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Interactions between and within Streptococcus mutans and Streptococcus sobrinus isolated from humans harboring both species.

The prevalence of Streptococcus mutans and Streptococcus sobrinus was examined in plaque samples from small discrete areas of the buccal tooth surfaces of seven subjects. Strains of S. mutans and S. sobrinus were isolated and tested for bacteriocin-mediated interactions between and within the two species, using the stab inoculation technique. S. mutans and S. sobrinus did not colonize each tooth surface uniformly and, in plaque from small discrete sites, S. mutans and S. sobrinus were either undetected or present in different interspecies proportions. Within the same subject, there were no bacteriocin-mediated interactions between strains of the same mutans species and no difference in bacteriocin activity was found between the strains of S. mutans and S. sobrinus from different sites. When bacteriocin interactions were tested between isolated strains from all seven subjects a somewhat higher inhibition ability was found for producer strains isolated from plaque compared with those isolated from saliva. S. mutans appeared to be more bacteriocinogenic than S. sobrinus. Replacing the glucose in the medium with sucrose enhanced the bacteriocin activity of S. mutans towards other S. mutans strains but reduced the inhibitory interaction towards strains of S. sobrinus.

Adult↗

Effect of chlorhexidine on the relative proportions of Streptococcus mutans and Streptococcus sanguis in hamster plaque.

The effect of chlorhexidine on the proportions of Streptococcus mutans and Streptococcus sanguis in plaque was studied in hamsters fed a diet containing 28% sucrose. In animals given chlorhexidine in their drinking water for 10 d a decrease in the population of S. mutans and an increase of S. sanguis occurred in the plaque. Following the removal of chlorhexidine the population of S. mutans increased again in the presence of sucrose and the number of S. sanguis returned to initial values. When animals were given a sucrose-free diet the low proportion of S. mutans observed following the short-term chlorhexidine period persisted. These data indicate that there is an inverse relationship between the number of S. sanguis and S. mutans in plaque and that the sensitivity in vivo of S. mutans to chlorheximide can be used to suppress the population of S. mutans with a concomitant rise in the proportion of S. sanguis.

Animals↗

The effects of basic and acidic synthetic polypeptides on the adherence of the oral bacteria, Streptococcus mutans and Streptococcus sanguis, to hydroxyapatite.

Two basic and two acidic synthetic polypeptides that bind strongly to hydroxyapatite at neutral pH were tested to determine their influence on adsorption of two Streptococcus mutans and two Streptococcus sanguis strains to hydroxyapatite. The adsorption of the strains was significantly enhanced or reduced by the basic and acidic agents, respectively. Study of acidic polypeptides provided evidence of competition between the polypeptides and the bacterial cells for hydroxyapatite adsorption sites.

Adsorption↗

Susceptibility of Streptococcus mutans and Streptococcus sobrinus to cell wall inhibitors and development of a novel selective medium for S. sobrinus.

Representative strains of Streptococcus mutans and Streptococcus sobrinus showed differences in susceptibility to members of the monobactam group of beta-lactam antibiotics: S. sobrinus was less sensitive than S. mutans. The minimum inhibitory concentrations of aztreonam (AZT) and carumonam, both of which belong to this group, were 2,000 microg/ml for S. sobrinus and 125 microg/ml for S. mutans. Further addition of fosfomycin, bacitracin and sodium chloride to Mitis Salivarius agar (MS) supplemented with AZT resulted in growth inhibition of S. mutans and oral streptococci other than S. sobrinus, and was therefore used as a selective medium for S. sobrinus (MS-SOB medium). The average growth recovery of laboratory and clinically isolated strains of S. sobrinus on MS-SOB medium was 74.1% compared to that on MS medium. Seventy-eight percent of clinical samples in which S. sobrinus was detected yielded pure growth of S. sobrinus on MS-SOB medium.

Anti-Bacterial Agents↗

Antineoplastic agents inhibit the growth of Streptococcus mutans and Streptococcus sanguis in vitro.

The effect of methotrexate (MTX) and doxorubicin on the growth, metabolism and ultrastructure of Streptococcus mutans and Streptococcus sanguis was studied in vitro. Both anticancer drugs exerted an inhibitory effect on the oral streptococci. MTX was more inhibitory than doxorubicin. The minimum inhibitory concentrations (MICs) of MTX to S. mutans were 0.25-2.5 micrograms/ml and that of doxorubicin 0.2 mg/ml. The MICs of MTX and doxorubicin to S. sanguis were 0.025 micrograms/ml and 2.0-0.02 mg/ml, respectively. When saliva samples of patients with malignant tumors receiving various doses of MTX were analyzed, MTX was found to be secreted into the oral cavity at concentrations ranging from 0.014 to 4.486 micrograms/ml. The saliva of these patients was also found to inhibit the growth of S. mutans, and the inhibition zones were in accordance with the MIC values observed. The results suggest that anticancer therapy must be taken into account when the salivary microbiological findings of cancer patients are interpreted.

Adult↗

In vitro inhibitory effects of Polygonum cuspidatum on bacterial viability and virulence factors of Streptococcus mutans and Streptococcus sobrinus.

OBJECTIVES: Polygonum cuspidatum has been used in Korean folk medicine to improve oral hygiene. This study was performed to evaluate the effects of methanol extract from root of P. cuspidatum (MEP) on bacterial viability and the virulence factors of Streptococcus mutans and Streptococcus sobrinus. METHODS: To test the effects of MEP on bacterial viability, we determined the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) against 20 bacterial strains, including S. mutans and S. sobrinus, using a micro-dilution assay. In case of S. mutans and S. sobrinus, the assays for time-kill and bacterial growth rate at sub-MIC concentrations were also performed. To determine effects of the extract on the virulence factors of S. mutans and S. sobrinus, the assays for sucrose-dependent adherence, water-insoluble glucan formation, glycolytic acid production, and acid tolerance were performed at sub-MIC levels. Phytochemical analysis for constituents of MEP was carried out. RESULTS: MEP showed a broad antibacterial range (MIC 0.5-4 mg/ml). The MBC was two to four times higher than the MIC. The time-kill curves showed S. mutans and S. sobrinus were significantly killed after 1h of incubation. At sub-MIC levels, doubling times of S. mutans and S. sobrinus dose-dependently increased up to 211% and 123%, respectively. At sub-MIC levels, MEP also showed inhibitory effects on the virulence factors of S. mutans and S. sobrinus in a dose-dependent fashion. Phytochemical analysis revealed the presence of alkaloids, sterol/terpenes, tannins, flavonoids, and carbohydrates. CONCLUSION: These data indicate that MEP has inhibitory effects on bacterial viability at higher concentrations (> or =MIC) and the virulence factors of S. mutans and S. sobrinus at sub-MIC concentrations, suggesting that it might be useful for the control of dental plaque formation and subsequent dental caries formation.

Anti-Bacterial Agents↗

Effects of oxygen on pyruvate formate-lyase in situ and sugar metabolism of Streptococcus mutans and Streptococcus sanguis.

The strictly anaerobic metabolism of sugar in strains of Streptococcus mutans and Streptococcus sanguis was studied because deep layers of dental plaque are strictly anaerobic. Galactose-grown cells of these streptococcal strains had higher pyruvate formate-lyase activity than did glucose-grown cells. Among these strains, two strains of S. mutans had a significantly higher pyruvate formate-lyase activity than did the others. This enzyme is extremely sensitive to oxygen, and even in situ the enzyme was inactivated by exposure of the cells to air. Lactate was less than 50% of the total end product of the strictly anaerobic incubation of the galactose-grown cells of S. mutans with excess glucose, and a significant amount of formate, acetate, and ethanol was produced through the catalysis of pyruvate formate-lyase. But the cells exclusively produced lactate when exposed to air for 2 min before the anaerobic incubation. The metabolism of sorbitol by S. mutans was seriously impaired by the exposure of the cells to oxygen, and the metabolic rate was reduced to less than 1/20 of that found under strictly anaerobic conditions because of the inactivation of pyruvate formate-lyase. S. sanguis produced a smaller amount of the volatile products from glucose than did S. mutans because of the low level of pyruvate formate-lyase. However, the pyruvate formate-lyase in situ in S. sanguis was less sensitive to oxygen than was that in S. mutans. Because of this low sensitivity, S. sanguis metabolized glucose more rapidly under aerobic conditions, whereas the rates of the aerobic and anaerobic metabolism of glucose by S. mutans were similar, which suggests that S. mutans rather than S. sanguis can sustain the rapid sugar metabolism in the deep layers of dental plaque.

Acetyltransferases↗

[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↗

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↗

Longitudinal study of dental caries incidence associated with Streptococcus mutans and Streptococcus sobrinus in pre-school children.

Streptococcus mutans and Streptococcus sobrinus are known to be associated with the development of dental caries. In this study these bacteria were detected in pre-school children (each with primary dentition, age range 3-5 years, n = 60) using a PCR method, and then their presence was compared with the incidence of dental caries over a 1-year period. Plaque samples were collected from all erupted tooth sites using a sterile toothbrush. Dental examinations at the beginning of the study (baseline) and after 1 year were also performed to determine decayed, missing, filled teeth (dmft) scores using WHO caries diagnostic criteria. The prevalences of S. mutans and S. sobrinus across all the subjects were 61.7% and 56.6%, respectively; 13 subjects (21.7%) were positive for S. mutans alone, 10 (16.6%) were positive for S. sobrinus alone and 24 (40.0%) were positive for both S. mutans and S. sobrinus, whereas 13 (21.7%) were negative for both S. mutans and S. sobrinus. dmft scores of subjects positive for both S. mutans and S. sobrinus at baseline and after 1 year were significantly higher than of those positive for S. mutans alone at the same stages (P < 0.01 and P < 0.001, respectively). The caries incremental increase was also significantly greater in those with both bacteria detected (P < 0.05). Our results indicate that pre-school children harbouring both S. mutans and S. sobrinus have a significantly higher incidence of dental caries than those with S. mutans alone.

Child, Preschool↗

Simple and rapid detection of Streptococcus mutans and Streptococcus sobrinus in human saliva by polymerase chain reaction.

Streptococcus mutans and Streptococcus sobrinus are major pathogens causing dental caries in humans. A simple and rapid method to detect these species in human saliva simultaneously was developed using the polymerase chain reaction (PCR). Chromosomal DNA was extracted by boiling bacterial cells in lysis solution containing 1% Triton X-100. Oligonucleotide primers specific for portions of the glucosyltransferase genes (gtfB of S. mutans and gtfI of S. sobrinus) were designed. After PCR using two sets of these primers, S. mutans and S. sobrinus were specifically identified. The method was capable of amplifying DNA fragments specific for these species from chromosomal DNA extracted from 1 x 10(3) cells, or from 10 microliters of clinical saliva samples containing 1 x 10(3) colony-forming units of either streptococcal species. A second PCR, using the first PCR product as a template with newly designed internal primers, made it possible to detect 1 x 10(2) colony-forming units of either streptococcal species in 10 microliters of saliva samples. These results indicate that the PCR method developed in this study is useful for detecting S. mutans and S. sobrinus in saliva and that it can be used in epidemiological studies to evaluate the prevalence level of these organisms.

Adult↗

Detection of Streptococcus mutans and Streptococcus sobrinus on the permanent first molars of the Mosuo people in China.

Streptococcus mutans and Streptococcus sobrinus are associated with the development of dental caries. The purpose of this study was to differentiate these bacteria by morphology, biochemical characteristics and PCR, and to compare their occurrence with the prevalence of dental caries in the Mosuo people. Plaque samples were collected from the permanent first molar in 126 Mosuo people (83 females, 43 males, aged 25-55 years, average age 36.1 +/- 7.73). Dental status was recorded as DMFT by WHO caries diagnostic criteria. Males had a significantly lower prevalence of caries and DMFT than females: 11.4 vs. 86.9% and 1.65 vs. 6.95, respectively (p<0.001). Morphological and biochemical tests gave unreliable results. The prevalence of S. mutans and S. sobrinus was 75.4 and 57.1%, respectively. 26.5% of females and 53.5% of males were positive for S. mutans alone, 18.1% of females and 16.3% of males were positive for S. sobrinus alone, while 50.6% of females and 18.6% of males were positive for both S. mutans and S. sobrinus and 4.8% of females and 11.6% of males were negative for both species. The DMFT scores of subjects positive for both S. mutans and S. sobrinus were significantly higher than of those positive for either S. mutans or S. sobrinus alone. These results indicate that subjects harboring both S. mutans and S. sobrinus have a significantly higher prevalence of dental caries than those with S. mutans or S. sobrinus alone.

Adult↗

Differentiation of Streptococcus mutans and Streptococcus sobrinus via genotypic and phenotypic profiles from three different populations.

Routine identification of Streptococcus mutans and Streptococcus sobrinus is generally based upon growth on various selective media, colony morphology and biochemical characteristics. We examined various approaches of differentiating these two species through a combination of the conventional phenotypic methodology with chromosomal DNA fingerprint (CDF) and arbitrarily primed polymerase chain reaction (AP-PCR) methods. Initially, ten ATCC type strains and 20 randomly selected clinical isolates of mutans streptococci (MS) were characterized and grouped into two major types based on patterns generated by the CDF using HaeIII digestion. The CDF's patterns with restriction fragments equal to or greater than 6.6 kb were defined as the CDF-1 group. The CDF's patterns with restriction fragments less than 6.6 kb were defined as the CDF-2 group. Both groups were then examined for biotype, serotype, and composition of DNA via thermal denaturation. AP-PCR was applied and evaluated for the capability of delineating S. mutans from S. sobrinus strains. Results of this study showed that all CDF-1 strains fit within a G+C range of 36.2% to 42.2%, whereas the CDF-2 strains had a G+C range of 45.8% to 47.0%. The serotyping assay exhibited 100% sensitivity, 90% specificity and 86.7% agreement with the CDF. The biotyping assay presented the poorest specificity (38.5%), indicating the highest variability. The capability of AP-PCR in differentiation of S. mutans from S. sobrinus was comparable to the CDF method, suggesting that either of these two approaches can and may serve as a viable alternative method to serotyping or biotyping of MS.

Analysis of Variance↗

Stability and activity of specific antibodies against Streptococcus mutans and Streptococcus sobrinus in bovine milk fermented with Lactobacillus rhamnosus strain GG or treated at ultra-high temperature.

Passive local immunization against dental caries is a promising approach to its prevention, as clinical evidence of active oral or nasal immunization is still limited and controversial. By means of systemic immunization of pregnant cows with a multivalent vaccine, high titres of IgG antibodies against human cariogenic bacteria, Streptococcus mutans and Streptococcus sobrinus, were produced in bovine colostrum. The purified immune product (IP) of this preparation has a number of anticariogenic properties, such as inhibition of streptococcal adherence to saliva-coated hydroxyapatite and inhibition of glucosyltransferase enzymes. This study investigated whether IP antibodies remained active and functional when added to ultra-high temperature (UHT)-treated milk or to Lactobacillus rhamnosus GG (LGG)-fermented milk stored for an extended time. LGG was chosen because of its widely known health benefits in humans and animals. A commercial UHT toddler's milk was supplemented with IP and stored for 2 months at 5, 21 and 30 degrees C. The antistreptococcal titres in UHT milk did not decline at any temperature during storage, and UHT-IP inhibited the adherence of S. mutans for up to 2 months. This was not the case with UHT toddler's milk without IgG antibodies. Milk was fermented with live LGG cells in the presence or absence of 5% IP. The antistreptococcal titres declined to about 30% of the original titres after storage. Fresh milk alone slightly enhanced streptococcal adhesion but fresh milk with IP inhibited the adherence of S. mutans by over 50%. LGG-positive fermented milk without antibodies also inhibited (P < 0.05) the adhesion by about 40%. In both LGG-fermented and UHT immune milk, the activity of antibodies against cariogenic streptococci was maintained during the expected shelf-life of these products. From the anticariogenic point of view it may be beneficial to add bovine-specific antibodies against mutans streptococci to probiotic LGG-containing milk products.

Animals↗

Longitudinal study of transmission, diversity, and stability of Streptococcus mutans and Streptococcus sobrinus genotypes in Brazilian nursery children.

The aim of this study was to perform a follow-up evaluation of the Streptococcus mutans and Streptococcus sobrinus colonization profile of children's oral cavities, which included the pattern of vertical transmission from mother to child, genotypic diversity, and stability of the strains. The subjects were 16 mother-child pairs, who were monitored for 20 months. Samples of saliva, tongue dorsum, alveolar ridge mucosa, and dental plaque from the children were collected bimonthly. Saliva samples from the mothers were also collected. After isolation and identification, the arbitrarily primed PCR method was performed for the genotypic characterization of S. mutans (968 isolates) and S. sobrinus (111 isolates). At the time the strains were acquired, the children harbored one to four distinct genotypes of S. mutans and only one genotype of S. sobrinus. Although S. mutans prevalence and genotypic diversity were greater than those of S. sobrinus, the presence of matching genotypes of S. mutans and S. sobrinus was similar (in 81.25 and 83.33% of mother-child pairs, respectively), suggesting vertical transmission for both species. This longitudinal study showed an increase in genotypic diversity of S. mutans in the oral cavity during the follow-up period: most of the initially acquired genotypes persisted, normally those genotypes transmitted by the mother, and some were lost during follow-up; new strains were also acquired. In conclusion, S. mutans and S. sobrinus genotypes acquired from maternal or alternative sources may show effective persistence in the oral cavity and/or transitory detection in the children's mouths, reflecting the continuous development of oral microbiota in children.

Adult↗

Adherence of Streptococcus mutans and Streptococcus sanguis to salivary components bound to glass.

Adherence of radiolabeled Streptococcus mutans and Streptococcus sanguis to saliva-treated glass surfaces was studied under conditions which minimized bacteria-glass interactions. Treatment of glass with an alkylsilane solution decreased nonspecific bacterial adherence and enhanced adsorption of radiolabeled salivary components to these surfaces. Addition of Triton X-100 to the bacterial suspensions also reduced nonspecific adherence to siliconized glass, but did not affect adherence to salivary components attached to siliconized glass. Calcium stimulated S. mutans adherence to saliva-free glass, but inhibited adherence to saliva-treated glass. S. sanguis adherence to either saliva-free or saliva-treated glass was inhibited slightly at high calcium ion concentrations. Adherence of streptococci to saliva-treated glass exhibited saturation kinetics, and the numbers of binding sites on the experimental salivary pellicle and the affinity constants for bacteria-saliva attachment were determined. Preincubation of the streptococci with whole saliva decreased their capacity to adhere to saliva-treated glass, but not to saliva-free glass. Bacteria adherent to saliva-treated glass surfaces were readily desorbed by washing with saliva. The addition of homologous antisera, ammonium sulfate-precipitated immunoglobulins, or Fab fragments to the bacterial suspensions inhibited cell adherence to saliva-treated glass.

Calcium↗

Competition and coexistence between Streptococcus mutans and Streptococcus sanguinis in the dental biofilm.

The human mucosal surface is colonized by the indigenous microflora, which normally maintains an ecological balance among different species. Certain environmental or biological factors, however, may trigger disruption of this balance, leading to microbial diseases. In this study, we used two oral bacterial species, Streptococcus mutans and Streptococcus sanguinis (formerly S. sanguis), as a model to probe the possible mechanisms of competition/coexistence between different species which occupy the same ecological niche. We show that the two species engage in a multitude of antagonistic interactions temporally and spatially; occupation of a niche by one species precludes colonization by the other, while simultaneous colonization by both species results in coexistence. Environmental conditions, such as cell density, nutritional availability, and pH, play important roles in determining the outcome of these interactions. Genetic and biochemical analyses reveal that these interspecies interactions are possibly mediated through a well-regulated production of chemicals, such as bacteriocins (produced by S. mutans) and hydrogen peroxide (produced by S. sanguinis). Consistent with the phenotypic characteristics, production of bacteriocins and H2O2 are regulated by environmental conditions, as well as by juxtaposition of the two species. These sophisticated interspecies interactions could play an essential part in balancing competition/coexistence within multispecies microbial communities.

Adaptation, Biological↗