Kinetic analysis of Streptococcus sanguis adhesion to artificial pellicle.

Studies of equilibria between Streptococcus sanguis and artificial pellicle have suggested that there are multiple binding sites for the organism. In the present study, adhesion of S. sanguis to saliva-coated hydroxylapatite was examined by means of kinetic methods. Cell-pellicle complex formation was measured from initiation of binding to equilibrium. Rate constants were calculated for forward reactions (adsorption) and reverse reactions (desorption). Initial binding obeyed reversible, first-order kinetics, whereas desorption of bound cells followed biphasic kinetics. Initial desorption proceeded approximately ten times faster than the slower second rate. The results are consistent with the mechanism C + P reversible CP* in equilibrium with CP in which CP* represents the reversible equilibrium that shifts at a discrete rate to the high-affinity CP state. Thus, the biphasic binding behavior that has been previously deduced from equilibrium studies may be attributed to a time-dependent shift from close apposition to pellicle, stabilized by low-specificity forces, to a higher-affinity binding.

Bonding of polymers to enamel: influence of deposits formed during etching, etching time and period of water immersion.

A technique for measuring the shear bond strength of composites to etched enamel was developed. The importance of thorough washing of the enamel surface to remove deposits formed during etching was demonstrated by measurements of bond strength and by scanning electron microscopy. One of three orthodontic adhesive systems showed a highly significant increase in bond strength after immersion in water for three months. Etching times as short as five seconds gave a bond strength not significantly different from that obtained after a one-minute etch.

Enamel solubility rate measurements in vivo on naval recruits.

Enamel solubility rates were determined in vivo to depths of 1 to 2 mum in the teeth from two samples of naval recruits that had been grouped, respectively, according to DMFT scores and visual indexes of oral hygiene status. No positive correlation between enamel solubility rates and DMFT scores were observed; rather there was a tendency for caries-resistant participants to have a highest apparent enamel solubility rates, even though mean differences among the DMFT groups were not generally significant. Intra-subject measurements on 12 teeth per participant for eight of the recruits showed, on the average, only about one half the variability founl incisors for more than 70 of the recruits. The least variability was found for measurements on paired maxillary central incisors. It was concluded that factors such as tooth morphology and, in particular, tooth-surface films could considerably affect enamel solubility rates measured in vivo.

Modification by salivary pellicles of in vitro enamel remineralization.

The effectiveness of inorganic calcifying solutions to remineralize enamel with subsurface demineralization decreased with saliva pre-treatments conducive to the formation of enamel pellicles. Greater reductions in remineralization rates occurred with longer pellicle formation times. It is suggested that enamel pellicles may act to control surface deposition and favor subsurface precipitation.

Effect of salivary pellicle formation time on in vitro attachment and demineralization by Streptococcus mutans.

Enamel subsurface demineralization induced by Streptococcus mutans was significantly reduced by seven-day saliva pre-treatments conducive to the formation of enamel pellicles. A two-hour saliva pre-treatment was ineffective. Results suggest that the protection provided by long-term pellicles may relate to changes in ionic transport rates rather than cell attachment.

In vitro enamel demineralization by Streptococcus mutans in the presence of salivary pellicles.

Salivary pellicles developed on extracted teeth favorably affected the degree and nature of enamel demineralization when the teeth were incubated in vitro with either of two pure strains of cariogenic Streptococcus mutans. The mechanism responsible for this protection may relate to the permselective properties of these salivary pellicles.

Demineralization and remineralization of dental enamel.

Experimentation in vitro using organic acid buffers as demineralizing media shows that caries-like lesions can be obtained which are very similar in morphology and developmental stages to early lesions formed naturally under oral conditions. The use of these chemical systems and of mechanistic models advanced to explain the unique histological features of incipient caries have yielded a good understanding of the processes involved in caries formation. The study of natural and induced factors influencing the demineralization process has been greatly facilitated by the use of bacteriological systems in which demineralization is produced by direct colonization of cariogenic microorganisms on the surfaces of extracted teeth. Comparison of results obtained with these latter systems and with chemical systems has allowed us, for example, to elucidate the mechanism by which acquired salivary pellicles and fluoride topical solutions decrease the rate of enamel demineralization. The pellicle retards transport of matter across the enamel surface, whereas the fluoride topical solutions decrease the cariogenicity of the colonizing bacteria.

Contribution of stereochemical interactions in the adhesion of Streptococcus sanguis C5 to experimental pellicles.

Parameters for the adsorption of Streptococcus sanguis C5 to experimental salivary pellicles were determined to assess the relative contributions of stereochemical and less specific hydrophobic-electrostatic interactions in this process. S. sanguis C5 cells possess hydrophobic surfaces and also an adhesin which exhibits stereochemical binding to sialic acid-containing salivary receptors. The adsorption of S. sanguis C5 to untreated pellicles was significantly better described by a two-site adsorption model than by a one-site model (p less than 0.001). One class of binding sites had an affinity for the organism that was more than 400 times that of the second class of binding sites. However, adsorption of the organism was better described by a one-site model to asialo pellicles prepared by neuraminidase treatment which destroyed the receptors for the organism's adhesin. The affinity of the organism to sites in asialo pellicles was low, and approximated that to the low-affinity sites in untreated pellicles. These observations suggest that, in the absence of stereochemical interactions involving the organism's adhesin and specific salivary receptors, S. sanguis C5 cells only exhibit low-affinity binding to experimental pellicles. This was substantiated by studies which showed that adsorption of the organism to albumin-coated hydroxyapatite and to polystyrene beads was also better described by a one-site model, and it occurred with only low affinity.

Adherent interactions which may affect microbial ecology in the mouth.

Bacteria attach to teeth and oral mucosal surfaces in a surprisingly selective manner, and attachment appears to be the first step in the colonization process. Attachment is thought to involve lectin-like and/or hydrophobic ligands, called adhesins, often present in bacterial surface appendages which interact with receptors on oral tissues. A variety of factors can influence bacterial attachment, and therefore have the potential to affect host-parasite interactions in the mouth. Factors discussed include salivary components, dietary lectins, and sublethal concentrations of antibiotics.

Computer modeling of the effects of chewing sugar-free and sucrose-containing gums on the pH changes in dental plaque associated with a cariogenic challenge at different intra-oral sites.

Variation in salivary access to different intra-oral sites is an important factor in the site-dependence of dental caries. This study explored, theoretically, how access is modified by chewing sugar-free and sugar-containing gums. A finite difference computer model, described elsewhere, was used. This allowed for diffusion and/or reaction of substrate, acid product, salivary buffers, and fixed-acid groups. Site-dependent saliva/plaque exchange was modeled in terms of a 100-microns-thick salivary film covering the plaque (a) flowing directly from the salivary ducts, (b) flowing from the intra-oral salivary pool, or (c) exchanging with the pool. Computed flow-velocities or rates of exchange were based on previous intra-oral measurements. The model was also tested against an in vitro study conducted by two of the authors. In addition, the three proposed models of saliva/plaque interaction were compared, and the effect of salivary film thickness investigate. Results suggested that: (1) although sugar-free gum chewed during a cariogenic challenge causes a rapid rise in plaque pH, sucrose-containing gums cause the pH, after a temporary rise resulting from increased salivary flow, to stay low for an extended period; (2) the computer model reproduced in vitro tests reasonably well; (3) although the three models of the plaque/saliva interaction start from different assumptions, two lead to closely related predictions; and (4) increasing the assumed salivary film thickness by a large amount (e.g., from 50 to 200 microns) caused no change in modeled Stephan curves, as long as these changes were accompanied by appropriate reductions in film velocity, in accord, theoretically, with the practical clearance data.

Calculus attachment. Review of the literature and new findings.

This study was undertaken in order to provide current information relative to the modes of calculus attachment to tooth surfaces. A total of 63 freshly extracted teeth were fixed, sectioned, and conventionally prepared for light, transmission electron, and scanning electron microscopic examination. Previously reported histologic findings of cuticular attachment, mechanical locking into undercuts, and direct attachment of calculus matrix to the tooth surface were affirmed. The claim of bacterial penetration as a mode of attachment has been rejected. Ultrastructural evidence of cuticular attachment has been presented for the first time via a series of scanning electron micrographs. The most frequently encountered method of attachment was found to be the apparent melding of calculus matrix to the surface of cementum. In many instances, these two substances were virtually indistinguishable.

Interactions between denture lining material, protein pellicles and Candida albicans.

The interactions between pellicles derived from saliva, serum, mucin and lysozyme deposited on lining material (tissue conditioner) and Candida albicans were investigated by monitoring pH changes associated with protein-free and protein-coated lining material and by ultrastructural observations of yeast colonization. No significant differences in pH reduction between culture media in contact with the protein-free, control lining materials and those coated with saliva, serum or mucin were observed after 120 h of incubation. However, scanning electron microscopy revealed that much greater numbers of the yeasts colonized the saliva- or serum-coated lining material than the lysozyme-, mucin-coated or control material. Hyphal invasion was observed in saliva-coated lining material. These results suggested that denture pellicle derived from saliva and/or serum may potentiate candidal colonization of denture lining materials.

An examination of strains of the bacterium Streptococcus vestibularis for relative cariogenicity in gnotobiotic rats and adhesion in vitro.

The cariogenicity and adhesion of six strains of Streptococcus vestibularis were compared with those of strains of Strep. salivarius. All strains of Strep. vestibularis produced low levels of caries, confined to the fissures, whereas the two strains of Strep. salivarius produced high levels of caries, with one strain producing approximal as well as fissure caries. The values for adhesion to saliva-coated hydroxyapatite of Strep. vestibularis in the absence (median 4.74%) and the presence (median 4.67%) of sucrose were not statistically different, nor did they differ significantly from those of Strep. salivarius. Strains of Strep. vestibularis were able to adhere to buccal epithelial cells (median 1.19%) as well as could Strep. salivarius strain HB (1.65%). Neither sucrose nor saliva greatly aggregated the strains of either species. Strep. vestibularis did not adhere to hexadecane (median 18.5%) to the same extent as did Strep. salivarius strains (median 69%). There was a significant correlation between the adhesion in the presence and absence of sucrose (p less than 0.01). Strep. vestibularis strains could not coaggregate with either actinomycetes or Veillonella spp. whereas Strep. salivarius strains were able to coaggregate with Veillonella spp.

Modulation of bacterial binding to salivary pellicle by treatment with hydrophilizing compounds.

Two hydrophilizing agents, a branched polyethylene glycol derivative and a non-ionic cellulose ether (EHEC) bind to buffer-treated hydroxyapatite and prevent attachment of Streptococcus mutans. EHEC gives a more efficient surface modification, presumably due to a more complete surface coverage. Neither of the 2 hydrophilizing agents were effective on hydroxyapatite which had been pre-treated with saliva. In a small clinical trial, EHEC was found to be moderately effective in preventing plaque formation.

Role of cryptic receptors (cryptitopes) in bacterial adhesion to oral surfaces.

Progress in characterizing the receptors that promote bacterial attachment to teeth and oral epithelial cells has suggested that hidden molecular segments may frequently be involved. Such cryptic receptors, referred to as 'cryptitopes', may become exposed by several mechanisms. Hidden segments of salivary acidic proline-rich proteins evidently become exposed when the molecules undergo a conformational change as they adsorb to apatitic mineral. Adhesins of Actinomyces viscosus and certain other prominent dental plaque bacteria are able to bind to these cryptitopes, and this enables these organisms to bind to proline-rich proteins on apatitic surfaces while avoiding interactions with these proteins in solution. Cryptitopes may also become exposed as a result of enzymatic action. Thus, several bacteria, including Fusobacterium nucleatum, Eikenella corrodens, A. viscosus, A. naeslundii and Bacteroides intermedius, have adhesins that bind to galactosyl receptors which become exposed after treatment with neuraminidase. Similarly, the adhesion of some Gram-negative bacteria, such as Bact. gingivalis, is enhanced when tissue surfaces are treated with certain proteases, or lysosomal enzymes derived from human polymorphonuclear leucocytes. It seems likely that elevated levels of enzymes present in gingival fluid as sequelae of poor oral hygiene and gingivitis may generate cryptitopes for potentially periodontopathic bacteria, and thereby contribute to modulation of the gingival flora.

Experimental salivary pellicles formed on titanium surfaces mediate adhesion of streptococci.

The goal of this study was to characterize salivary components of titanium pellicles and to determine how experimental pellicles affect adhesion of several strains of streptococci to titanium surfaces. Titanium experimental pellicles were formed by incubation of fresh human parotid or human submandibular-sublingual saliva on pure titanium beads. Pellicle was recovered from the beads using sodium dodecyl sulfate buffer and was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting to identify adsorbed salivary components. Streptococcus anginosus, S. oralis, and S. salivarius recovered from in vivo titanium plaque and five reference strains of streptococci were used in adhesion assays to titanium beads with and without experimental salivary pellicles. The experimental pellicle formed on titanium was found to be composed of selected proteins from human parotid and human submandibular-sublingual saliva. Salivary alpha-amylase and proline-rich proteins were found in all experimental pellicles, while sIgA, high-molecular weight mucin, and proline-rich glycoproteins were detected in one of the experimental pellicles examined. Adhesion of fresh isolates and reference stains of S. anginosus, S. oralis, and S. salivarius to saliva-coated titanium was reduced compared to that of titanium without saliva coating. However, adhesion of laboratory strains of S. gordonii and S. sanguis was found to be significantly greater to experimental pellicles of human submandibular-sublingual saliva than was the adhesion of the fresh isolates, suggesting that streptococci-colonizing implant surfaces may be inherently less adhesive than other bacterial strains. This study found that salivary pellicles are selectively formed on titanium and mediate in vitro adhesion of streptococci.

The effect of saliva or serum on Streptococcus mutans and Candida albicans colonization of hydroxylapatite beads.

OBJECTIVE: Several recent reports imply the possibility of cariogenicity of denture plaque containing Candida albicans. Hence the purpose of this study was to investigate the effects of salivary and serum pellicles on C. albicans and Streptococcus mutans colonization on hydroxylapatite beads. METHODS: The colonization of three isolates of C. albicans and two isolates of S. mutans was examined by the use of a bioluminescent adenosine triphosphate (ATP) assay based on the firefly luciferase-luciferin system. RESULTS: In the preliminary study, a good correlation was observed between the cell number and ATP amount of each isolate tested, and the results yielded a level of significance (P < 0.001; Student's t-test), confirming the validity of this method. When the relative ATP content of the 48 h colonization of both isolates of S. mutans were compared, a saliva pellicle was significantly more effective in promoting bacterial colonization than either uncoated or serum pellicle (ANOVA; P < 0.01). In contrast, in the case of colonization of C. albicans isolates, a serum pellicle was significantly more effective in promoting the colonization of C. albicans GDH 18 and GDH 19, than both uncoated specimens and saliva pellicle (ANOVA; P < 0.01). Similar trends were observed with C. albicans GDH 16, though significant differences were not observed (ANOVA; P > 0.05). CONCLUSION: The results suggest that the mechanism involved in fungal colonization on hydroxylapatite (HAP) should be different from that of mutans streptococci.

Syndromes with salivary dysfunction predispose to tooth wear: Case reports of congenital dysfunction of major salivary glands, Prader-Willi, congenital rubella, and Sjögren's syndromes.

Four cases-of congenital dysfunction of the major salivary glands as well as of Prader-Willi, congenital rubella, and Sjögren's syndromes-were identified in a series of 500 patients referred for excessive tooth wear. Although there was evidence of consumption of highly acidic drinks, some occlusal parafunction, and unacceptable toothbrushing habits, salivary dysfunction was the salient factor predisposing a patient to tooth wear in these syndromal cases. The 500 subjects have been characterized either as having medical conditions and medications that predispose them to xerostomia or lifestyles in which workplace- and sports-related dehydration lead to reduced salivary flow. Normal salivation, by buffering capacity, clearance by swallowing, pellicle formation, and capacity for remineralization of demineralized enamel, protects the teeth from extrinsic and intrinsic acids that initiate dental erosion. Thus, the syndromes, unrelated in many respects, underline the importance of normal salivation in the protection of teeth against tooth wear by erosion, attrition, and abrasion.

Adhesive bonding of various materials to hard tooth tissues--solubility of dentinal smear layer in dilute acid buffers.

While the acid etch technique has largely solved the problem of bonding restorative materials to enamel, attempts to improve adhesive bonding to dentin have so far had only limited success. One aspect to which insufficient attention has been paid is the presence of a smear layer on cut surfaces and of pellicle on surfaces exposed to saliva, both of which would reduce or prevent durable adhesive bonds. The present study attempts to evaluate the usefulness of various acidic solutions in removing these surface layers while causing minimal damage to the vital contents of the dentinal tubules. A series of twelve 0.16 M buffer solutions were applied to cut dentin surfaces which were then washed, dried and sputter-coated with gold/palladium in preparation for examination by scanning electron microscopy. It is concluded that the smear layer on cut dentin surfaces can be substantially removed in vitro by a 30 second exposure to an isotonic solution of buffered monobasic acids having pKa values between 3.8 and 2.5. Since these solutions did not always remove material from grooves in the prepared surface clinical techniques should aim at producing a smooth cut surface. The response of the vital pulp to this treatment still awaits investigation.

The hard tissue lesion revisited.

Recent research indicates that the thorough removal of calculus is still a critical step in the treatment of periodontal disease. Surgical access facilitates this in deep pockets. Repair is mediated by a long junctional epithelium as microscopically total calculus removal is rarely achieved. In intrabony defects the apical portion of the root surface which has not attracted deposits of calculus has the potential for connective tissue reattachment and regeneration if care is taken to preserve collagen fibres still attached to the root.