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PubMed1982

Effect of microbial interaction on the colonization rate of Actinomyces viscosus or Streptococcus mutans in the dental plaque of rats.

The resident oral microflora of conventional Osborne-Mendel rats was challenged with Actinomyces viscosus or Streptococcus mutans strains. The adherence of the inoculated organism to the tooth surface and the subsequent growth were studied by means of viable counts determination. The initial growth rate of S. mutans in conventional rats was lower than in mono-associated gnotobiotic rats (doubling time, td = 5 h versus td = 1.1 h). The delayed start of growth and the low initial growth rate indicated that a competitive interaction between S. mutans and the resident microflora occurred. The initial growth rate of A. viscosus in conventional rats (td = 3.1 h) was approximately the same as that in gnotobiotic rats (td = 2.8 h). The start of growth of A. viscosus was only slightly delayed compared with the start in gnotobiotic rats. These results suggest a neutralistic relationship between A. viscosus and the resident microflora. A. viscosus reached a stationary level about 7 days after inoculation, whereas the S. mutans strains did not reach stationary levels until 2 weeks after inoculation.

Actinomyces↗

PubMed1980

Effects of antibacterial microenvironment on in vitro plaque formation of Streptococcus mutans as observed by scanning electron microscopy.

This study presents in vitro techniques to follow the adsorption and subsequent surface proliferation of Streptococcus mutans on hydroxyapatite surfaces leading to artificial plaque. Inhibition of proliferation as well as extracellular dextrane formation can be achieved by pretreatments of the hydroxyapatite surface with aqueous solutions of sodium ricinoleate. The antibacterial activity is restricted to an antibacterial microenvironment adjacent to the hydroxyapatite surface.

Adsorption↗

PubMed2006

Enhancement of erythrosine-mediated photodynamic therapy of Streptococcus mutans biofilms by light fractionation.

OBJECTIVES: We aimed to increase the bacterial cell killing efficacy of erythrosine-mediated photodynamic therapy (PDT) of Streptococcus mutans biofilms by fractionating the delivered light dose into a series of shorter pulses. METHODS: S. mutans biofilms of 200 microm thickness were grown in a constant-depth film fermenter (CDFF). Biofilms were incubated with 22 microM erythrosine before being irradiated with white light for increasing periods of time. We also used light dose fractionation to deliver the same overall dose of light in a series of shorter pulses separated by dark periods. Bacterial cell killing as a result of each killing protocol was quantified by colony counting. RESULTS: A 2 log10 of bacterial cell killing was achieved with 5 min of continuous white light irradiation. For time periods longer than 5 min the amount of killing increased more slowly, which was probably due to photobleaching of the erythrosine. Fractionation of the light dose into 5 x 1 min doses separated by dark recovery periods of 5 min increased the amount of bacterial killing by 1 log10 compared with 5 min continuous irradiation. Further fractionation of the light dose into 10x 30 s doses separated by 2 min recovery periods resulted in 3.7 log10 of cell kill, an improvement of 1.7 log10 compared with the continuous irradiation protocol. CONCLUSIONS: Erythrosine-mediated PDT of S. mutans biofilms can be further enhanced by fractionation of the applied light dose.

Biofilms↗

PubMed1982

Effects of lysozyme and inorganic anions on the morphology of Streptococcus mutans BHT: electron microscopic examination.

The effects of hen egg white lysozyme and the inorganic salt sodium thiocyanate on the integrity of Streptococcus mutans BHT were studied by transmission electron microscopy. Both control cells and cells exposed to NaSCN possessed thick outer cell walls and densely staining inner cell walls juxtaposed to the plasma membranes. In the presence of NaSCN, however, the S. mutans BHT nucleoid was coagulated into thick electron-dense filaments. Exposure of S. mutans BHT to 150 mug of hen egg white lysozyme per ml resulted in the progressive destruction of both the cell walls and the plasma membranes. The enzyme appeared to affect the region of the cell wall septum, and exposure to 150 mug of hen egg white lysozyme per ml for as short a time as 10 min resulted in visible morphological cell wall alterations. At 30 min, ultrastructural observations revealed that the majority of the cells were in the process of expelling a portion of their cytoplasmic contents from the septal and other regions of the cells at the time of fixation. After 3 h of incubation in the presence of this high lysozyme concentration, gelled protoplasmic masses, which were free from the cells, were evident. In addition, extensive damage to the outer and inner cell walls and to the plasma membranes was apparent, although the cells maintained their shape. On some areas of the cell surface, the outer cell wall and plasma membrane were completely absent, whereas at other locations the outer cell wall was either split away from the inner cell wall and plasma membrane or distended from an area free of inner cell wall and plasma membrane. Upon addition of NaSCN to the hen egg white lysozyme-treated cells, both the gelled protoplasmic masses and the damaged cells exhibited an exploded appearance and existed as membrane ghosts, cell wall fragments, or dense aggregates of cytoplasmic components. The effects of a low lysozyme concentration (22.5 mug/ml) on S. mutans morphology were less pronounced at short incubation times (i.e., 10 and 30 min) than those that were observed with a high enzyme concentration; however, breaks in the cell walls and dissolution of the plasma membranes with resulting cell lysis were visible after a prolonged (3-h) incubation and after subsequent addition of NaSCN.

Cell Membrane↗

PubMed1989

Correlating Streptococcus mutans with dental caries in young children using a clinically applicable microbiological method.

The present study used a clinically applicable microbiological method to correlate Streptococcus mutans counts and dental caries prevalence in young children. The study population consisted of 37 subjects, between 16 and 60 months of age. Using a sterile tongue depressor, samples of unstimulated saliva were obtained from subjects and inoculated onto elevated agar plates containing a selective medium. Plates were incubated anaerobically at 37 degrees C for 48 h. chi 2 and multiple regression analyses gave highly significant relationships (p less than 0.01) between dental caries prevalence and the number of S. mutans colonies.

Child, Preschool↗

PubMed2006

Quantitative detection of Streptococcus mutans in the dental plaque of Japanese preschool children by real-time PCR.

AIMS: To detect quantitatively the total bacteria and Streptococcus mutans in dental plaque by real-time PCR with prbac, Sm and GTF-B primers, and to compare their presence with the prevalence of dental caries in Japanese preschool children. METHODS AND RESULTS: Human dental plaque samples were collected from the labial surfaces of the upper primary central incisors of 107 children. The dental status was recorded as dft by WHO caries diagnostic criteria. Positive dt and dft scores by the Sm or GTF-B primer were significantly higher than negative scores (P < 0.01). The proportions of Strep. mutans to the total bacteria from sound, and sound and/or filled upper primary incisors were significantly lower than those from decayed or filled, and decayed incisors, respectively (P < 0.01). CONCLUSIONS: The ratios of Strep. mutans to total bacteria in plaque detected by real-time PCR with Sm and GTF-B primers were closely associated with the prevalence of dental caries in Japanese preschool children. SIGNIFICANCE AND IMPACT OF THE STUDY: These assays may be useful for the assessment of an individual's risk of dental caries.

Child↗

PubMed1990

Development and establishment of fluoride-resistant strains of Streptococcus mutans in rats.

Fluoride-resistant strains of mutans streptococci have been found to be less cariogenic than their parent strains in the rat model system. Therefore, it is important to determine whether the use of fluorides in vivo will increase the fluoride resistance of mutans streptococci and thereby possibly decrease their cariogenic potential. We studied (1) the fluoride resistance of Streptococcus mutans C180-2 in rat dental plaque exposed to various fluoride treatments (experiment 1), (2) the cariogenic potential of the exposed plaque after transmission to other animals (experiment 2), and (3) whether the fluoride treatments favoured the establishment of in vitro induced fluoride-resistant S. mutans in competition with a fluoride-sensitive strain in rat dental plaque (experiment 3). In all experiments, specific pathogen-free Osborne-Mendel rats were used, and all animals received diet SSP 20/5, containing 20% (w/w) sucrose and 5% (w/w) glucose, and tap water ad libitum. In experiment 1, the rats were inoculated with S. mutans C180-2. For an experimental period of 44 days, the rats either received no F treatment or received NaF mixed in the diet and drinking water (20 or 60 ppm F) or were brushed 6 days per week with an aqueous NaF solution containing either 0.1% (w/v) or 1% (w/v) F, respectively. Bacteriological analyses at the end of the experiment revealed that none of the fluoride treatments had caused a detectable increase in the inherent fluoride resistance of S. mutans C180-2.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗

PubMed1984

Characterization of monoclonal antibodies to Streptococcus mutans antigenic determinants I/II, I, II, and III and their serotype specificities.

Monoclonal antibodies (McAb) were developed to four protein components of Streptococcus mutans serotype c, some of which are significant in the protection against dental caries. The six McAb used in this investigation support the identities of streptococcal antigens (SA) I/II, I, II, and III. The specificities of these antigenic determinants were established both by direct binding and inhibition with the pure SA with a solid-phase radioassay. Whereas conventional antisera to S. mutans serotype c cross-react with serotypes c, e, and f (and g), McAb to serotype c-derived SA I/II react predominantly with serotype c and show some low-titer reactivity with serotype f. The slight cross-reactivity between S. mutants cells of serotypes c and f could be further differentiated by absorption of any of the three McAb to SA I/II with cells of serotype c. Parallel studies of McAb with cells of S. mutans and their ammonium sulfate-precipitated culture supernatants suggest that some SA determinants are retained predominantly on the cell surface, but others are readily shed into the culture medium, so that they are detected both on the cell surface and culture medium. Unlike polyclonal antibodies, McAb are capable of discriminating single antigenic determinants and can be applied to the study of shedding of antigens from microorganisms into the environment, such as the gut or gingival sulcus.

Animals↗

PubMed1986

Role of the cell membrane in pH-dependent fluoride inhibition of glucose uptake by Streptococcus mutans.

The effect of pH on the fluoride sensitivity of glucose uptake by whole cells and glucose transport by permeabilized cells of Streptococcus mutans was compared. Whole cells exhibited a marked pH-dependent sensitivity to fluoride over the pH range 7.0 to 5.0. As the pH was decreased, fluoride sensitivity increased. In contrast, no significant effect of pH on the fluoride sensitivity of glucose transport (e.g., phosphorylation) by permeabilized cells energized with 2-phosphoglycerate was noted. The relative effect of pH on the fluoride sensitivity of whole cell glucose uptake and fermentation was similar. These data are consistent with the notion that the cell membrane is impermeable to the fluoride anion and that intracellular accumulation of fluoride depends on translocation of hydrogen fluoride across the membrane.

Cell Membrane↗

PubMed1979

Inflammatory lesions and bone resorption induced in the rat periodontium by lipoteichoic acid of Streptococcus mutans.

Severe inflammatory lesions were induced in the periodontal tissues of the rat following the intragingival injection of lipoteichoic acid (LTA) from Streptococcus mutans. There was no difference in the severity and distribution of the lesions between nonimmunized rats and animals immunized against LTA after antigenic challenge. The lesions are characterized by the occurrence of granulation tissue, massive infiltration of PMNs, abscess formation, bone resorption, and new bone formation. Deacylated LTA and saline caused relatively mild inflammation, and no significant bone resorption or new bone formation was evident. The peak response was reached after 3 intragingival infections. The mechanisms by which LTA caused the pathological alterations in the rat periodontium and the possible relations of this experimental model to periodontal disease in the human are discussed.

Animals↗

PubMed1979

Determination, by carbon-13 nuclear magnetic resonance spectroscopy, of the composition of glucans synthesized by enzymes of the cariogenic organism Streptococcus mutans.

The 13C-n.m.r. spectra of water-soluble and -insoluble glucans synthesized by enzymes isolated from six strains of Streptococcus mutans are interpreted. The glucans are shown to be composed primarily of alpha-(1 linked to 3)- and alpha-(1 linked to 6)-linked glucosyl residues, and the relative abundance of each linkage is estimated from peak areas. Treatment of water-insoluble glucans with dextranase is found to result in water-soluble and -insoluble products, the former enriched in alpha-(1 linked to 6)-linkages and the latter in alpha-(1 linked to 3)-linkages. The structural conclusions arrived at by 13C-n.m.r. spectroscopy are consistent with data from methylation analysis and 1H-n.m.r. spectroscopy.

Chemical Phenomena↗

PubMed1975

Serological purification of polysaccharide antigens from Streptococcus mutans serotypes a and d: characterization of multiple antigenic determinants.

The polysaccharide antigen preparations from serotype a and serotype d strains of Streptococcus mutans contained both a serotype-specific antigenic determinant and a common a-d antigenic determinant, as demonstrated by agar gel diffusion studies and a quantitative cross-precipitin assay. The chromatographically purified antigens were isolated by a method which depended on their serological specificity to determine if these two antigenic determinants were located on the same molecule. The a and d polysaccharides were recovered from specific antigen-antibody complexes and characterized with respect to their immunological specificity and chemical composition. Agar gel diffusion tests demonstrated that, in both the a and d preparations, the serotype-specific antigenic determinant and the common a-d antigenic determinant were present in one molecule.

Animals↗

PubMed1975

Dextranases from oral bacteria: inhibition of water-insoluble glucan production and adherence to smooth surfaces by Streptococcus mutans.

The effect of dextranases (EC 3.2.1.11) from the oral isolates Actinomyces israelii and Bacteroides ochraceus on water-insoluble glucan production by the Streptococcus mutans dextransucrase (EC 2.4.1.5) and sucrose-dependent adherence to smooth glass surfaces by S. mutans was studied. Collection on membrane filters of water-insoluble polysaccharides synthesized from radioactive sucrose was used to demonstrate the marked sensitivity of insoluble glucan formation to the presence of dextranase. Concentrations of A. israelii dextranase as low as 0.002 U/ml inhibited insoluble glucan formation by 60%. Similar results were obtained the the B. ochraceus enzyme. An assay for sucrose-stimulated adherence of S. mutans to smooth surfaces involved attachment of radioactively labeled nongrowing cells to the bottom of glass scintillation vials. This facile and sensitive assay was utilized to demonstrate that sucrose-dependent adherence was affected by low levels of dextranase from either A. israelii or B. ochraceus. Enzyme at 0.005 U/ml reduced adherence of S. mutans by 80%. Treatment of S. mutans cells previously attached to glass with low concentrations of the dextranases resulted in removal of 50% to 60% of the bacteria. The results indicate that dextranase-producing oral bacteria may affect sucrose-dependent colonization of S. mutans on the tooth surface and offer a possible explanation for both the difficulties involved in implanting this bacterium into the human mouth and the limited intraoral transmission of S. mutans from one tooth surface to another.

Actinomyces↗

PubMed1992

Streptococcus mutans serotype c tagatose 6-phosphate pathway gene cluster.

DNA cloned into Escherichia coli K-12 from a serotype c strain of Streptococcus mutans encodes three enzyme activities for galactose utilization via the tagatose 6-phosphate pathway: galactose 6-phosphate isomerase, tagatose 6-phosphate kinase, and tagatose-1,6-bisphosphate aldolase. The genes coding for the tagatose 6-phosphate pathway were located on a 3.28-kb HindIII DNA fragment. Analysis of the tagatose proteins expressed by recombinant plasmids in minicells was used to determine the sizes of the various gene products. Mutagenesis of these plasmids with transposon Tn5 was used to determine the order of the tagatose genes. Tagatose 6-phosphate isomerase appears to be composed of 14- and 19-kDa subunits. The sizes of the kinase and aldolase were found to be 34 and 36 kDa, respectively. These values correspond to those reported previously for the tagatose pathway enzymes in Staphylococcus aureus and Lactococcus lactis.

Aldehyde-Lyases↗

PubMed1988

A simple purification method for a glucosyltransferase complex from Streptococcus mutans OMZ 176 with a high yield.

1. Glucosyltransferase was purified from Streptococcus mutans OMZ 176 by ammonium sulfate fractionation and gel filtration. 2. The overall purification was about 15,000-fold with a yield of 53%. 3. The purified enzyme appeared to be a multienzyme complex consisting of three different components with molecular weights of 158,000, 153,000 and 149,000, respectively.

Ammonium Sulfate↗

PubMed2004

Peptide mapping of a novel discontinuous epitope of the major surface adhesin from Streptococcus mutans.

Guy's 13 is a mouse monoclonal antibody that specifically recognizes the major cell-surface adhesion protein SA I/II of Streptococcus mutans, one of the major causative agents of dental caries. Passive immunization with Guy's 13 prevents bacterial colonization in humans. To help elucidate the mechanism of prevention of colonization conferred by this antibody, the SA I/II epitope recognized by Guy's 13 was investigated. It was previously established that the epitope is conformational, being assembled from two non-contiguous regions of SA I/II. In the current study, using recombinant fragments of SA I/II and, ultimately, synthetic peptides, the discontinuous epitope was localized to residues 170-218 and 956-969. This work describes the mapping of a novel discontinuous epitope that requires an interaction between each determinant in order for epitope assembly and recognition by antibody to take place. Guy's 13 binds to the assembled epitope but not to these individual epitope fragments. The assembled epitope results from the interaction between the individual antigenic determinants and can be formed by mixing together determinants present on separate polypeptide chains. The data are consistent with one of the epitope fragments adopting a polyproline II-like helical conformation.

Adhesins, Bacterial↗

PubMed1983

The action of selected agents on the accumulation of 18F by Streptococcus mutans.

The action of certain substances known to induce cellular alterations, or encounted in the oral cavity, on the accumulation of 18F by Streptococcus mutans GS-5 has been investigated. A 62-67% inhibition in the number of 18F atoms bound per mg dry weight of cells could be induced by a 15 min pretreatment with 2.7 X 10(-4) M cetyltrimethylammoniumbromide, 1 X 10(-1) M acetic anhydride, or 7 X 10(-2) M HCl. Plate counts indicated that alteration of the cellular composition rather than viability was responsible for this diminution in 18F accumulation. Prior exposure for 15 min of this organism to 1 M HCHO or 0.1 M NaOH did not alter 18F accumulation. Of the common salts encountered in the oral cavity, CaCl2 enhanced 18F binding. Pretreatment of the assay cells for 15-160 min with 0.1-10 mg/ml of trypsin, pronase, protease, alpha-glucosidase, dextranase, or lactoferrin had no significant effect on the accumulation of 18F. However, pre-exposure of cells for 60 min to 1-10 mg/ml of either amylase or lipase induced a 40-67% inhibition in the binding of 18F, while lysozyme enhanced the binding of 18F by the cells. It would appear then that the binding of 18F by S. mutans may be altered by certain substances encountered in the oral cavity.

Acetic Anhydrides↗

PubMed1975

Antigens of Streptococcus mutans: characterization of a polysaccharide antigen from walls of strain GS-5.

A cell wall-associated polysaccharide antigen was isolated from Streptococcus mutans GS-5 and appeared to determine serotype c specificity. Ouchterlony double-diffusion analysis of crude formamide extracts derived from purified cell walls of two serotype c strains (GS-5 and JC-2) showed complete identify when reacted with anti-GS-5 sera. Immunoelectrophoresis of this extract demonstrated the typical mobility for this serotype as described by others. Column chromatography on BioGel P-100 of the crude formamide extracts derived from GS-5 walls resulted in a single antigenic peak being resolved. This material, when loaded onto a diethylaminoethylcellulose column and eluted with a linear gradient of ammonium carbonate (0.0 to 0.2 M), was resolved further into two serologically reactive peaks (I and II). Only two consituents, rhamnose and glucose, were detected in the purified column fractions. Peak 1 had a rhamnoseto-glucose molar ratio of 0.9:1.0, and peak II, the major resolvable fraction, had a molar ratio of 1.7:1.0, The peak II ratio was very similar to that found in the formamide extract residue pellet (1.6:1.0)9 Ouchterlony analysis of the crude formamide extract and the purified fractions revealed only partial identify between peaks I and II but complete identify between peak II and the crude extract. Likewise, immunoelectrophoresis showed no differences in mobility of peak II and the crude extract, whereas peak I moved towards the cathode. Possible structural relationships between the two antigenic fractions are discussed below. Hapten inhibition studies suggested that an alpha-glucosyl group is at the immunodeterminant site of the antigen.

Antigens, Bacterial↗