Trypsin-susceptible cell surface characteristics of Streptococcus sanguis

The adherence of Streptococcus sanguis to saliva-coated hydroxylapatite was markedly reduced by treatment of the cells with trypsin. In Scatchard plots of adherence data, protease-treated S. sanguis did not exhibit the characteristic positive slopes, suggesting that trypsin prevented cooperative interactions between the cells and artificial pellicle. Trypsin also reduced the tendency of S. sanguis to bind to hexadecane and to octyl-Sepharose. When sodium dodecyl sulfate was used to elute S. sanguis from columns of octyl-Sepharose, it was observed that the elution profiles of trypsin-treated cells were more complex than those of control cells. Water and salts were incapable of removing the cells from octyl-Sepharose. The results suggest that adherence to saliva-coated hydroxylapatite, binding to hexadecane and to octyl-Sepharose depend on trypsin-susceptible cell surface molecules.     

Salivary film expresses a complex, macromolecular binding site for Streptococcus sanguis

Teeth in the oral cavity are coated with a salivary film or pellicle, which lacks apparent intermolecular organization. This heterogeneous film facilitates binding of early commensal colonizing bacteria, including Streptococcus sanguis. To test the hypothesis that sufficient intermolecular organization exists in salivary films to form binding sites for S. sanguis, an in vitro model of saliva-coated teeth was probed with murine anti-idiotypical monoclonal antibodies (mAb2, anti-ids). The anti-ids were harvested from hybridomas that were developed in response to first generation murine hybridomas that produced anti-S. sanguis adhesin monoclonal antibodies (mAb1). The anti-ids (i) reacted with experimental salivary films and inhibited S. sanguis adhesion in a dose-dependent fashion. In Western blots, the anti-ids (ii) recognized a high molecular weight salivary antigen and (iii) secretory IgA (sIgA) light chain and alpha-amylase. After isolation by gel filtration from whole saliva or mixed secretory IgA and alpha-amylase, the high molecular weight component, containing amylase activity and sIgA, bound to hydroxyapatite to promote adhesion of S. sanguis. Therefore, a complex enriched in secretory immunoglobulin A and alpha-amylase forms a S. sanguis-binding site.     

Inhibition of the cooperative adhesion of Streptococcus sanguis to hydroxylapatite

The adhesion of Streptococcus sanguis to hydroxylapatite is a process involving several adhesins and receptors. Binding isotherms and Scatchard plots of the adhesion suggest that cooperative interactions occur at low cell densities. It was found that sulfolane, a hydrophobic-bond diluent, was capable of inhibiting the cooperative adhesion of S. sanguis to saliva-coated hydroxylapatite beads. Sodium thiocyanate, a chaotropic agent, inhibited not only cooperative adhesion, but also the adhesion thought to result from noncooperative interactions. It is suggested that strong chaotropic agents may not only inhibit adhesin-receptor complexes, but also may influence the secondary/tertiary structures of interacting species.     

Calorimetric studies of the binding of Streptomyces subtilisin inhibitor to subtilisin of Bacillus subtilis strain N'

The binding of Streptomyces subtilisin inhibitor (SSI) to subtilisin of Bacillus subtilis strain N' (subtilisin BPN', EC 3.4.21.14) was studied by isothermal calorimetry at pH 7.0 and at various temperatures ranging from 5 to 30 degrees C. Thermodynamic quantities for the binding reaction were derived as a function of temperature by combining the data reported for the dissociation constant with the present calorimetric results. At 25 degrees C, the values are delta G degrees = -57.9 kJ mol-1, delta H = -19.8 kJ mol-1, delta S degree = 0.13 kJ K-1 mol-1, and delta Cp = -1.02 kJ K-1 mol-1. The entropy and the heat capacity changes are discussed in terms of the contributions from the changes in vibrational modes and in hydrophobic interactions.     

In vitro inhibition of Streptococci binding to enamel acquired pellicle by plant lectins

AIM: Initial colonization of the tooth surface by streptococci involves the attachment of these bacteria to adsorbed salivary components of the acquired pellicle. In dental biofilm this adhesion may also involve lectin-like components, present on the surface of the organisms, which bind to complementary carbohydrates on the surface of the tooth. Therefore, this work aimed to evaluate the potential of six lectins, extracted from seeds of Leguminosae family members, to inhibit the adherence of five streptococci species to acquired pellicle in vitro. METHODS AND RESULTS: The lectins used in this work were extracted from Canavalia ensiformis, Canavalia brasiliensis, Dioclea violacea, Dioclea grandiflora, Cratylia floribunda and Vatairea macrocarpa. Fluorescence micrography was employed to visualize the ability of FITC-labeled lectins to attach to acquire pellicle. Adherence inhibition was performed on saliva-coated microtiter plates at which lectins solutions were previously incubated followed by incubation with the oral streptococci. Glucose-mannose specific lectins attached to acquired pellicle with high intensity, while galactose specific lectins, from V. macrocarpa, exhibits low intensity attachment. CONCLUSIONS: All lectins were able to inhibit the adherence of the microorganisms tested (p < 0.01). SIGNIFICANCE AND IMPACT OF THE STUDY: Our results suggest that lectins may be useful in anti adhesion therapeutics.     

Crystalline anatase-rich titanium can reduce adherence of oral streptococci

Dental implant abutments that emerge through the mucosa are rapidly covered with a salivary protein pellicle to which bacteria bind, initiating biofilm formation. In this study, adherence of early colonizing streptococci, Streptococcus gordonii, Streptococcus oralis, Streptococcus mitis and Streptococcus sanguinis to two saliva-coated anodically oxidized surfaces was compared with that on commercially pure titanium (CpTi). Near edge X-ray absorption (NEXAFS) showed crystalline anatase was more pronounced on the anodically oxidized surfaces than on the CpTi. As revealed by fluorescence microscopy, a four-species mixture, as well as individual bacterial species, exhibited lower adherence after 2?h to the saliva-coated, anatase-rich surfaces than to CpTi. Since wettability did not differ between the saliva-coated surfaces, differences in the concentration and/or configuration of salivary proteins on the anatase-rich surfaces may explain the reduced bacterial binding effect. Anatase-rich surfaces could thus contribute to reduced overall biofilm formation on dental implant abutments through diminished adherence of early colonizers.     

The conditioning role of saliva in streptococcal attachment to hydroxyapatite surfaces

The solution properties of saliva and its role in conditioning both the substrate and the bacterial surface have been investigated with regard to the attachment of oral streptococci to hydroxyapatite surfaces. Saliva from eight subjects was used and the attachment of three organisms, Streptococcus mutans strains FA-1 (serotype b) and KPSK2 (serotype c) and S. sanguis T175-1, was studied. An adsorbed salivary layer on a hydroxyapatite surface substantially reduced the affinity of the organisms for the surface. Adsorbed saliva on the bacterial surfaces, however, tended to increase the organisms' affinity for saliva-coated apatite. The source of saliva was important in determining the extent of inhibition of attachment. The data indicated that the negatively charged and hydrophilic nature of salivary conditioning films was important in controlling bacterial adsorption to hydroxyapatite. The results also suggested that hydrophobic salivas could promote binding of the more hydrophobic bacteria known to be early colonizers of the teeth.     

In vitro inhibition of oral streptococci binding to the acquired pellicle by algal lectins

AIMS: The initial colonization of the tooth by streptococci involves their attachment to adsorbed components of the acquired pellicle. Avoiding this adhesion may be successful in preventing caries at early stages. Salivary mucins are glycoproteins that when absorbed onto hydroxyapatite may provide binding sites for certain bacteria. Algal lectins may be especially interesting for oral antiadhesion trials because of their great stability and high specificity for mucins. This work aimed to evaluate the potential of two algal lectins to inhibit the adherence of five streptococci species to the acquired pellicle in vitro. METHODS AND RESULTS: The lectins used were extracted from Bryothamnion triquetrum (BTL) and Bryothamnion seaforthii (BSL). Fluorescence microscopy was applied to visualize the ability of fluorescein isothiocyanate-labelled lectins to attach to the pellicle and revealed a similar capability for both lectins. Streptococcal adherence assays were performed using saliva-coated microtitre plates. BSL inhibited more than 75% of Streptococcus sanguis, Streptococcus mitis, Streptococcus sobrinus and Streptococcus mutans adherence, achieving 92% to the latter. BTL only obtained statistically significant results on S. mitis and S. sobrinus, whose adherence was decreased by 32.5% and 54.4%, respectively. CONCLUSION: Algal lectins are able to inhibit streptococcal adherence. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results support the proposed application of lectins in antiadhesion therapeutics.     

Thermodynamic properties of ligand binding by monoclonal anti-fluorescyl antibodies

The effects of temperature on the binding of fluorescein by three monoclonal anti-fluorescyl antibodies (4-4-20, 20-19-1, and 20-20-3) were assessed by measurements of affinity constants (Ka) over a temperature range of 2-70 degrees C. Values for Ka were determined from the degree of ligand association by using fluorescence methodology. Curvilinear van't Hoff plots (ln Ka vs. T-1) were observed for all three antibodies, indicating that their standard enthalpy changes (delta Ho) were temperature dependent. This phenomenon was further investigated by plotting the changes in unitary free energy (delta Gu), standard enthalpy (delta Ho), and unitary entropy (delta Su) vs. temperature. Strong temperature dependencies were observed for enthalpy and entropy values, while free energy plots were only weakly dependent on temperature. At low temperatures (4 degrees C), entropy played a major role in the binding of fluorescein by all three antibodies, while enthalpy dominated at higher temperatures. This was a consequence of the negative heat capacity changes (delta Cpo approximately equal to -320 cal K-1 mol-1) observed for these antibodies, which produced a negative trend in both enthalpy and entropy values with increasing temperature. The negative heat capacity values also indicated that the hydrophobic effect was instrumental in the binding of fluorescein. Entropy changes were lower than expected for hydrophobic binding alone, suggesting that other forces were acting to mitigate the hydrophobic effect. One possibility was that the binding of fluorescein acted to restrain vibrational fluctuations in the active-site region, producing negative changes in both heat capacity and entropy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reaction heats and heat capacity changes for intermediate steps of the ATP hydrolysis catalyzed by myosin subfragment 1

The interaction of myosin Subfragment 1 with ATP in 0.1 M KCl containing 0.01 M MgCl2 and 0.02 M Tris/HCl (pH 8.0) was studied by microcalorimetry at temperatures of 4, 12, and 23 degrees C so that values of the heat capacity change (delta Cp) could be obtained for intermediate steps of the ATPase cycle. The delta Cp values are large compared to the value for the overall cycle, indicating that large changes in the hydrophobic effect are involved in transitions between different intermediate states. However, the heat capacity changes themselves show peculiar temperature dependences. Thus bindings of ATP and ADP to Subfragment 1, both of which are strongly exothermic processes, take place with large negative delta Cp of about -3 kJK-1 mol-1 between 4 and 12 degrees C but with very small delta Cp of 0.3-0.4 kJ K-1 mol-1 between 12 and 23 degrees C. On the contrary, the delta Cp for the endothermic hydrolysis of ATP bound to Subfragment 1 is positive (congruent to kJK-1 mol-1) in the lower temperature range but strongly negative (congruent to -4 kJK-1 mol-1) in the higher temperature range. The magnitude of delta Cp for the slow Pi dissociation process is similar but its sign is just opposite to that for the hydrolysis. These anomalous changes in the heat capacity may be due to the temperature-induced changes in a balance between large opposing effects which result from distinct, local conformation changes within the Subfragment 1 molecule.     

Nonclassical hydrophobic effect in membrane binding equilibria.

The enthalpy of transfer of four different amphiphilic molecules from the aqueous phase to the lipid membrane was determined by titration calorimetry. The four molecules investigated were the potential-sensitive dye 2-(p-toluidinyl)naphthalene-6-sulfonate (TNS), the membrane conductivity inducing anion tetraphenylborate (TPB), the Ca2+ channel blocker amlodipine [B?uerle, H. D., & Seelig, J. (1991) Biochemistry 30, 7203-7211], and the positively charged local anesthetic dibucaine. All four amphiphiles penetrate into the hydrophobic part of the membrane, and their binding constants, after correcting for electrostatic effects, range between 600 M-1 for dibucaine and 60,000 M-1 for tetraphenylborate. The corresponding changes in free energy were about -6 to -9 kcal/mol. Binding of the amphiphiles to membrane vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine was accompanied by exothermic heats of reaction for all four molecules. For TNS, TPB, and amlodipine, the enthalpies of transfer were almost identical and corresponded to delta H approximately -9 kcal/mol, essentially accounting for the total free energy change. Thus, the binding of these charged amphiphiles to the hydrophobic membrane was driven by enthalpy. This is in contrast to the classical hydrophobic effect, where the transfer is considered to be entropy driven. For dibucaine, the enthalpy of transfer was smaller with delta H approximately -2 kcal/mol but was still about one-third of the total free energy change. All enthalpies of transfer exhibited a distinct temperature dependence with molar heat capacities delta Cp of -30 to -100 cal mol-1K-1 for the transfer from water to the membrane.(ABSTRACT TRUNCATED AT 250 WORDS)     

Aggregation of Streptococcus sanguis biotypes I and II by parotid saliva: a comparison between peritrichously fibrillar and tufted strains

Twelve strains of Streptococcus sanguis biotype I and seven strains of Streptococcus sanguis biotype II carrying either peritrichous fibrils or tufts of fibrils, were examined for their susceptibility to aggregation by parotid saliva. Salivary aggregation was evaluated using a spectrophotometric measurement of sedimentation to assess clump size. A clear distinction emerged between structural sub-groups. Irrespective of biotype, strains carrying peritrichous fibrils aggregated strongly whilst tufted strains were little affected. The one strain with peritrichous fimbriae as well as fibrils, was not aggregated by saliva. Pre-treatment of two peritrichously fibrillar strains with parotid saliva reduced their ability to adhere to parotid saliva-coated hydroxyapatite, whereas adhesion of two tufted strains was not inhibited. Inhibition of adhesion may have been due to steric hindrance, but blocking of bacterial adhesins by saliva components could not be discounted.     

Moderation of oral bacterial adhesion on saliva-coated hydroxyapatite by polyaspartate

AIMS: Synthetic sodium alpha,beta-polyaspartate (PA) has been investigated as a moderator of adhesion and the subsequent biofilm formation by oral bacteria. METHODS AND RESULTS: The inhibition of bacterial adhesion by PA was assessed by (i) a 30-min incubation with Streptococcus sanguis in a microtitre assay with the wells coated with hydroxyapatite (HAP) and (ii) an 18-h challenge with human salivary microflora in a HAP disc assay. In contrast to HAP-coated surfaces, clean polystyrene surfaces in the microtitre assay exhibited no anti-adhesion properties. It has been found that PA significantly and similarly adsorbs onto HAP surfaces in the presence and absence of salivary coating. The HAP disc assay also showed that PA, both in aqueous solutions and in toothpaste, reduced the level of adhered microflora and this effect was enhanced by added propylene oxide-ethylene oxide copolymers. CONCLUSION: The principal finding from this work is the potential role for PA as an inhibitor of dental plaque formation. PA may significantly modify the salivary pellicle. SIGNIFICANCE AND IMPACT OF THE STUDY: This work indicates the use of PA in controlling the development of dental plaque and the formation of bacterial biofilm in general.     

Thermodynamic analysis of the folding of the streptococcal protein G IgG-binding domains B1 and B2: why small proteins tend to have high denaturation temperatures.

We have cloned, expressed, and characterized two naturally occurring variations of the IgG-binding domain of streptococcal protein G. The domain is a stable cooperative folding unit of 56 amino acids, which maintains a unique folded structure without disulfide cross-links or tight ligand binding. We have studied the thermodynamics of the unfolding reaction for the two versions of this domain, designated B1 and B2, which differ by six amino acids. They have denaturation temperatures of 87.5 degrees C and 79.4 degrees C, respectively at pH 5.4, as determined by differential scanning calorimetry. Thermodynamic state functions for the unfolding reaction (delta G, delta H, delta S, and delta Cp) have been determined and reveal several interesting insights into the behavior of very small proteins. First, though the B1 domain has a heat denaturation point close to 90 degrees C, it is not unusually stable at physiologically relevant temperatures (delta G = 25 kJ/mol at 37 degrees C). This behavior occurs because the stability profile (delta G vs temperature) is flat and shallow due to the small delta S and delta Cp for unfolding. Related to this point is the second observation that small changes in the free energy of unfolding of the B-domain due to mutation or change in solvent conditions lead to large shifts in the heat denaturation temperature. Third, the magnitude and relative contributions of hydrophobic vs nonhydrophobic forces (per amino acid residue) to the total free energy of folding of the B-domain are remarkably typical of other globular proteins of much larger size.     

Thermodynamic analyses of triplex formation with homopurine oligonucleotide.

We analyzed the thermodynamics of purine motif triplex formation by isothermal titration calorimetry. The signs of calorimetric enthalpy change, delta Hcal, and entropy change, delta S, of the triplex formation were negative in the temperature range between 15 and 35 degrees C. Since an observed negative delta S was unfavorable for the triplex formation, the triplex formation was driven by a large negative delta Hcal. delta Hcal decreased with increasing temperature, yielding a negative heat capacity change, delta Cp, of approximately -1.2 kcal mol-1 K-1. We found that the binding constant, Ka, increased with increasing temperature, leading to an apparent positive van't Hoff enthalpy change, delta Hvh, which was in sharp contrast with the large negative delta Hcal. The analyses of the observed temperature dependence of Ka and delta Hcal and the negative delta Cp suggest that the purine motif triplex formation near room temperature is not a simple two-state binding process but exhibits multiple states, which was previously observed for the pyrimidine motif triplex formation near room temperature.     

Role of Type 1 Fimbriae in the Adhesion of Escherichia coli to Salivary Mucin and Secretory Immunoglobulin A

Saliva is known to modulate the adhesion of bacteria in the oral cavity. The present work was performed to assess the effect of salivary components on the adhesion of Escherichia coli to a model oral surface. Several genetically engineered E. coli strains were used to examine the role of type 1 fimbriation in the interaction of these strains with salivary components in solution or adsorbed to hydroxyapatite. High (MG1) and low (MG2) molecular weight salivary mucins, and secretory immunoglobulin A (sIgA), were found to interact with the surface of E. coli, and these interactions were independent of the expression of fimbriae or capsule. In contrast, fimbriated strains of E. coli adhered to a greater extent to saliva-coated synthetic hydroxyapatite (HAP) than did nonfimbriated strains. Testing of salivary components separated by gel filtration chromatography revealed that only high-molecular-weight components promoted adhesion of E. coli to HAP. Additional studies found that purified MG2 and sIgA promoted the adhesion of E. coli to HAP. Expression of type 1 fimbriae enhanced adhesion, while mannose inhibited adhesion of fimbriated strains, to saliva-coated HAP and to HAP coated with MG2 and sIgA. We conclude that salivary MG2 and sIgA may provide receptors for the adhesion of type 1 fimbriated E. coli to oral surfaces. Received: 10 February 1996 / Accepted: 11 March 1996     

Stability mutants of staphylococcal nuclease: large compensating enthalpy-entropy changes for the reversible denaturation reaction

By use of intrinsic fluorescence to determine the apparent equilibrium constant Kapp as a function of temperature, the midpoint temperature Tm and apparent enthalpy change delta Happ on reversible thermal denaturation have been determined over a range of pH values for wild-type staphylococcal nuclease and six mutant forms. For wild-type nuclease at pH 7.0, a Tm of 53.3 +/- 0.2 degrees C and a delta Happ of 86.8 +/- 1.4 kcal/mol were obtained, in reasonable agreement with values determined calorimetrically, 52.8 degrees C and 96 +/- 2 kcal/mol. The heat capacity change on denaturation delta Cp was estimated at 1.8 kcal/(mol K) versus the calorimetric value of 2.2 kcal/(mol K). When values of delta Happ and delta Sapp for a series of mutant nucleases that exhibit markedly altered denaturation behavior with guanidine hydrochloride and urea were compared at the same temperature, compensating changes in enthalpy and entropy were observed that greatly reduce the overall effect of the mutations on the free energy of denaturation. In addition, a correlation was found between the estimated delta Cp for the mutant proteins and the d(delta Gapp)/dC for guanidine hydrochloride denaturation. It is proposed that both the enthalpy/entropy compensation and this correlation between two seemingly unrelated denaturation parameters are consequences of large changes in the solvation of the denatured state that result from the mutant amino acid substitutions.     

Thermodynamics of follitropin binding to solubilized calf testis receptor

Thermodynamic parameters of follitropin binding to solubilized testicular receptors were measured in order to assess the forces involved in the binding reaction. Reversibility of follitropin binding to solubilized receptor decreased only 20% over the temperature range 4-24 degrees C, whereas earlier studies indicated reversibility of binding to membrane-bound receptor decreased by more than 40% over the same range [Anderson, T. T., Curatolo, L. M., & Reichert, L. E., Jr. (1983) Mol. Cell. Endocrinol. 33, 37-52]. Thermodynamic analysis of follitropin binding to solubilized receptors showed that the hydrophobic effect was important in the binding reaction. The mean values, at 25 degrees C, for delta H and delta S were -31.8 kcal/mol and -66.0 cal mol-1 K-1, respectively, and delta Cp was -3.0 kcal mol-1 K-1. This is an unusually large heat capacity for protein-protein association reactions, indicating an enhanced role for the hydrophobic effect with the solubilized (compared to membrane-bound) receptor. Since glycerol was necessary to stabilize the solubilized receptor, we determined whether glycerol affected the thermodynamic parameters measured for the binding reaction. Control experiments, performed with membrane-bound receptor in the presence or absence of glycerol, indicated that delta Cp actually decreased upon addition of glycerol (-0.8 kcal mol-1 K-1 in the presence of glycerol compared to -2.3 kcal mol-1 K-1 in the absence of glycerol). Thus, the large negative delta Cp observed for the soluble receptor was a result of its removal from the membrane and was not due to the presence of glycerol.(ABSTRACT TRUNCATED AT 250 WORDS)     

A comparison of the adhesive properties and surface ultrastructure of the fibrillar Streptococcus sanguis 12 and an adhesion deficient non-fibrillar mutant 12 na

Streptococcus sanguis 12 and a naturally occurring mutant, 12 na, were screened for their ultrastructure and adhesive properties in vitro. Negative staining showed that Strep. sanguis 12 carried three types of surface structure. The majority of cells carried long fibrils that in different batches ranged in length from 80 to 207 nm, and shorter fibrils which were 51.0 +/- 15.7 nm long. Both types of fibrils were primarily located at the poles of the cells. Occasionally cells were seen that carried fimbriae, which are structurally distinct from fibrils, and were 3-4 nm wide and less than 1.0 micron long. Strain 12 na carried no detectable surface structures. Ruthenium red staining revealed that both strains carried a loose, amorphous, extracellular polysaccharide layer attached to the cell wall. Streptococcus sanguis 12 na was 83% less adhesive than strain 12 in a saliva-coated hydroxyapatite assay, and 50% less adhesive in a buccal epithelial cell adhesion assay. In contrast, strain 12 na was more sensitive to aggregation by parotid saliva than strain 12, and both strains were equally aggregated by whole saliva. The cell surface hydrophobicity of the two strains was similar. Extraction of surface proteins by sodium lauroyl-sarcosinate followed by sodium dodecylsulphate polyacrylamide gel electrophoresis demonstrated that Strep. sanguis 12 expressed more high mol.wt proteins on its surface than strain 12 na. Using immunogold labelling, the fibrils of strain 12 labelled well with antiserum directed against the long fibrils, but so did the cell surfaces of both Strep. sanguis 12 and 12 na. High molecular weight proteins and cell surface fibrils may be associated with adhesion in this strain.     

A comparison of the adhesive properties and surface ultrastructure of the fibrillar Streptococcus sanguis 12 and an adhesion deficient non-fibrillar mutant 12 na

Streptococcus sanguis 12 and a naturally occurring mutant, 12 na, were screened for their ultrastructure and adhesive properties in vitro. Negative staining showed that Strep. sanguis 12 carried three types of surface structure. The majority of cells carried long fibrils that in different batches ranged in length from 80 to 207 nm, and shorter fibrils which were 51mD0 PT 15mD7 nm long. Both types of fibrils were primarily located at the poles of the cells. Occasionally cells were seen that carried fimbriae, which are structurally distinct from fibrils, and were 3mD4 nm wide and <1mD μm long. Strain 12 na carried no detectable surface structures. Ruthenium red staining revealed that both strains carried a loose, amorphous, extracellular polysaccharide layer attached to the cell wall. Streptococcus sanguis 12 na was 83% less adhesive than strain 12 in a saliva-coated hydroxyapatite assay, and 50% less adhesive in a buccal epithelial cell adhesion assay. In contrast, strain 12 na was more sensitive to aggregation by parotid saliva than strain 12, and both strains were equally aggregated by whole saliva. The cell surface hydrophobicity of the two strains was similar. Extraction of surface proteins by sodium lauroyl-sarcosinate followed by sodium dodecylsulphate polyacrylamide gel electrophoresis demonstrated that Strep. sanguis 12 expressed more high mol.wt proteins on its surface than strain 12 na. Using immunogold labelling, the fibrils of strain 12 labelled well with antiserum directed against the long fibrils, but so did the cell surfaces of both Strep. sanguis 12 and 12 na. High molecular weight proteins and cell surface fibrils may be associated with adhesion in this strain.