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PubMed1993

Distribution of Streptococcus mutans and Streptococcus sobrinus at sub-sites in human approximal dental plaque.

The distribution and prevalence of Streptococcus mutans and Streptococcus sobrinus were determined at three sub-sites in human approximal plaque: away from (A), to the side of (S) and below (B) the contact area. Small plaque samples were taken from all three sub-sites on clinically sound approximal surfaces of a single premolar from each of 21 schoolchildren. S. mutans was detected significantly more often and in higher proportions than S. sobrinus from sub-sites A (p = 0.019), S (p = 0.034) and B (p = 0.004). S. mutans was detected in highest proportions from the B site compared to the A site (p = 0.025); there were no significant differences in the isolation frequency or prevalence of S. sobrinus from any of the sub-sites. S. mutans and S. sobrinus were never isolated together from the A sub-sites and were recovered together most commonly from the B sub-sites (p < 0.01). It is concluded that S. mutans and S. sobrinus preferentially colonise the most caries-prone site apical to the contact area.

Bacteria, Anaerobic↗
PubMed2000

PCR for detection and identification of Streptococcus sobrinus.

Oligonucleotide primers were designed based upon a comparison of the dextranase gene (dex) sequences from Streptococcus sobrinus and S. mutans. The primers amplified a 1610-bp long DNA fragment on the dex gene by a PCR. The pair of primers was specific to S. sobrinus as the other members of the mutans streptococci - S. mutans, S. downei, S. cricetus, S. rattus, S. macacae and S. ferus - gave no PCR products. Other gram-positive oral bacteria (15 strains of 10 species of cocci and 18 strains of 12 species of rods) and gram-negative oral bacteria (3 strains of 3 species of cocci and 31 strains of 22 species of rods) also gave negative results in the PCR. The PCR procedure was able to detect as little as 100 fg of purified chromosomal DNA or as few as 9 cfu of S. sobrinus NIDR6715. Seven clinical isolates of S. sobrinus were also positive in the dex PCR. This laboratory developed the S. mutans-specific PCR (dexA PCR) method with the primers specific for a portion of the dextranase gene of S. mutans Ingbritt. Primers for the dex and dexA PCR methods detected two species exclusively from the mutans streptococci. Furthermore, these two species were effectively differentiated by the species-specific amplicons with different lengths. The application of the PCR method to human dental plaque showed that the prevalence of S. sobrinus (83%) in oral cavities was higher than currently supposed (0-50%). These results suggest that the described PCR method is suitable for the specific detection and identification of human cariogenic bacteria, S. sobrinus and S. mutans.

Animals↗
PubMed1999

[Effects of monoclonal antibody against the major cell surface protein on salivary agglutinin mediated aggregation of Streptococcus sobrinus].

OBJECTIVE: To investigate the effects of monoclonal antibody on salivary agglutinin-mediated aggregation of Streptococcus sobrinus in vitro. METHODS: Bacterial aggregation inhibition was measured under OD650 at 10 minutes intervals for 1 hour. RESULTS: It showed that the McAb ZS2/286 raised against the PAg of S. sobrinus 6715 could inhibit the aggregation by some degree, but the effect was not significant. CONCLUSION: The McAb ZS2/286 might not bind directly to the functionally important region of PAg related to aggregation.

Agglutinins↗
PubMed1991

Chronic peroral administration of Streptococcus sobrinus to conventional laboratory rats produces cycling levels of salivary antibodies.

Conventional outbred rats were fed Streptococcus sobrinus for 24 weeks and ELISA was used to identify isotypes of antibodies against bacteria in saliva and serum. Quantities of antibodies from experimental rats were compared with values derived from the control population. Saliva IgM and IgA anti-S. sobrinus from experimental rats were greater than controls at week 3, were much less at week 9, but normal levels were found by week 13. IgG antibodies in saliva peaked at weeks 5 and 9 but fell to control levels by week 13. Relative levels of antibodies in saliva of experimental animals continued to cycle during weeks 13-24 but did not differ greatly from controls. Serum IgM and IgG antibodies to S. sobrinus were essentially like controls throughout the experiment. Serum IgA increased briefly during the first 12 weeks then returned to normal levels. The results suggest that prolonged peroral exposure to cariogenic bacteria ultimately leads to modulation of antibody around unimmunized control levels even though antigenic stimulation persists.

Animals↗
PubMed1987

Nucleotide sequence of a glucosyltransferase gene from Streptococcus sobrinus MFe28.

The complete nucleotide sequence was determined for the Streptococcus sobrinus MFe28 gtfI gene, which encodes a glucosyltransferase that produces an insoluble glucan product. A single open reading frame encodes a mature glucosyltransferase protein of 1,559 amino acids (Mr, 172,983) and a signal peptide of 38 amino acids. In the C-terminal one-third of the protein there are six repeating units containing 35 amino acids of partial homology and two repeating units containing 48 amino acids of complete homology. The functional role of these repeating units remains to be determined, although truncated forms of glucosyltransferase containing only the first two repeating units of partial homology maintained glucosyltransferase activity and the ability to bind glucan. Regions of homology with alpha-amylase and glycogen phosphorylase were identified in the glucosyltransferase protein and may represent regions involved in functionally similar domains.

Amino Acid Sequence↗
PubMed2000

Streptococcus sobrinus antigens that react to salivary antibodies induced by tonsillar application of formalin-killed S. sobrinus in rabbits.

We previously found that tonsillar application of antigen induces a strong antibody response to Streptococcus sobrinus in saliva and blood plasma. Rabbits immunized against S. sobrinus by tonsillar application were highly resistant to experimental dental caries triggered by oral inoculation of living S. sobrinus organisms with sucrose. In the present study, we examined the reaction of S. sobrinus antigens to the antibodies induced by the tonsillar application of S. sobrinus AHT-k in rabbits and compared them to those antibodies induced by intramuscular injection. In an enzyme-linked immunosorbent assay using ultrasonic fragments from mutans group streptococci, the saliva and blood plasma selectively reacted to S. sobrinus AHT-k (serotype g) and serologically related streptococci (serotypes a, d, and h) in the sixth week after tonsillar application, whereas the blood plasma in the sixth week after intramuscular injection reacted to the unrelated streptococci (serotypes b, c, e, and f) in addition to the aforementioned streptococci. The antibody reactivity induced after tonsillar application was not lost after treatment of the antigen with heat or proteinase digestion, whereas these treatments resulted in a 70% decrease of the antibody reactivity induced by intramuscular injection. The inhibition by haptenic sugars and the decrease in immunoreactivity by heat treatment and proteinase digestion suggested that 80% of the antibodies induced by tonsillar application reacted to saccharides. These saccharide antigens appeared to be involved in a specific reaction with S. sobrinus-specific streptococci and a selective reaction with serologically related streptococci. These antigens are probably involved in anticaries reactions in experimental dental caries.

Animals↗
PubMed1987

Oral immunization of humans with Streptococcus sobrinus glucosyltransferase.

The effect of oral administration of glucosyltransferase (GTF) from Streptococcus sobrinus 6715 on levels of immunoglobulin A (IgA) antibody to GTF in parotid saliva and on the number of indigenous Streptococcus mutans in the whole saliva was studied in young adult males. GTF combined with aluminum phosphate (AP) was administered in capsules to 14 subjects, while sodium phosphate buffer combined with AP was administered in the same way to 11 control subjects. Thirteen administrations were given during the first immunization regimen, and five administrations, approximately 3 months later, constituted the second immunization regimen. All subjects were given professional dental prophylaxis immediately prior to each immunization. Each subject served as his own control by using antibody and bacterial data collected prior to antigen administration for comparison. After the first immunization regimen, the GTF vaccine group exhibited a significantly higher distribution (P less than 0.05) of normalized parotid saliva IgA antibody elevations than observed in the placebo group. Between the first and second immunization regimens a significant increase (P less than 0.05) in parotid salivary anti-GTF activity also occurred in the GTF vaccine but not the placebo group. No significant differences between these two groups were observed on any occasion when serum IgG or IgA antibody to GTF was analyzed. Comparison of the group mean log ratios (post- to prevaccine administration) of S. mutans to total streptococci in whole saliva revealed that the GTF vaccine group values were always lower than those of the placebo group. These differences reached significance (P less than 0.01) on three of the last four sampling occasions (days 21, 35, and 42) following initiation of the first immunization regimen. The mean log ratios of the GTF vaccine group were also lower than those of the placebo group after the second immunization regimen but did not reach significance. These data indicate that oral administration of GTF from the mutans streptococci has the potential to elicit a salivary IgA antibody response when combined with an aluminum-based adjuvant and that this response can interfere with the reaccumulation of indigenous S. mutans following dental prophylaxis.

Administration, Oral↗
PubMed1992

Mechanism of water-insoluble glucan synthesis in Streptococcus sobrinus.

Synthesis of water-insoluble glucan (IG) by 1,3-alpha-D-glucan synthase from Streptococcus sobrinus was examined using methylation analysis. The purified enzyme was incubated with sucrose and dextran T2000 (DT2000) for a given time and only IG was harvested by centrifugation. The remaining supernatant was incubated again, and IG was obtained. By repeating the above method using the residual supernatant, we obtained 5 varieties of IG precipitated in different periods. These IGs were methylated and examined using gas chromatograph mass spectrometry. The DT2000 water-insolubilized in the early reaction stage tended to have a highly ramified structure, with 1,3-alpha-D-glucan on a 1,6-alpha-D-glucan chain as the main chain. On the contrary, the DT2000 water-insolubilized in the late stage tended to have sparse side chains of 1,3-alpha-D-glucan which elongated with incubation. Specifically, the greater the number of side chains, the sooner the DT2000 was insolubilized. These results suggest that water-insolubilization of the water-soluble glucan not only depends on the increase of the ratio of 1,3-alpha-glucoside linkages to 1,6-alpha-glucoside linkages but also on the degree of branching of the 1,3,6-alpha-branched glucoside linkages.

Glucans↗
PubMed1998

Root caries and some related factors in 88-year-old carriers and non-carriers of Streptococcus sobrinus in saliva.

The prevalence of caries on exposed root surfaces in 88-year-old subjects with and without salivary levels of Streptococcus sobrinus was studied. Ninety-two individuals were examined with regard to root caries lesions and fillings. The root caries index (RCI) was calculated and related to salivary flow rate and buffer capacity, plaque score and salivary counts of Streptococcus mutans, S. sobrinus and lactobacilli. In 89 subjects with exposed root surfaces, all but 2 harbored mutans streptococci; 51 subjects carried S. mutans only, 35 both S. sobrinus and S. mutans, and 1 S. sobrinus only. The RCI was significantly higher in persons with than those without S. sobrinus (p < 0.05). Subjects with both S. sobrinus and S. mutans had higher counts of total mutans streptococci and lactobacilli than subjects with only S. mutans (p < 0.05). The RCI was significantly correlated to S. sobrinus and S. mutans (p < 0.05). The positive correlation between the RCI and S. sobrinus was still significant when the other tested variables were kept constant, whereas the correlation between the RCI and S. mutans was weaker when S. sobrinus and lactobacilli were kept constant. The D-component of the RCI (DSr%) was significantly correlated to S. sobrinus, S. mutans and lactobacilli, whereas the F-component of the RCI showed no significant correlation to any of the tested variables. A stepwise multiple correlation showed that the variance of DSr% was best explained in the S. sobrinus carriers by S. sobrinus and the salivary buffer effect, and in the non-carriers by S. mutans.

Aged↗
PubMed2003

Development of a 5' nuclease-based real-time PCR assay for quantitative detection of cariogenic dental pathogens Streptococcus mutans and Streptococcus sobrinus.

A 5' nuclease TaqMan PCR assay was developed for the quantitative detection of the major cariogenic bacteria Streptococcus mutans and Streptococcus sobrinus. The absolute and relative numbers of bacteria were measured by this method. This assay will be useful for quantifying these organisms in oral specimens and for analyzing biofilm formation.

Biofilms↗
PubMed1991

Dental location of Streptococcus mutans and Streptococcus sobrinus in humans harboring both species.

The distribution and prevalence of Streptococcus mutans and Streptococcus sobrinus were determined in plaque samples from the cervical areas of all buccal, lingual and approximal tooth surfaces and from the fissures of all occlusal sites in 40 subjects harboring both species. S. mutans was detected more often and in higher numbers than S. sobrinus. There were more teeth detected with S. mutans only than with S. sobrinus only. Most teeth harbored both of these mutans streptococci species, indicating a positive association. The highest numbers of CFU for both species were detected on the molars, with the lowest incidence on the anterior teeth. The presence of S. mutans was relatively similar on all teeth tested, while the presence of S. sobrinus was relatively higher on the molars compared to the anterior teeth. S. mutans and S. sobrinus were found to colonize the buccal surfaces in almost equal numbers. On all other surfaces, S. mutans was detected more frequently or in higher numbers compared to S. sobrinus. No significant differences could be found in the relative proportions of S. mutans and S. sobrinus between sound, decayed or filled tooth surfaces.

Adolescent↗
PubMed1994

Effect of seaweed extracts on Streptococcus sobrinus adsorption to saliva-coated hydroxyapatite.

The inhibitory effect of the extracts of seaweeds on the adsorption of Streptococcus sobrinus B13 to the tooth surface was investigated in vitro. Using saliva-coated hydroxyapatite beads (S-HA) as experimental salivary pellicle, extracts from Gloiopeltis furcata and Gigartina teedii (Rhodopheceae) strongly inhibited the adsorption of S. sobrinus B13 to S-HA. The G. furcata extract, which strongly inhibited the adsorption of S. sobrinus B13 to S-HA, was a high molecular weight sulfated polysaccharide. The result of I.R. analysis indicated that the active component in this extract of G. furcata was funoran.

Bacterial Adhesion↗
PubMed2003

[Rapid detection of Streptococcus mutans and streptococcus sobrinus in human saliva by nested polymerase chain reaction].

OBJECTIVE: To establish a simple and rapid method to detect Streptococcus mutans and streptococcus sobrinus simultaneously in human saliva. METHODS: Chromosomal DNA from the bacteria was obtained by the extraction method with phenol-chloroform. A nested PCR method with two sets of primers specific for portions of the glucosyltransferase genes (gtfB of S. mutans and gtfI of S. sobrinus), was optimized to detect S. mutans and S. sobrinus from standard strains, clinical strains and directly in human saliva. RESULTS: The first process of nested PCR was capable of amplifying DNA fragments specific for these species from chromosomal DNA extracted from 10(5) CFU cells of standard and clinical strains, or from 1 ml clinical saliva samples containing 10(5) CFU cells of either species. a second process of nested PCR, using the first PCR product as a template with new internal primers to detect 10(3) CFU of either streptococcal species in 1ml saliva samples. CONCLUSION: Nested PCR could detect S. mutans and S. sobrinus rapidly and simply in human saliva. This finding would be important to studies of elucidation the role of these two streptococcal species in the etiology of dental caries.

Humans↗
PubMed1991

A simple biochemical scheme for the differentiation of Streptococcus mutans and Streptococcus sobrinus.

A collection of wild-type and reference strains of Streptococcus mutans and Streptococcus sobrinus was compared using a set of fermentation and enzymic tests in order to establish a simple scheme for the differentiation of these two species isolated from selective media. Tests for the production of acid from N-acetylglucosamine, arbutin and melibiose, and the presence of alpha-galactosidase and alpha-glucosidase activities were useful in differentiating these species. Strains of Streptococcus mutans unable to ferment melibiose were also frequently unable to ferment other sugars; a consideration of the genetic control and organization of the genes involved in melibiose metabolism indicates that certain of the conventional characterization tests are not measurements of independent biochemical capabilities.

Adolescent↗
PubMed1993

Effects of N-acetylglucosamine on carbohydrate fermentation by Streptococcus mutans NCTC 10449 and Streptococcus sobrinus SL-1.

We have investigated the ability of two species of streptococci isolated from the human oral cavity (Streptococcus mutans NCTC 10449 and Streptococcus sobrinus SL-1) to metabolize N-acetylglucosamine (GlcNAc), a naturally occurring amino sugar present in saliva and human glycoproteins, when provided as the sole fermentable carbohydrate and determined the effects of the presence of GlcNAc on the fermentation of other carbohydrates. S. mutans used GlcNAc at concentrations of up to 10 mM to increase cell numbers, but S. sobrinus was unable to ferment the amino sugar alone and its uptake only occurred in the presence of a fermentable carbohydrate. GlcNAc had a marked inhibitory effect on the ability of S. sobrinus to produce lactic acid from glucose, sucrose, and fructose, at the same time increasing the lag period and doubling time of batch-grown cells. Such patterns of inhibition were found with S. mutans, but the effects were less than those seen in S. sobrinus. In mixed culture studies of the two species, S. sobrinus became the predominant organism when 10 mM glucose was supplied as the sole fermentable carbohydrate, with a concomitant decrease in the numbers of S. mutans cells, but supplementation of the broth with 10 mM glucose and 10 mM GlcNAc resulted in the emergence of S. mutans as the predominant organism. S. mutans and S. sobrinus grown in media containing glucose possessed the ability to transport glucose and GlcNAc, probably via the same glucose-phosphotransferase system at similar rates. However, intracellular levels of N-acetylglucosamine-6-phosphate deacetylase and glucosamine-6-phosphate deaminase were markedly higher in S. mutans grown on glucose and GlcNAc than in S. sobrinus: 34 and 398 and 8 and 17 nmol of NADPH formed per mi per mg of protein for S. mutans and S. sobrinus, respectively. We propose that GlcNAc inhibited growth of S. sobrinus in media containing glucose and GlcNAc by competing with glucose for the glucose phosphotransferase, depleting intracellular levels of phosphoenolpyruvate, and possessing, in contrast to S. mutans, low levels of N-acetyl-glucosamine-6-phosphate deacetylase and glucosamine-6-phosphate deaminase activity. Together, these data suggest that in dental plaque, S. sobrinus when exposed to GlcNAc will have a reduced ability to compete with S. mutans for dietary carbohydrates, contributing to the greater frequency of isolation of S. mutans from human populations.

Acetylglucosamine↗
PubMed2006

In vitro antibacterial activity of the peptide PsVP-10 against Streptococcus mutans and Streptococcus sobrinus with and without glycocalyx.

The antibacterial activity of the peptide PsVP-10 obtained from Pseudomonas sp. R10 against Streptococcus mutans and Streptococcus sobrinus was investigated. One hundred and twenty strains of S. mutans and 120 strains of S. sobrinus with and without glycocalyx were isolated from saliva samples in trypticase-yeast-cysteine-sucrose-bacitracin (TYCSB) agar. Bacterial identification was made by polymerase chain reaction. Glycocalyx production was observed in modified TYCSB agar and confirmed with a modified version of the microplate adherence assay. The minimum inhibitory concentration (MIC) of PsVP-10 bacteriocin was determined by means of the agar dilution method, and the time of bacterial death was calculated by means of colony-forming unit counts. The MIC of the bacteriocin PsVP-10 for both bacterial species with and without glycocalyx was < 2 mg/L and the time of bacterial death was less than 240 s for all the studied bacterial strains. Thus, bacteriocin PsVP-10 could be an interesting possibility to combat these cariogenic bacterial species.

Anti-Bacterial Agents↗
PubMed2000

Signal sequence and alanine-rich region of streptococcal protein antigen A of Streptococcus sobrinus can direct localization of alkaline phosphatase to the periplasm of Escherichia coli.

Streptococcal protein antigen A (SpaA) of Streptococcus sobrinus is expressed on the surface of cells and extracellularly. TnphoA which lacks signals for transcription and membrane transport of Escherichia coli alkaline phosphatase was used to analyze the sequences necessary for transport of a SpaA/PhoA fusion protein across the cytoplasmic membrane to the periplasm of E. coli cells. Of 15 alkaline phosphatase-producing isolates analyzed, all were found to localize more than 85% of the SpaA/PhoA hybrid protein to the periplasm of E. coli cells. From DNA sequence analysis, all were found to have TnphoA inserted into an identical site. The insertion site of TnphoA was downstream from the coding sequence that generates four tandemly repeated alanine-rich sequences of 82 amino acid residues. These results suggest that in addition to the signal sequence, mature protein sequences containing alanine-rich repeat sequences may play a role in the export of the SpaA protein across a bacterial membrane.

Alanine↗
PubMed2000

Single-molecule imaging of interaction between dextran and glucosyltransferase from Streptococcus sobrinus.

Using total internal reflection fluorescence microscopy, we directly observed the interaction between dextran and glucosyltransferase I (GTF) of Streptococcus sobrinus. Tetramethylrhodamine (TMR)-labeled GTF molecules were individually imaged as they were associating with and then dissociating from the dextran fixed on the glass surface in the evanescent field. Similarly dynamic behavior of TMR-labeled dextran molecules was also observed on the GTF-fixed surface. The duration of the stay on the surface (dwell time) was measured for each of these molecules by counting the number of video frames that had recorded the image. A histogram of dwell time for a population of several hundred molecules indicated that the GTF-dextran interaction obeyed an apparent first-order kinetics. The rate constraints estimated for TMR-labeled GTF at pH 6.8 and 25 degrees C in the absence and presence of sucrose were 9.2 and 13.3 s(-1), respectively, indicating that sucrose accelerated the dissociation of GTF from dextran. However, the accelerated rate was still much lower than the catalytic center activity of GTF (> or = 25 s(-1)) under comparable conditions.

Dextrans↗