Comparative opsonic activity for Steptococcus mutans in oral fluids, and phagocytic activity of blood, crevicular, and salivary polymorphonuclear leucocytes in rhesus monkeys

The opsonic activity for Streptococcus mutans was assayed in serum, gingival crevicular fluid, parotid saliva and mixed saliva from rhesus monkeys immunized against dental caries with a vaccine of Streptococcus mutans in Freund's incomplete adjuvant. The phagocytic activities of polymorphonuclear leucocytes from the blood and gingival crevice were compared, and the effects of gingival crevicular fluid and mixed saliva on blood polymorphonuclear leucocyte viability and phagocytic activity were assessed. Heat-stable opsonic activity was detected in sera, crevicular fluid, and mixed saliva of immunized animals. Polymorphonuclear leucocytes from the gingival crevice and saliva retained viability, although this was lower than in cells from blood. Exposure of blood polymorphonuclear leucocytes to crevicular fluid or mixed saliva for 30 min resulted in no loss of cell viability or phagocytic activity, but saliva was cytotoxic on prolonged exposure. These results support the hypothesis that the opsonization and phagocytosis of cariogenic bacteria might be a mechanism involved in the immunological protection against dental caries.     

Effect of peroral immunization of humans with Streptococcus mutans on induction of salivary and serum antibodies and inhibition of experimental infection

Naturally occurring antibodies reactive with Streptococcus mutans whole cells were assayed in whole saliva, parotid saliva, and blood samples collected from eight human volunteers. The levels and serotypes of indigenous S. mutans in plaque and whole saliva samples were also determined. After baseline sampling the teeth were cleaned and the subjects were inoculated with streptomycin-resistant S. mutans strains Ingbritt (serotype c) and OMZ65 (serotype g). The level of implantation and duration of colonization were determined in plaque and saliva, and antibodies reactive with these strains were monitored in saliva and serum. After the implanted bacteria were shed, the subjects wee immunized by the daily ingestion of an enteric-coated capsule containing 25 mg of Formalin-killed, freeze-dried OMZ65 cells for 3 days and inoculation was repeated. The levels of antibodies and of implantation and the duration of colonization were monitored as before. One month after the bacteria could no longer be detected, the immunization and inoculation cycle was repeated except that the subjects were immunized for 7 days. Five of the eight subjects were successfully colonized by strains Ingbritt and OMZ65. The remaining three did not become colonized with either strain. Strain OMZ65 implanted at a higher level than did strain Ingbritt. Oral immunization did not result in a detectable antibody response in saliva or serum to whole bacterial cells. However, after both the first and second immunizations there were marked reductions in the peak levels of infection and the duration of colonization of both OMZ65 and Ingbritt.     

Study of humoral immunity to commensal oral bacteria in human infants demonstrates the presence of secretory immunoglobulin A antibodies reactive with Actinomyces naeslundii genospecies 1 and 2 ribotypes

The mouths of three human infants were examined from birth to age 2 years to detect colonization of Actinomyces naeslundii genospecies 1 and 2. These bacteria did not colonize until after tooth eruption. The diversity of posteruption isolates was determined by ribotyping. Using immunoblotting and enzyme-linked immunosorbent assay, we determined the reactivity of secretory immunoglobulin A (SIgA) antibodies in saliva samples collected from each infant before and after colonization against cell wall proteins from their own A. naeslundii strains and carbohydrates from standard A. naeslundii genospecies 1 and 2 strains. A. naeslundii genospecies 1 and 2 carbohydrate-reactive SIgA antibodies were not detected in any saliva sample. However, SIgA antibodies reactive with cell wall proteins were present in saliva before these bacteria colonized the mouth. These antibodies could be almost completely removed by absorption with A. odontolyticus, a species known to colonize the human mouth shortly after birth. However, after colonization by A. naeslundii genospecies 1 and 2, specific antibodies were induced that could not be removed by absorption with A. odontolyticus. Cluster analysis of the patterns of reactivity of postcolonization salivary antibodies from each infant with antigens from their own strains showed that not only could these antibodies discriminate among strains but antibodies in saliva samples collected at different times showed different reactivity patterns. Overall, these data suggest that, although much of the salivary SIgA antibodies reactive with A. naeslundii genospecies 1 and 2 are directed against genus-specific or more broadly cross-reactive antigens, species, genospecies, and possibly strain-specific antibodies are induced in response to colonization.     

Adherence of Porphyromonas (Bacteroides) gingivalis to Streptococcus sanguis in vitro.

Intergeneric bacterial adherence is responsible for the complexity of the microbiota in human dental plaque and is believed to enable some extraneous bacteria to initially colonize the human oral cavity. Some current evidence indicates that Streptococcus sanguis, an early colonizer of teeth, enhances subsequent colonization by Porphyromonas (Bacteroides) gingivalis, a bacterium associated with advanced adult periodontitis. In this study, selected strains of P. gingivalis and S. sanguis were tested for their adherence activities in vitro. A differential filtration assay was devised in which one member of the test pair was radiolabeled. Heterogeneous aggregates that formed in mixed suspensions were collected on polycarbonate filters (8-microns pore size) and were washed free of individual bacteria and small homologous clumps. P. gingivalis 381, W50, JKG7, and 33277 adhered to S. sanguis G9B, M5, Challis 6, and 38. P. gingivalis A7A1-28 did not adhere well to S. sanguis under these conditions. More precise measurements of intergeneric adherence were obtained with an alternative assay with radiolabeled P. gingivalis and an artificial dental plaque composed of S. sanguis coupled to cyanogen bromide-activated agarose beads. CNBr-agarose was selected as the supporting matrix for the plaque because it was uniformly and permanently coated with S. sanguis and because P. gingivalis had negligible adherence activity for streptococcus-free beads. P. gingivalis W50 grown to the early stationary phase adhered to S. sanguis-coated beads in higher numbers than either midlogarithmic- or late-stationary-phase cells. Intergeneric adherence was not inhibited or reversed by the presence of lactose or other monosaccharides or disaccharides. Pretreatment of either bacterium with trypsin or proteinase K reduced subsequent adherence by 86 to 100%. Neuraminidase treatment of P. gingivalis caused 98% reduction of adherence, whereas similar treatment of S. sanguis caused only a 2% loss. Preincubation of P. gingivalis at 60 degrees C for 30 min decreased subsequent adherence to S. sanguis-coated beads by 94%. Adherence was reduced by 96% when bacteria were assayed while suspended in human whole saliva or when pretreated with saliva and subsequently assayed in buffer. The concentration of whole human saliva required to inhibit 50% adherence in this assay was 23 micrograms per ml (1:200 dilution). Suspension of the bacteria in normal rabbit serum resulted in 94% inhibition of adherence. These data indicate that saliva and serum may be important host defense factors for controlling Porphyromonas-Streptococcus adherence.     

Protective secretory immunoglobulin A antibodies in humans following oral immunization with Streptococcus mutans.

Ingestion of a vaccine containing killed Streptococcus mutans, originally isolated from each volunteer, daily for 10 consecutive days induced increased levels of specific secretory immunoglobulin A (sIgA) antibodies to S. mutans cells and two cell surface proteins, glucosyltransferase and surface antigen I/II, in parotid saliva and tears of four healthy males and in parotid saliva, tears, colostrum, and milk of a pregnant woman. In addition, these antibodies inhibited glucosyltransferase activity. Both IgA1 and IgA2 antibodies were induced. The levels of IgA antibodies in all secretions remained significantly above preimmunization levels for more than 50 days after oral administration of antigen. A second series of immunizations for 7 consecutive days resulted in even higher levels of sIgA antibodies, which peaked earlier and persisted longer than those observed after the primary immunizations. No increase in levels of antibodies in serum were detected in any subject. Antibodies reactive with human heart and kidney antigens could not be detected in saliva, tears, colostrum, milk, or serum samples collected at any time during the immunization regimen. The numbers of viable S. mutans organisms in dental plaque and whole saliva decreased after each series of immunizations, which correlated with increased levels of IgA antibodies in saliva, suggesting that IgA antibodies in saliva were responsible for the reduced adherence of this bacterium. These results indicate that ingested S. mutans antigen induces secretion of specific IgA1 and IgA2 antibodies in saliva, tears, colostrum, and milk, providing further evidence for the existence of a common mucosal immune system.     

Study of Humoral Immunity to Commensal Oral Bacteria in Human Infants Demonstrates the Presence of Secretory Immunoglobulin A Antibodies Reactive with Actinomyces naeslundii Genospecies 1 and 2 Ribotypes

The mouths of three human infants were examined from birth to age 2 years to detect colonization of Actinomyces naeslundii genospecies 1 and 2. These bacteria did not colonize until after tooth eruption. The diversity of posteruption isolates was determined by ribotyping. Using immunoblotting and enzyme-linked immunosorbent assay, we determined the reactivity of secretory immunoglobulin A (SIgA) antibodies in saliva samples collected from each infant before and after colonization against cell wall proteins from their own A. naeslundii strains and carbohydrates from standard A. naeslundii genospecies 1 and 2 strains. A. naeslundii genospecies 1 and 2 carbohydrate-reactive SIgA antibodies were not detected in any saliva sample. However, SIgA antibodies reactive with cell wall proteins were present in saliva before these bacteria colonized the mouth. These antibodies could be almost completely removed by absorption with A. odontolyticus, a species known to colonize the human mouth shortly after birth. However, after colonization by A. naeslundii genospecies 1 and 2, specific antibodies were induced that could not be removed by absorption with A. odontolyticus. Cluster analysis of the patterns of reactivity of postcolonization salivary antibodies from each infant with antigens from their own strains showed that not only could these antibodies discriminate among strains but antibodies in saliva samples collected at different times showed different reactivity patterns. Overall, these data suggest that, although much of the salivary SIgA antibodies reactive with A. naeslundii genospecies 1 and 2 are directed against genus-specific or more broadly cross-reactive antigens, species, genospecies, and possibly strain-specific antibodies are induced in response to colonization.     

Use of monoclonal antibodies with neutralizing effects on toxic antigens from human bacterial plaque to detect specific bacteria by colony blotting.

Inflammatory periodontal diseases are provoked by bacteria which adhere to teeth at the gingival margin and form plaques containing toxins detectable by their effect on mammalian cells in culture. The aim of this study was to make toxin-neutralizing monoclonal antibodies and determine whether they detect antigen in specific oral bacteria. Bacterial plaque was collected from teeth and homogenized, and the fluid phase (plaque extract) was boiled or first fractionated over Sephacryl S-300. Hybridomas from immunized mice secreted immunoglobulin M (IgM) antibodies which reacted to plaque antigens. Neutralization was detected by an increase in the growth of HL60 cells which were exposed to plaque toxins in the presence of IgM from hybridoma culture or ascitic fluids. However, the neutralization was obvious only when the plaque toxins reduced growth by 50% or less. Plaque toxin preparations were found to contain proteases which hydrolyzed all of the IgM in ascitic fluids within 24 h. Replenishing the IgM daily preserved protection compared with protection from IgM from other hybridomas or saline only. The decrease in the specific activity of plaque proteins caused by replenishing one such antibody (3hE5) was 2.5-fold compared with activity with unreplenished 3hE5, 3.8-fold compared with activity with saline only, and 10.7-fold compared with activity with replenished, unrelated antibody. The neutralizing IgM detected an array of 14,000- to 22,000-molecular-weight antigens. The native toxins may be aggregates of these antigens, or the array may indicate fragments of an undetected, larger antigen or a common, nonpeptide adduct. Only 0.5 to 0.8% of the bacteria from sites with periodontitis and grown on blood agar contained antigen. One group of reactive bacteria was identified as Actinomyces odontolyticus serotype I. Other isolates were identified as Staphylococcus epidermidis, but antigen disappeared from the these isolates within 6 weeks of subculture. Epitope-containing antigens were also found in streptococcal and Eikenella isolates, and it is likely that the antigens from only some of these bacteria are toxic.     

Passive immunization with milk produced from an immunized cow prevents oral recolonization by Streptococcus mutans.

Cell surface protein antigen (PAc) and water-insoluble glucan-synthesizing enzyme (GTF-I) produced by cariogenic Streptococcus mutans are two major factors implicated in the colonization of the human oral cavity by this bacterium. We examined the effect of bovine milk, produced after immunization with a fusion protein of functional domains of these proteins, on the recolonization of S. mutans. To prepare immune milk, a pregnant Holstein cow was immunized with the fusion protein PAcA-GB, a fusion of the saliva-binding alanine-rich region (PAcA) of PAc and the glucan-binding (GB) domain of GTF-I. After eight adult subjects received cetylpyridinium chloride (CPC) treatment, one subgroup (n = 4) rinsed their mouths with immune milk and a control group (n = 4) rinsed with nonimmune milk. S. mutans levels in saliva and dental plaque decreased after CPC treatment in both groups. Mouth rinsing with immune milk significantly inhibited recolonization of S. mutans in saliva and plaque. On the other hand, the numbers of S. mutans cells in saliva and plaque in the control group increased immediately after the CPC treatment and surpassed the baseline level 42 and 28 days, respectively, after the CPC treatment. The ratios of S. mutans to total streptococci in saliva and plaque in the group that received immune milk were lower than those in the control group. These results suggest that milk produced from immunized cow may be useful for controlling S. mutans in the human oral cavity.     

Examination of human stomach biopsies, saliva, and dental plaque for Campylobacter pylori.

To examine possible sources of Campylobacter pylori and to determine the routes by which it is transmitted to the human stomach, samples of dental plaque and saliva from 71 patients undergoing endoscopy in addition to stomach biopsies were collected and cultured on selective noninhibitory Skirrow medium. A total of 29 (40.8%) of the stomach biopsies yielded C. pylori. None of the saliva samples and only one of the dental plaque samples was found positive for C. pylori, and thus neither saliva nor dental plaque could be implicated as a significant reservoir of this organism.     

Survival of Bacteria from Human Dental Plaque Under Various Transport Conditions

The effects of transport media, temperature, and anaerobiosis on the survival of bacteria from human supragingival dental plaque were studied. Individual samples were obtained by passing sterile, unwaxed dental floss through the interproximal spaces. The plaque-bearing portion of floss was immediately placed in vials containing reduced transport fluid, viability-preserving microbistatic medium, or reduced salt solution transport fluid. Plaque samples were dispersed by ultrasonic oscillation, serially diluted, and plated in duplicate on MM10-sucrose-blood agar, mitis salivarius bacitracin agar, and Rogosa tomato juice agar. Initial viable counts (time 0) were compared with viable count determinations after 48- and 72-h storage. Quantitative recovery (>30%) of various groups of oral bacteria was accomplished from both reduced transport fluid and viability-preserving microbistatic medium after 48- and 72-h storage. Storage of dental plaque in reduced salt solution proved unsatisfactory for most bacteria (less than 10% survival). Since growth of some bacteria may occur in viability-preserving microbistatic medium and the charcoal present interferes with colonly enumeration on low-dilution plates, we found reduced transport fluid to be the most suitable medium for transport and recovery of bacteria from supragingival dental plaque. Subzero storage (-196 and -40 degrees C) did not enhance the survival of bacteria from dental plaque; storage at moderate (5 and 20 degrees C) temperatures gave better recovery of viable bacteria. Survival after anaerobic or aerobic storage was comparable for total colony-forming units; however, anaerobic storage enhanced survival of Streptococcus mutans and Lactobacillus. Since these organisms are specifically associated with dental caries, anaerobic techniques are preferred for caries activity testing of plaque.     

Age-related microbiological changes in the salivary and plaque microflora of healthy adults

The effect of age on quantitative or qualitative differences in selected bacteria of dental significance and on the carriage of opportunistic pathogens and transient oral species was determined in 79 healthy, non-denture wearing individuals divided into four age groups: 20-39 years (group A), 40-59 years (group B), 60-79 years (group C) and greater than or equal to 80 years (group D). Samples of dental plaque and whole saliva were cultured on appropriate selective and non-selective bacteriological media. The total numbers of viable bacteria in saliva, and the prevalence of mutans streptococci in plaque and saliva were similar in all age groups. Similarly, there was no correlation between the numbers of spirochaetes in plaque and age. In contrast, statistically significantly higher mean proportions (p = 0.004), mean log10 viable counts (p = 0.001) and isolation frequencies (p less than 0.01) of lactobacilli were found in the saliva of those aged greater than or equal to 70 years compared to subjects in group A. The isolation frequency (p less than 0.05) and proportions (p = 0.056) of staphylococci in saliva were also higher in those aged greater than or equal to 70 years. Yeasts were isolated most often and in higher numbers from saliva in those aged greater than or equal to 80 years and the proportion of yeasts was higher after 60 years of age, but these differences were not significant in comparison with results from individuals in group A.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enumeration of mutans streptococci in clinical samples by using monoclonal antibodies.

Mutans streptococci have been strongly associated with dental caries. Two members of this group of bacteria, Streptococcus mutans and Streptococcus sobrinus, are often found in human dental plaque. Identification of mutans streptococci on the basis of sugar fermentation is troublesome and easily leads to erroneous conclusions. Furthermore, the recovery on selective media differs for different species. This causes incorrect enumeration of S. mutans and S. sobrinus in clinical samples. The aim of this study was to develop a method for simultaneous identification and enumeration of S. mutans and S. sobrinus in dental plaque and saliva samples. With this immunoblot technique (IBT), significantly more plaque samples containing S. sobrinus were detected than on the selective medium Trypticase-yeast-cysteine-sucrose-bacitracin agar (TYCSB) (P less than 0.01). The numbers of plaque samples harboring S. mutans were equal on TYCSB and by IBT. However, the numbers of CFU of S.mutans as well as of S. sobrinus detected with the IBT were significantly higher than those obtained on TYCSB (P less than 0.001). The recovery of primary isolations of S. sobrinus on TYCSB seems to have been inhibited in 26 of the 45 S. sobrinus-containing plaque samples. False-positive or false-negative reactions with the IBT were not found.     

In Vitro Attachment of Streptococci to the Tooth Surface

The ability of Streptococcus strains to adhere to the tooth surface in vitro was investigated. Polished enamel slabs, with and without acquired pellicles, were incubated with buffer suspensions of oral streptococci, and attached bacteria were counted under a microscope using incident light. Low numbers of bacteria adhered to uncoated enamel; the presence of an acquired pellicle significantly enhanced the attachment of all strains tested. The adherence of Streptococcus sanguis was significantly greater than that of Streptococcus salivarius, and both of these strains adhered in greater numbers than did Streptococcus mutans. When bacteria were suspended in whole saliva, the adherence of S. salivarius and S. mutans was inhibited, whereas the adherence of S. sanguis was enhanced in some experiments and inhibited in others. The adherence of S. sanguis and S. salivarius was consistently inhibited by parotid fluid; this inhibitory effect persisted after thorough washing and resonication of the bacterial cells. Incubation in oral fluids was associated with the attachment of bacterial clumps to the pellicle, and parallel investigation revealed agglutination of S. sanguis and S. salivarius by whole saliva and, in particular, parotid fluid. The results are discussed in terms of surface microecology, and are related to the development of dental plaque.     

Passage of intact IgG from plasma to the oral cavity via crevicular fluid

The aim of this investigation was to determine whether IgG could pass from the blood to the oral cavity. Pure IgG was prepared from monkey serum, by ion exchange chromatography and gel filtration, and was radiolabelled with ¹²⁵I. This was injected intravenously into eight Rhesus monkeys. Radioactivity could be detected in crevicular fluid washings, and in mixed and parotid saliva samples 30 min after injection. Ultracentrifugation on sucrose density gradients revealed that most of the radioactivity in crevicular fluid washings was associated with proteins having a sedimentation coefficient similar to marker IgG. Radioactivity in parotid saliva was not found in the IgG zone, but was present in zones with sedimentation coefficients of approximately 4·5S and 1S. The results suggest that IgG passes as an intact molecule from plasma to crevicular fluid, and support the hypothesis that serum antibodies could play a role in protection against dental caries.     

The normal gastric microflora and Helicobacter pylori; before, during and after treatment with omeprazole and amoxycillin

Objective: To study the microflora in patients with Helicobacter pylori infection during treatment with omeprazole alone and in combination with amoxycillin, to study transmission of relapsing H. pylori strains by fingerprinting and to investigate if H. pylori is detectable in saliva and dental plaque by polymerase chain reaction (PCR).
Methods: Twenty-eight dyspeptic patients with H. pylori infection were divided into two treatment groups omeprazole 20 mg plus amoxycillin 1000 mg, or omeprazole 20 mg plus placebo twice a day for 14 days. Biopsies were taken before, during and after treatment. The biopsies were cultivated in order to study the commensal microflora and H. pylori PCR was used to detect H. pylori in the biopsies, saliva and dental plaque. The H. pylori strains were fingerprinted with arbitrary primed PCR.
Results: Twenty-five patients harbored H. pylori , of whom 22 also harbored a normal mucosal microflora. H. pylori was present in all patients in the omeprazole-placebo group and in 39% of the patients in the omeprazole-amoxycillin group 4 weeks after treatment. There were no significant differences in the number of bacteria in antrum and corpus.
Conclusions: There was an inverse relationship between the growth of commensal microflora and H. pylori during treatment in both groups. H. pylori was detectable in saliva and dental plaque by PCR. The original H. pylori strain was present in all but one relapsing patient.     

Passive Immunization with Milk Produced from an Immunized Cow Prevents Oral Recolonization by Streptococcus mutans

Cell surface protein antigen (PAc) and water-insoluble glucan-synthesizing enzyme (GTF-I) produced by cariogenic Streptococcus mutans are two major factors implicated in the colonization of the human oral cavity by this bacterium. We examined the effect of bovine milk, produced after immunization with a fusion protein of functional domains of these proteins, on the recolonization of S. mutans. To prepare immune milk, a pregnant Holstein cow was immunized with the fusion protein PAcA-GB, a fusion of the saliva-binding alanine-rich region (PAcA) of PAc and the glucan-binding (GB) domain of GTF-I. After eight adult subjects received cetylpyridinium chloride (CPC) treatment, one subgroup (n = 4) rinsed their mouths with immune milk and a control group (n = 4) rinsed with nonimmune milk. S. mutans levels in saliva and dental plaque decreased after CPC treatment in both groups. Mouth rinsing with immune milk significantly inhibited recolonization of S. mutans in saliva and plaque. On the other hand, the numbers of S. mutans cells in saliva and plaque in the control group increased immediately after the CPC treatment and surpassed the baseline level 42 and 28 days, respectively, after the CPC treatment. The ratios of S. mutans to total streptococci in saliva and plaque in the group that received immune milk were lower than those in the control group. These results suggest that milk produced from immunized cow may be useful for controlling S. mutans in the human oral cavity.     

Vascular endothelial growth factor is constitutively expressed in normal human salivary glands and is secreted in the saliva of healthy individuals

The expression of vascular endothelial growth factor (VEGF), a specific mitogen for endothelial cells, was examined in salivary glands and in normal saliva. In normal salivary glands, VEGF mRNA and protein were strongly present in acinar cells, whereas little or no VEGF was found in ductal cells. In chronically inflamed glands, VEGF protein was in addition present in ductal elements and in infiltrating mononuclear cells. No difference of VEGF expression was observed between benign and malignant salivary gland tumours. By ELISA, whole saliva of 24 healthy individuals contained up to 2·5 ng/ml (mean 1·4 ng/ml; SD 0·77 ng/ml) of VEGF, confirming the constitutive secretion of this cytokine by human salivary glands. Western blot analysis of normal saliva under non-reducing conditions detected anti-VEGF reactive protein moieties of ≈46 kD, corresponding to VEGF secreted by cells in tissue culture. Additional anti-VEGF reactive proteins of ≈60 and 90 kD were detected in the saliva of some individuals. The presence of considerable quantities of VEGF in normal human saliva suggests an important role for this cytokine in the maintenance of the homeostasis of mucous membranes, with rapid induction of neoangiogenesis by salivary VEGF helping to accelerate wound healing within the oral cavity. Moreover, salivary VEGF may permeabilize intraglandular capillaries and thus participate in the regulation of saliva production itself. Copyright © 1998 John Wiley & Sons, Ltd.     

Epithelial cell response to challenge of bacterial lipoteichoic acids and lipopolysaccharides in vitro

Accumulating dental plaque at the gingival margin contains lipoteichoic acids (LTAs) from the cell walls of gram-positive bacteria. In subgingival plaque associated with periodontal disease the amount of lipopolysaccharides (LPSs) from gram-negative bacteria increases. As the gingival junctional epithelium (JE) is an important structural and functional tissue, participating in the first line defence against apical advancement of dental plaque, this study examined the direct effects of LTAs (from Streptococcus mutans and S. sanguis) and LPSs (from Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola and Escherichia coli) on two epithelial cell lines (HaCaT and ERM) and a culture model for human JE. The cells were exposed to the LTAs or LPSs (10-50 microg/ml) for variable periods of time. None of the bacterial surface components had any effect on primary adhesion or on the epithelial attachment of the JE cultures. However, cell growth and mitotic activity were consistently reduced in all cultures treated with LTAs. In contrast, LPSs showed only slight or no effects on cell growth and mitotic activity depending on the epithelial cells used. This suggests that LPSs, despite their established role as modulators of inflammation, do not have direct harmful effects - at the concentrations found in dental plaque and gingival crevicular fluid - which would explain the mechanism of epithelial degeneration and detachment from the tooth surface. However, the LTAs appear to inhibit the renewal of epithelium and may thus contribute to degeneration of coronal JE and subgingival colonisation by periodontal pathogens.     

The effect of orthodontic bonding materials on dental plaque accumulation and composition in vitro

The aim of this study was to investigate the accumulation and composition of microcosm dental plaque on different orthodontic bonding materials using an in vitro model. Microcosm plaques were grown on discs of a range of bonding materials in a constant depth film fermentor. The biofilms were derived from human saliva and supplied with artificial saliva as a source of nutrients. The number of viable bacteria in the biofilms was determined and the streptococci present were identified to species level. The results showed that there was no significant difference in bacterial accumulation between different bonding materials, however, biofilms grown on materials which were fluoride releasing, did not contain Streptococcus mutans. This in vitro study has shown that the use of fluoride-releasing bonding materials may support the growth of supragingival plaque, which does not contain S. mutans.     

Phosphorylcholine-dependent cross-reactivity between dental plaque bacteria and oxidized low-density lipoproteins

Antibodies reactive with phosphorylcholine (PC) are ubiquitous in human sera, but the antigens stimulating their production and their function are not clear. Previous studies have shown that a significant proportion of dental plaque bacteria contain PC as determined by reactivity with PC-specific mouse myeloma proteins and monoclonal antibodies. Additionally, serum antibody concentrations of immunoglobulin (IgG) G anti-PC are higher in sera of individuals who have experienced periodontal attachment loss than those who are periodontally healthy. These data implicate the oral microflora as a source of antigen-stimulating anti-PC responses. Recent data also indicate that antibodies with specificity for PC are elevated in ApoE-deficient mice, a model for studies of athersclerosis, and that such antibodies bound oxidized low-density lipoproteins (LDL) (oxLDL) in atherosclerotic plaques. These data prompted the hypothesis that human anti-PC could bind to both oral bacteria and human oxLDL, and that these antigens are cross-reactive. We therefore examined the ability of human anti-PC to bind to PC-bearing strains of oral bacteria using enzyme-linked immunosorbent inhibition assays and by assessment of direct binding of affinity-purified human anti-PC to PC-bearing Actinobacillus actinomycetemcomitans. Our results indicated that PC-bearing strains of Streptococcus oralis, Streptococcus sanguis, Haemophilus aphrophilus, Actinomyces naeslundii, Fusobacterium nucleatum, and A. actinomycetemcomitans, as well as a strain of Streptococcus pneumoniae, absorbed up to 80% of anti-PC IgG antibody from human sera. Furthermore, purified anti-PC bound to a PC-bearing strain of A. actinomycetemcomitans but only poorly to a PC-negative strain. OxLDL also absorbed anti-PC from human sera, and oxLDL but not LDL reacted with up to 80% of the anti-PC in human sera. Furthermore, purified anti-PC bound directly to oxLDL but not to LDL. The data indicate that PC-containing antigens on a variety of common oral bacteria are cross-reactive with neoantigens expressed in oxLDL. We propose that PC-bearing dental plaque microorganisms may induce an antibody response to PC that could influence the inflammatory response associated with atherosclerosis.