingested, a gradient will be set up within the plaque (Fig. 2.11b), with the concentration falling toward the interior, and bacterial metabolism will steepen this gradient because utilization near the outer surface will make less nutrient available for inward diffusion (Fig. 2.12). This phenomenon has important effects on plaque ecology. For instance, most plaque bacteria are anaerobic (surviving only in the absence of oxygen), such as Veillonella or facultative (preferring to live in absence of oxygen), such as streptococci and Actinomyces, probably because oxygen is consumed by aerobic bacteria such as Neisseria at the plaque surface, and none reaches the inner plaque.3 A similar process probably limits exposure of plaque bacteria to antimicrobial agents, because such agents will be immobilized by strong interaction with bacteria in the outer plaque, and the concentration reaching the inner plaque may be too low to be effective. Metabolic end products clear slowly from plaque because their movement is similarly regulated by diffusion. These phenomena are central to the process of dental caries because they control the different phases of the cariogenic challenge.
Fig. 2.10 Diagram to represent some aspects of plaque structure. Tooth surfaces exposed to saliva are covered by an acquired pellicle consisting of adsorbed salivary proteins. Initial bacterial colonists attach to the pellicle, and the plaque increases in bulk and complexity by attachment of further bacteria to the initial colonists. Attachment is mediated by specific receptors on bacterial surfaces (indicated by geometrical lock-and-key shapes). However, plaque cohesion is further enhanced by nonspecific interactions (dotted lines), e.g., calcium bridging, and by interactions between polymers forming the plaque matrix: proteins and extracellular polysaccharides (EPS) such as glucans.
Fig. 2.11a, b Diffusion.
a A single molecule (small red circle) in solution makes frequent small ‘jumps’ (single arrows). Even though each jump is in a random direction, a series of jumps results in net movement of the molecule (double arrow, d = net distance moved).
b Since more molecules move from regions of high concentration into regions of low concentration than vice versa, there is net transfer of molecules down gradients of concentration. This is illustrated for molecules diffusing from saliva (left) into plaque, via the plaque fluid.
Fig. 2.12 Diffusion-with-reaction in plaque. Molecules of a nutrient such as sugar or oxygen (small red circles) diffusing into the surface of the plaque (left) are utilized immediately by superficial bacteria and this leaves less to diffuse deeper into the plaque. This results in a much steeper fall in concentration than in the case of simple diffusion (cf. Fig. 2.11). If metabolism is rapid, little or no nutrient will reach the deepest parts of the plaque. In plaque containing little extracellular polysaccharide (EPS) (above), the bacteria are closely packed and quickly utilize sugar molecules diffusing in from the saliva, allowing little to reach the interior. If EPS is abundant (below) the bacteria are more widely spaced, more sugar can diffuse into the interior and bacteria near the tooth surface can produce acid, which causes a greater pH drop in the environment of the tooth mineral.
The interstitial fluid bathing the matrix, referred to as plaque fluid, is the component of plaque in direct contact with the tooth surface and is therefore the medium for the flux of H+ ions and mineral ions during the caries process. Analysis of plaque fluid isolated by centrifugation shows that its electrolyte composition differs markedly from that of saliva37 (Table 2.2), mainly because exchange of ions between the two fluids is diffusion-dependent and slow. Thus, the elevated potassium concentration is due to slow clearance of K+ ions released by bacterial lysis. In plaque that has not recently been exposed to nutrients, the predominant organic acid is acetic acid (Table 2.2), which is an end product of metabolic pathways that derive the maximal ATP from the low amounts of available carbohydrate3.
A cariogenic challenge is initiated by exposure to fermentable carbohydrate, which provokes a characteristic pattern of change in plaque pH, known as the Stephan curve3,4 (Fig. 2.13), which can be recorded by micro-electrodes inserted into the plaque. The duration of a Stephan curve varies, but is typically 30–60 minutes. The curve can be divided into two phases: an initial rapid pH fall from the resting value (approximately pH7), followed by a slower recovery of pH. These phases reflect the underlying pattern of bacterial metabolism. In the initial phase, the high intraoral sugar concentration typical of confectionery or sweetened drinks drives diffusion of sugar into the plaque, where it is rapidly metabolized to produce energy. The main metabolic pathway is conversion of sugar by glycolysis to pyruvic acid and then directly to lactic acid (see Chapter 11), which reduces the pH within the plaque fluid3 (Table 2.2). The rate of pH fall is slowed by combination of H+ ions with plaque buffers, made up mostly of macromolecules associated with the bacterial cell walls. Simple sugars are cleared from the mouth by saliva within 1–2 minutes but the pH fall in plaque lasts for somewhat longer because the bacteria continue to metabolize the sugar that diffused inward initially. Eventually the sugar is used up and acid production ceases, so the initial phase of the Stephan curve comes to an end. The minimum local pH during a Stephan curve is about 4.0, which represents the lowest value at which even the most aciduric bacteria can produce acid, but average values will be higher than this.
Although plaque pH is lower than under “resting” conditions for the whole duration of a Stephan curve, only part of it represents the cariogenic