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Lec. 3 Microbial deposits

الكلية كلية طب الاسنان     القسم التقويم والاطفال وطب الاسنان الوقائي     المرحلة 5
أستاذ المادة حسن فليح فرحان السلطاني       6/7/2011 12:48:43 PM

د. حسن الوطيفي                                   Prevention                                                                 Lec. 3

Microbial deposits
Bacterial colonization of the mouth
One milliliter of whole saliva may contain more than million organisms representing more than 250 different bacterial species. These microorganisms constitute a very complex microbiota which itself dose not result in disease as they exist in equilibrium and in balance with the host.
Microorganism initially colonizes the mouth after birth being naturally acquired from the mother, thereafter; bacteria come to reside in the mouth from sources such as the atmosphere, food and human contact.
All surfaces within the oral cavity will constantly be subject to microbial colonization, the pattern of which will be partly dependent on the surface properties. As the sequamous epithelium of the mucous membranes of the oral cavity is constantly being renewed as a result of turnover in the epithelium, the surface epithelial cells will be constantly being desquamated, together with the colonizing microorganisms and swallowed. On tooth surfaces no surface renewal takes place and therefore microorganisms are able colonize and grow on these surfaces unless removed mechanically or chemically. Colonization may be somewhat different on tooth surface that are exposed to chewing forces and abrasion from foods on lips and cheeks than in areas which are not easily accessible, such as proximal site, tooth surfaces along the gingival margin, and in irregularities of tooth surface, such as occlusal fissures. These areas are often designated stagnation areas.
The development of dental plaque occurs in two phases: acquired pellicle formation and dental plaque formation

Acquired pellicle
Microorganism do not deposit or adhere directly an apatite crystal surface.
Within seconds after saliva first contact the external tooth surface, a coating of salivary materials called the acquired pellicle begins to develop on the tooth therefore, saliva is very seldom in direct contact with the tooth surface, the saliva compartment is separated from the tooth by a thin layer of material called the acquired pellicle. This layer of salivary protein adheres strongly to the enamel that is not removed during tooth brushing.
The pellicle is defined as a cellular layer of adsorbed salivary proteins and other macromolecules on the dental mineral surface that is continually deposited on those surfaces of the tooth exposed to the oral cavity.
The formation rate varies between individuals probably due to differences in salivary composition. If the pellicle is displaced by prophylaxis, it begins to reform immediately.
The pellicle layer even if thin, has an important role in protecting the enamel against mechanical and chemical insults (abrasion and attrition), moreover, this film can take up stain and in other way influence the coating of the teeth.
Pellicle in other wards can be defined as protective diffusion barrier formed on enamel from salivary protein. Because of its perm selective nature, restricting transport of ions in and out of the dental hard tissues, through this matrix network surface, fluids and small sized molecules can slowly diffuse throughout the enamel; therefore, the pellicle may play an important modifying role in caries process. Experiments has shown that the pellicle delay the initiation of caries and the dissolution of the enamel when teeth are placed in low pH soft drink.
There is a competition for the binding sites on the pellicle, not only by receptors on the bacteria, but also from host protein, such as immunoglobulins; ie antibodies, proteins of the complement system, and the enzyme. These host protein originate from the saliva and the gingival (crevicular fluid). Once one of the competing entities occupies a pellicle site, occupancy by another is interdicted, not only dose competition arise for occupancy of binding sites but an antagonistic relationship often exist between different types of bacteria occupying the binding sites. For example, it has been shown that some streptococci synthesize and release bacteriocins, which can inhibit some strains of Actinomyces.
The bacterial colonization of the acquired pellicle can be beneficial for the bacteria because the pellicle components can serve as nutrients.

Plaque formation
Dental plaque is not a static entity but should be regarded as constantly subject to force for change - reforming and changing in composition and properties under the influence of environmental stresses.
The supragingival plaque Formation envisage a succession of stages (succession means microbial population shift), each of them involving a different range of bacterial species, but all stages being characterized by a set specific molecular interaction, This microbial succession may be summarized as follows:
Initial microbial colonization
In the initial stage (pioneer) species of bacteria colonize the tooth surface by binding to component of the salivary pellicle, among these initial colonizers are believed to constitute a highly selective part of the oral microflora, mainly Streptococcus sanguis, streptococcus oralis and Streptococcus mitis. In addition, the initial microflora comprises minor proportion of Actinomyces spp. And gram"rie;a;a.tive bacteria, e.g. Haemophilus spp. It must be noted that the mutans streptcicocci are not considered to be amongst these pioneer species and only represent a very small proportion of the total number of bacteria present.
The selective manner in which the bacteria attach to the tooth surface is thought to reflect the fact bacteria on their surfaces contain a recognition system, which enables component on the bacterial surface (adhesions) to bind to complementary molecules (receptors) in the pellicle.
Another molecular mechanism of bacterial adhesion may be the phenomenon termed calcium bridging, which links negatively charged bacterial cell surfaces to the negatively charged acquired pellicle via interposed, positively charged divalent calcium ions from me saliva.
For many years, it was believed that mutans streptococci constituted a significant part of the initial microflora because of their ability to elaborate sticky extracellular polysaccharide from sucrose. However recent studies have shown that mutans streptocopcci comprise only 2% or less of the initial streptococcal microflora, irrespective of an individual s exposure to sucrose. The low recovery of mutans streptococci in initial dental plaque in vivo is most probably a result of the relatively low concentration of these species in saliva A positive correlation has been demonstrated between the concentration of specific microorganism available in saliva for adsorption and the actual number of adsorbed microbial cells.
Also Streptococcus mutans is much less efficient than S. sanguis in adhering to the tooth surface, approximately 104-105 cells of the S. mutans per ml of saliva has to be present before one cell can be recovered from a cleaned tooth surface, while the equivalent concentration for S. sanguis is about 103 cells per ml.
Plaque then develops by both:
• Growth of the pioneer species to form microcolonies.
• The further accretion of more bacterial cells of the same or other species.
• This accretion can be direct bacterium-bacterium binding in coaggregation pairs due to specific receptor ligand binding or May be mediated by salivary macromolecules that cross –link because they are bound more than one bacterium.
Along with these process, plaque build-up is facilitated by the production of bacterial extracellular polymers which serve to entrap the cells, the polymer of a particular interest are those synthesized from sucrose by various species of streptococci.

Microbial succession
The principal of microbial succession is that pioneer bacteria create an environment which is either more attractive for secondary invaders or is increasingly unfavorable to themselves because of lack of nutrients, accumulation of inhibitory metabolic products etc. In this way, resident microbial community is gradually replaced by other species more suited to the modified habitat. However, it is remarkable that S. oralis, a typical initial colonizer, is hardly ever present in mature supra gingival Plaque.
As the bacterial deposits become thicker, lowering of the oxygen concentration is one of the factors responsible for the microbial succession. Thus, in developing coronal plaque, a progressive shift is observed from mainly aerobic and facultative anaerobic species in the early stages, to a situation m which facultative anaerobic and anaerobic organisms predominate after 9 days.
However, even if the species composition of dental plaque may remain reasonably constant over time, this stability dose not result from a static set of circumstances, but rather reflects a highly dynamic situation in which dying bacteria are continuously replaced by new ones.
Mature plaque with its (climax community) of microbial species is dependent upon the maintenance of a fine balance between many species present ((homeostasis) that tend to eject invading species not previously present, with metabolic interactions and food chains taking an increased importance over adhesive interaction.                               .
The relative proportion of different species (the pathological potential of the plaque) can be altered drastically by shift in the environmental condition (particularly dietary sugar).
Along the gingival margin, the dental plaque adjacent to the gingival crevice will constantly be exposed to the gingival fluid. Because of the presence of the microorganisms, this tissue fluid (in addition to various nonspecific and specific antimicrobial components) contains numerous polymorph nuclear leukocytes, which actively combat the microorganisms. Clinically this result in a very narrow zone close to the opening of the gingival crevice, which is kept free of dental plaque, this is a good example of the protective nature of the gingival fluid. As long as the balance exist in this area any attempt by microorganisms to colonize within the crevice and thus grow down between junctional epithelium and the tooth surface creating the gingival pocket may be prevented. Thus the presence of the dental plaque along the gingival margin results in a protective reaction in the attached marginal gingiva which clinically presents itself as gingivitis.
With time, more bacteria migrate subgingivally and the process continues more aggressively, the critical locus of activity is the subgingival place, the bacteria residing in the pocket and the host cells that defend it determine the clinical outcome.
The microbiota of the gingival crevice exists in two distinct locations and m two distinct form.
1- The first of these the adherent plaque deposits at or below the height of the gingiva, differ quantitatively and qualitatively in composition from plaque on the coronal surfaces.
2- The second type of plaque seen in the gingival crevice is what ha s been called free-floating plaque, this plaque is composed of an unattached mass of motile organisms residing between the outer portion of the adherent plaque and the crevicular epithelium.
The pathogencity of each of the plaque can vary independently of the others. The diseased periodontal pocket harbors both attached subgingival plaque biofilms and non-attaching motile subgingival rnicroflora (spirochetes, vibrios,and straight rods with flagella).

Fissure plaque
Studies of the microflora in occlusal pits and fissure have always been hampered by inaccessibility to sampling. This problem can partly be overcome by example: the use of in-situ models, in which blocks of natural human fissures prepared from unerupted third molars are mounted into large amalgam fillings of subjects.
The salivary concentration of S. mutans was crucial for colonization of artificial fissures. Thus reducing the salivary concentration of S. mutans (< 103 CPU/ml) around the time of insertion, this organism did not colonize the fissures even if the salivary concentration was allowed to increase during subsequent experimental period. Once established, however experimental reduction m saliva did not influence the proportional distribution of S. mutsms in the fissures, these finding indicate that the initial inoculums may be the main determinant for fissure colonization.
When comparing the microflora in fissures after 7days and 200-270 days its striking that the qualitative changes caused by microbial succession in smooth surface plaque are not found in the fissures
Streptococcal adhesions
Mutans streptococci are able to attach to the tooth surface by either of two mechanisms:
1. Sucrose - independent adsorption, in which the bacteria attach lo the acquired pellicle through specific extracellular proteins (adhesions) that bind to salivary glycoprotein on tooth surface.
2. Sucrose dependent mechanisms, in which bacteria require the presence of sucrose to produce sticky extra cellular polysaccharides, or glucan, which allow attachment and accumulation.
While many fermentable carbohydrates may be utilized by plaque bacteria to generate the acids which attack enamel substance, sucrose is recognized, as being particularly important in caries process because not only can it be fermented, but it also serves as a substrate for extracellular enzymes of plaque bacteria which synthesize sucrose - derived polymers.
These polymers are of central important:
• A dhesive interaction in plaque where they mediate attachment of bacteria to the tooth surface and to other bacteria.
• They stabilize the plaque biofilm.
• Serve as energy stores aiding the survival of plaque bacteria.
• Volume of dental plaque is increased through its content of glucans, which may provide a barrier to diffusion of acids away from the tooth surface.
One of the key enzymes in the conversion of the glucose moiety of sucrose to glucan is glucosyltransferase enzyme (GTF) making water soluble and insoluble glucans, as well as the glucan-binding proteins (GBP).
A glucan which is water soluble is referred to as dextran and in contrast mutan is water -insoluble glucas. Both types of glucans are believed to be important in dental plaque, with dextran mediating bacterial aggregation and serving as a storage polymer, while mutan has been shown to be the major contributors to adherence.
Mutans streptococci, S. salivarius and Actinomyces viscosus may also form high molecular weight and soluble extracellular fructose polymers (fructans) in the presence of sucrose. Fructan can act as an energy source for any bacteria having the enzyme, levanase to use as an energy source.

Metabolism of plaque
For metabolism to occur, a source of energy is required, for streptococcus mutans and many other acid forming organisms this energy source can be asucrose. Almost immediately following exposure of these organisms to sucrose, they produce:
1" acids 2- an intracellular polysaccharide which provide a reserve source of energy for each bacterium. 3- extracellular polysaccharide glucan (e.g. dextran) and fructan (levan).
The streptococci are of a central importance in the metabolism of plaque because they are well equipped to survive in the fluctuating conditions.
Within plaque, it has been estimated that the glucose concentration may vary over a 10000-fold range; the pH may shift from 7.5-4,0 and while oxygen is freely available at the plaque surface, condition are entirely anaerobic close to the tooth surface.
The streptococcal are well suited to flourishing under varying condition of oxygen availability (facultative anaerobic). With regard to acid, S. mutans is notable in combining the properties of being extremely acidogenic and also aciduric. As consequence when there is a good supply of carbohydrate, S. mutans will produce a large amount of acids mainly lactic acids that will lower the plaque pH, continues to metabolize under the low pH condition while other competing species are disadvantage (different plaque species differ in the range of pH that they can tolerate), with the net result that the relative proportion of S. mutans in plaque population increase.


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