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PubMed1983

Effective immunity to dental caries: enhancement of salivary anti-Streptococcus mutans antibody responses with oral adjuvants.

In the present study, we compared the ability of the soluble adjuvants concanavalin A (ConA), muramyl dipeptide (MDP), and peptidoglycan (PG) to enhance immune responses to orally administered particulate antigens of Streptococcus mutans 6715 in gnotobiotic rats. The isotype and levels of antibody in saliva and in serum from experimental rats were determined by an enzyme-linked immunosorbent assay using S. mutans whole cells (WC) as the coating antigen. The specificities of salivary and serum immunoglobulin A (IgA) antibodies to particulate S. mutans antigens, lipoteichoic acid, S. mutans serotype g carbohydrate, and dextran were also determined. When 50 micrograms of ConA was used as the oral adjuvant with S. mutans 6715 WC immunogen, a slight enhancement of immune responses was obtained. A higher dose of ConA suppressed humoral responses to the immunogen. Enhanced immune responses, especially of the IgA isotype, in both serum and saliva were induced in gnotobiotic rats given MDP and either S. mutans 6715 WC or purified cell walls (CW) by gastric intubation. Elevated IgA antibody levels to CW, lipoteichoic acid, and carbohydrate were observed in rats given S. mutans WC and MDP by gastric intubation, whereas oral immunization with S. mutans CW and MDP resulted in higher antibody levels to CW and carbohydrate and lower levels to lipoteichoic acid when compared with the antibody levels in rats given antigen alone. Rats orally immunized with either S. mutans WC or CW and MDP and challenged with virulent S. mutans 6715 exhibited significantly (P less than or equal to 0.05) lower plaque scores, numbers of viable S. mutans in plaque, and caries scores than did rats immunized with antigen alone or in infected-only controls. In another series of experiments, a PG fraction derived from S. mutans 6715 CW was assessed for adjuvant properties. The oral administration of PG and either S. mutans WC or CW induced good salivary and serum IgA antibody responses. The specificity of the antibodies was similar to that obtained in rats given antigen and MDP. Rats receiving either S. mutans WC or CW and PG and challenged with virulent S. mutans 6715 had lower plaque scores, fewer numbers of viable S. mutans in plaque, and lower caries activity than did infected rats receiving S. mutans WC or CW immunogen alone. These results provide evidence that soluble adjuvants derived from the gram-positive bacterial CW, e.g., MDP and PG, are effective oral adjuvants and augment IgA immune responses to particulate S. mutans antigens which are protective against the mucosally associated disease, dental caries.

Adjuvants, Immunologic↗
PubMed1999

The effect of extracellular polysaccharides from Streptococcus mutans on the bactericidal activity of human neutrophils.

Extracellular polysaccharides (PS) synthesized by oral bacteria constitute one of their major virulence factors. The PS, synthesized from sucrose, facilitate adhesion and colonization by bacteria to tooth surfaces. The study was designed to test the effect of in situ production of extracellular PS by Streptococcus mutans on the bactericidal activity of human neutrophils. These effects were tested on bacteria pre-exposed to sucrose (PS-positive Strep. mutans) and compared to bacteria not exposed to sucrose (PS-negative Strep. mutans). The interactions between neutrophils and Strep. mutans were tested in suspension and on bacteria in an experimental model of dental plaque. Viability of Strep. mutans was measured by [3H]-thymidine incorporation into the bacteria. Degranulation of neutrophils was evaluated by the release of lysozyme, and the production of reactive oxygen products was measured by chemiluminescence. When neutrophils were incubated with suspended bacteria, the viability of PS-negative Strep. mutans was 20% of that of bacteria not incubated with neutrophils (control), while the viability of PS-positive Strep. mutans was 40% of the control. In the experimental dental-plaque model, 50% of the PS-negative Strep. mutans were killed by neutrophils while the viability of PS-positive Strep. mutans was not different than of the control. Degranulation of neutrophils was not affected by the presence of extracellular PS of Strep. mutans. Artificial stimulation of neutrophils with phorbol myristate acetate also did not enhance the bactericidal effect of neutrophils on PS-positive Strep. mutans. However, PS-positive Strep. mutans elicited oxygen-reactive products from neutrophils, 2-fold less than with PS-negative Strep. mutans. The results indicate that in situ production of bacterial extracellular polysaccharides might be a major virulence factor of Strep. mutans, enabling PS-positive Strep. mutans in the dental-plaque biofilm to evade killing by human neutrophils.

Anti-Bacterial Agents↗
PubMed1990

[Detection of mutans streptococci by latex agglutination test and its application as a caries-activity test].

The number of mutans streptococci in saliva and dental plaque has been reported to correlate with the incidence of dental caries. This report describes a simple and rapid diagnostic method for the detection of mutans streptococci in dental plaque using latex agglutination (LA) test. Latex particles (0.876 microns, diameter) were sensitized with partially purified antibodies against whole cells of Streptococcus mutans MT8148 (c), MT703R (e) and OMZ175 (f) and Streptococcus sobrinus B13 (d) and 6715 (g). Whole cells of mutans streptococci or dental plaque was extracted with a mixture of 8M sodium nitrite (5 microliters) and 2M acetic acid (5 microliters) for five minutes and neutralized with 2M sodium hydroxide (10 microliters), and the extract and the sensitized latex suspension (20 microliters) were mixed and the grade of agglutination reaction was read macroscopically after ten minutes standing at room temperature. The LA tests could detect 1.0 10ng of purified serotype antigen and 10(5)-10(6) CFU of live cells of mutans streptococci and specifically distinguish not only the mutans streptococci from the other streptococci but also S. mutans from S. sobrinus. However, cross-reactions were still observed among the serotypes c, e and f of S. mutans or between the serotypes d and g of S. sobrinus. Plaque samples were collected from 168 children (2 to 12 years of age) and the 0.1 mg (wet weight) were applied to the LA tests. At the same time, the total number of mutans streptococci in plaque and the serotypes of each isolate were determined. The results of LA reaction correlated significantly with the number of mutans streptococci in plaque (chi-square analysis; p less than 0.0001). The LA tests discriminated between S. mutans and S. sobrinus in dental plaque. It was found that the latex particles sensitized with anti-serotype c and/or e S. mutans antibodies were most effective in demonstrating mutans streptococci, and they were used in the following studies. The results of LA reaction correlated significantly with both the number of decayed tooth surfaces and the number of decayed and filled tooth surfaces. These results suggest that the LA test in mutans streptococci can be useful for the detection of mutans streptococci in dental plaque and also as a caries-activity test at dental clinics.

Chi-Square Distribution↗
PubMed2000

A role for Lewis a antigens on salivary agglutinin in binding to Streptococcus mutans.

Streptococcus mutans is a major etiological agent in dental caries. Salivary agglutinin is one of the main salivary components binding to S. mutans. To learn more about the interaction of salivary agglutinin with S. mutans, parotid, submandibular, sublingual and palatal saliva samples were incubated with S. mutans suspension. Both depleted saliva samples and bacterial extracts were analyzed by SDS-PAGE and immunoblotting. Salivary agglutinin was present in all types of glandular saliva and in all cases bound to S. mutans, also to PC337C, a P1 mutant of S. mutans. Agglutinin was separated by SDS-PAGE under reducing and non-reducing conditions and then transferred to nitrocellulose. Non-reduced agglutinin bound S. mutans, but reduced agglutinin did not. Adhesion of S. mutans to agglutinin-coated microplates was inhibited by amine-containing components, 1 M NaCl or KCl and EDTA. Adhesion decreased with decreasing pH with no adhesion below pH 5.0. These data suggest that calcium-dependent electrostatic interactions play a role in binding. By immunoblotting was demonstrated that blood group antigens and Lewis antigens were present on agglutinin. Synthetic blood group antigens and Lewis antigens covalently coupled to polyacrylamide were tested for binding to S. mutans. Only Le(a)(Gal beta 1,3(Fuc alpha 1,4)GlcNAc) bound to S. mutans, whereas the blood group antigens Le(b), Le(x), Le(y), H1, H2, A, B and sialylated Le(a) did not. Lea without galactose (Fuc alpha 1,4GlcNAc) still bound to S. mutans, but Le(a) without fucose (Gal beta 1,3GlcNAc) did not. Binding of agglutinin to S. mutans was not inhibited by Le(a). In conclusion, S. mutans can bind to Le(a) carbohydrate epitopes in which the fucose is an essential residue. Le(a) carbohydrate epitopes are present on salivary agglutinin but play no major role in binding.

Agglutinins↗
PubMed1996

Cariogenicity depends more on diet than the prevailing mutans streptococcal species.

This review aims to compare the occurrence and distribution of mutans streptococci in Africa, Europe, and North America and in addition will try to offer explanations for existing relationships among salivary mutans streptococci counts, dietary patterns, and dental caries. The literature reveals that salivary mutans streptococci counts in child populations of the three continents are comparable. The distribution of mutans streptococci species, with a predominance of S. mutans followed by S. sobrinus, and the virtual absence of other mutans streptococci species are also comparable. Although it is widely believed that diet has an important effect on mutans streptococci counts, this review provides evidence that this does not hold true when variations in dietary patterns are moderate, as they normally are in real-life situations. Since the diets of the child populations in the three continents vary moderately, a strong dietary-induced effect on salivary mutans streptococci counts cannot be expected. The observed analogous salivary mutans streptococci counts in these child populations are thus 'not surprising' but are in accordance with the conceptual expectation. The differences in caries experience in children of the three continents cannot be explained by the prevailing mutans streptococci species but instead should be attributed to differences in the cariogenicity of the various diets. The fact that the cariogenicity of the diet determines the development of dental caries while hardly affecting the mutans streptococci counts explains the limited value of the latter as an indicator of dental caries. The reviewed literature shows that mutans streptococci are ubiquitous in children aged 7 years and older in Africa, Europe, and North America. Mutans streptococci should therefore be considered as belonging to the indigenous microflora of the human mouth.

Adolescent↗
PubMed1998

Association of salivary immunoglobulin A antibody and initial mutans streptococcal infection.

We explored the relationship between mutans streptococcal infection and the development of salivary IgA antibody during initial colonization. Repetitive swabbing (n = 292) of the teeth of 33 children revealed that 45% became infected with mutans streptococci between 13 and 36 months of age. In contrast, mutans streptococci could not be detected in 18 children whose last sample was taken at 39-81 months of age (median age = 62 months). During the period of mutans streptococcal infectivity, immunoglobulin A (IgA) antibody to several mutans streptococcal antigens appeared in most children, whether or not infection had been demonstrated. Robust responses to mutans streptococcal components occurred during or shortly after, but not before the period of mutans streptococcal infectivity. No consistent differences were observed among the summarized patterns of response of infected and uninfected groups of children, although the IgA Western blot patterns of individual subjects were often quite distinct. For example, sets of siblings, who would be presumed to be challenged with similar maternal mutans streptococcal clonotypes, were shown to develop qualitatively different salivary IgA responses to mutans streptococcal components. These results support a discrete period for mutans streptococcal infection and may suggest that the level of maternal infection is a factor in the success of infection of the child during this period. The data also suggest that exposure to mutans streptococci is a sufficient condition for robust mucosal IgA responses to mutans streptococcal antigens during the period of infectivity and that these responses may be different, even among siblings.

Antibodies, Bacterial↗
PubMed1988

Suppression of Streptococcus sobrinus 6715 (g) in plaques by Streptococcus mutans 32K (c).

The dental plaque of 96 healthy donors was screened for the production of such antibacterial substances as mutalipocin and bacteriocin and 192 strains of mutans streptococci isolated: 28 produced mutalipocin and 22 produced bacteriocin. Mutalipocin produced by these 28 S. mutans strains possessed similar biochemical and biological characteristics of well-characterized mutalipocin-producing strain S. mutans 32K (serotype c). When equal amounts of S. mutans 32K and S. sobrinus 6715 (g) were cultured together, cells of S. sobrinus 6715 were completely killed in 18 h. In addition, S. mutans 32K inhibited in vitro plaque formation by S. sobrinus 6715, and S. mutans 32K also eliminated in vitro plaque preformed by S. sobrinus 6715. In rat experiments, S. mutans 32K could pre-emptively colonize in plaque preformed by S. sobrinus 6715. On the other hand, S. sobrinus 6715 could not colonize in plaque preformed by S. mutans 32K. The results indicate that S. mutans serotype c which produces antibacterial substances is able to invade dental plaque and replace the other mutans streptococci. This investigation offers one of the possible explanation why S. mutans serotype c is a predominant species among mutans streptococci in human plaque.

Animals↗
PubMed2001

Mutans streptococci oral colonization in 12-30-month-old Brazilian children over a one-year follow-up period.

OBJECTIVES: The infection levels of mutans streptococci were investigated during a one-year follow-up in children aged 12 to 30 months attending school nurseries where a sucrose-rich diet was provided. METHODS: Oral levels of mutans streptococci obtained from 101 children at baseline and after a one-year follow-up were compared by age, number of teeth, feeding habits, and presence of visible plaque at baseline. Baseline predictors and changes in mutans streptococci levels during the study were compared to caries incidence after one year. RESULTS: Fluctuations in mutans streptococci levels during the follow-up period were not related to feeding habits or presence of visible plaque. Mutans streptococci levels increased after one year among children aged 12 to 24 months, while a significantly higher proportion of those aged 25-30 months showed a decrease in mutans streptococci levels during the study. Multiple logistic regression analysis suggested that high levels of mutans streptococci (> or = 100 cfu) at baseline were associated with a higher caries increment, while reduction in mutans streptococci was negatively associated with caries incidence. CONCLUSIONS: Our data suggest that despite early mutans streptococci infection and high exposure to sucrose, mutans streptococci may achieve relatively stable levels after 2 years of age. Heavy colonization by mutans streptococci in an early age was related to an extremely high caries incidence during childhood, while decreasing levels of mutans streptococci can be associated to the decrease in caries activity.

Age Factors↗
PubMed2001

Passive transfer of immunoglobulin Y antibody to Streptococcus mutans glucan binding protein B can confer protection against experimental dental caries.

Active immunization with Streptococcus mutans glucan binding protein B (GBP-B) has been shown to induce protection against experimental dental caries. This protection presumably results from continuous secretion of salivary antibody to GBP-B, which inhibits accumulation of S. mutans within the oral biofilm. The purpose of this study was to explore the influence of short-term (9- or 24-day) passive oral administration of antibody to S. mutans GBP-B on the longer-term accumulation and cariogenicity of S. mutans in a rat model of dental caries. Preimmune chicken egg yolk immunoglobulin Y (IgY) or IgY antibody to S. mutans GBP-B was supplied in lower (experiment 1) and higher (experiment 2) concentrations in the diet and drinking water of rats for 9 (experiment 1) or 24 (experiment 2) days. During the first 3 days of IgY feeding, all animals were challenged with 5 x 10(6) streptomycin-resistant S. mutans strain SJ-r organisms. Rats remained infected with S. mutans for 78 days, during which rat molars were sampled for the accumulation of S. mutans SJ-r bacteria and total streptococci. Geometric mean levels of S. mutans SJ-r accumulation on molar surfaces were significantly lower in antibody-treated rats on days 16 and 78 of experiment 2 and were lower on all but the initial (day 5) swabbing occasions in both experiments. Relative to controls, the extent of molar dental caries measured on day 78 was also significantly decreased. The decrease in molar caries correlated with the amount and duration of antibody administration. This is the first demonstration that passive antibody to S. mutans GBP-B can have a protective effect against cariogenic S. mutans infection and disease. Furthermore, this decrease in infection and disease did not require continuous antibody administration for the duration of the infection period. This study also indicates that antibody to components putatively involved only in cellular aggregation can have a significant effect on the incorporation of mutans streptococci in dental biofilm.

Animals↗
PubMed1999

Acid-induced acid tolerance and acidogenicity of non-mutans streptococci.

Acid tolerance and acidogenicity of non-mutans streptococci and their capacity of acid adaptation were studied. The cells of non-mutans streptococci (Streptococcus sanguis [Streptococcus sanguinis], Streptococcus gordonii, Streptococcus oralis and Streptococcus mitis) grown at pH 7.0 showed 0.0088% to 71% viability after acidification at pH 4.0 for 60 min, whereas the cells of mutans streptococci (Streptococcus mutans) were not killed by the acidification. Washed cells of non-mutans streptococci lowered pH to 4.04-4.33 in the presence of glucose, while mutans streptococci cells lowered pH to 3.70. When the growth pH was shifted to 5.5 for 30-90 min, the viability of non-mutans streptococci after the acidification at pH 4.0 for 60 min increased (0.25% to 91%) and the minimum pH values of the cells in the presence of glucose decreased (3.90 4.19). Along with the increase in acid tolerance and acidogenicity, non-mutans streptococci increased activities of H(+)-ATPase and arginine deiminase and amounts of stress proteins cross-reacting with 60 kDa and 70 kDa heat shock proteins (Hsp60 and Hsp70). These results indicate that non-mutans streptococci were capable of increasing their acid tolerance and acidogenicity in response to environmental acidification. Furthermore, it is suggested that the acid adaptation observed in non-mutans streptococci cells could involve the induction of H(+)-ATPase, arginine deiminase and stress protein syntheses. The strains of non-mutans streptococci, which are pioneer bacteria for dental plaque formation and predominant in plaque microbial flora, may play a significant role in shifting the dental plaque environment toward acidic and consequently promoting the colonization of more acid-tolerant and acidogenic bacteria such as mutans streptococci and lactobacilli.

Acids↗
PubMed1983

An enzyme-linked immunosorbent assay (ELISA) for quantification of antibodies to Streptococcus mutans surface antigens.

An ELISA was developed to quantitate the level of antibodies to various cell surface antigens of the Gram positive bacterium, Streptococcus mutans. Whole cells and purified cell wall components of S. mutans, lipoteichoic acid (LTA) from Streptococcus pyogenes, and dextran T 2000 were employed as coating antigens in this study. Cell walls of S. mutans were purified by mechanical disruption of whole cells followed by differential centrifugation and proteolytic enzyme treatment. Serotype-specific carbohydrate was purified from an autoclaved, lyophilized S. mutans whole cell preparation by column chromatography. LTA was prepared by Sepharose 4B chromatography of a phenol-water extract of S. pyogenes and used for detection of anti-polyglycerophosphate (PGP) antibodies. A rabbit antiserum to S. mutans 6715 (serotype g), which precipitated with purified carbohydrate antigen (RR g), gave good reactions with purified cell walls and whole cells of S. mutans 6715, less activity with RR g and low activity to LTA and dextran when tested by ELISA. Adsorption of this antiserum with whole cells of S. pyogenes resulted in antibody activity with specificity only to the serotype carbohydrate. The specificity of the antibody for homologous coating antigen was RR g greater than cell wall greater than whole cells. An antiserum to S. mutans MT573 (serotype e) contained antibody predominantly to LTA, whereas, anti-S. mutans MT703 (serotype e) reacted with both dextran and LTA; however, the activity to LTA was removed by prior adsorption of the antiserum with S. pyogenes cells. This treatment did not alter the antibody activity to dextran. To establish the sensitivity of ELISA, a purified IgG anti-serotype carbohydrate antibody was prepared by adsorption of anti-S. mutans 6715 antiserum with a mutant of S. mutans which lacks serotype carbohydrate followed by adsorption and elution of specific antibodies from S. mutans 6715 whole cells. The minimum level of sensitivity of ELISA was 12.5 ng of IgG anti-serotype carbohydrate.

Antibodies, Bacterial↗
PubMed2001

Mutans streptococci in subgingival plaque of treated and untreated patients with periodontitis.

BACKGROUND: The etiology of root caries is thought to be identical to coronal caries, though root caries seem to be more complicated because of the higher susceptibility of exposed roots (dentin) by periodontal therapy to demineralization than intact enamel. This implies that mutans streptococci are the most likely pathogens in the development of root caries. Although it is known that both the numbers of mutans streptococci and the frequency of isolation in root caries lesions are negatively correlated with the distance from the gingival margin, the subgingival sulcus has not been considered a possible habitat for mutans streptococci. However, subgingival occurrence of mutans streptococci in both untreated and treated periodontal patients has not been documented well in the literature. OBJECTIVE: To investigate the presence and levels of mutans streptococci in the subgingival plaque of patients (n=154) in different stages of periodontal therapy. The subgingival sulcus may be a possible habitat for mutans streptococci. This localisation of mutans streptococci may be of importance in the development of root caries after periodontal surgery. MATERIALS AND METHODS: In this cross-sectional study, subgingival plaque samples from 154 consecutive adult periodontitis patients were tested for presence and levels of mutans streptococci and putative periodontal pathogens by anaerobic cultures. These patients were divided into 4 groups based on their stage of periodontal treatment: (1) untreated patients; (2) patients after initial periodontal therapy only; (3) patients in the maintenance phase who not underwent periodontal surgery; (4) patients after periodontal surgery. RESULTS: The prevalence of mutans streptococci in the 4 study groups varied from 82% in untreated patients to 94% in maintenance patients. The mean proportion of mutans streptococci was 6.65% in maintenance patients versus 1.86% in untreated patients (p=0.005) and 2.51% in patients after scaling and root planing (p=0.041). CONCLUSIONS: The subgingival area is a microbial habitat for mutans streptococci that may be of importance in the development of root caries in periodontitis patients.

Adult↗
PubMed1991

Caries prevalence and salivary mutans streptococci in 0-2-year-old children of Japan.

The prevalence of caries and the number and species distribution of salivary mutans streptococci in 356 children (aged 0-2 yr old) in Japan were examined twice at an interval of 1 yr. No mutans streptococci were isolated from the predentate children. The detection rate of mutans streptococci and the prevalence of caries increased with age. Mutans streptococci were isolated from 39.9% of the subjects. The majority of isolates were identified as Streptococcus mutans. The concentration of mutans streptococci correlated with the number of erupted teeth (r = 0.339). In addition, the concentration of mutans streptococci was more closely correlated to the caries prevalence of the next year (r = 0.465) than that of the year when the salivary concentration of mutans streptococci was first evaluated (r = 0.368). The children who had no caries but harbored mutans streptococci in the first year showed significantly higher caries prevalence in the next year than did the children with neither caries nor mutans streptococci. These results indicate that the establishment of mutans streptococci is associated with caries initiation in early childhood.

Chi-Square Distribution↗