Inhibitory effects of humic acid on insoluble glucan synthesis and cellular adherence of Streptococcus mutans (sobrinus) 6715.
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Bacterial Adhesion↗
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Colostral proteins from cows immunised with Streptococcus mutans/S. sobrinus support the phagocytosis and killing of mutans streptococci by human leucocytes.
Passive immunisation, based on bovine colostral preparations, is an area of active research. Specific bovine antibodies inhibit the virulence factors of target pathogens but the interactions between whey preparations and human immune defence cells are not well known. Bovine colostrum inhibits the phagocytic activity of bovine leucocytes and this may reflect the biological activity of immunoglobulins in it. Therefore, this study aimed to examine the effects of bovine whey protein preparations from the colostrum of Streptococcus mutans/S. sobrinus-immunised and sham-immunised cows on binding, ingestion and killing of these bacteria by human leucocytes. Binding and ingestion of FITC-labelled bacteria were estimated by flow cytometry and leukocyte activation was measured as chemiluminescence. Killing rate was estimated by plate counting and by measuring bioluminescence from S. mutans- containing the insect luciferase gene. Colostral whey protein preparation from hyperimmunised cows activated human leucocytes by opsonising specific bacteria. Neutrophils, eosinophils and monocytes weakly phagocytosed non-opsonised bacteria and bacteria opsonised with control product. On the contrary, binding and ingestion were efficient in the presence of the preparation from immunised cows. Thus, these results show that bovine colostral whey proteins are able to support the activation of human phagocytes against pathogenic microbes and that this property is related to specific antibodies in whey preparations. These whey proteins may also be clinically useful, especially in preventing the colonisation of newly erupted teeth by mutans streptococci.
Animals↗
Differentiation of mutans streptococci by intact cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
It is difficult to distinguish mutans streptococci on the species level, and even more so on the subspecies level. Intact cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) (ICM) was applied to reference strains of five of the species of the mutans group (Streptococcus criceti, Streptococcus downei, Streptococcus mutans, Streptococcus ratti, Streptococcus sobrinus), nonmutans streptococci (Streptococcus oralis, Streptococcus mitis, Streptococcus salivarius, and Streptococcus sanguinis), and 177 mutans streptococci isolated from saliva of 10 children. From the analysis of the reference strains, readily distinguishable ICM mass spectra were obtained for the different species. Based on multivariate statistical analysis, a correct and unambiguous assignment was made of the spectra of 159 isolated mutans streptococci to S. mutans and 16 isolates to S. sobrinus. Two isolates were sorted out and were identified by sequencing of their 16S rRNA genes as Streptococcus anginosus. In addition, ICM indicated a misclassification for some reference strains (AHT, V 100 and E 49) and re-classified AHT and E 49 as S. ratti and V 100 as S. sobrinus. This was confirmed by 16S rDNA sequencing. Based on a statistical similarity analysis of the spectra of reference strains and a quantitative assessment of the reproducibility of ICM, the isolates identified as either S. mutans or S. sobrinus were phenotyped on the subspecies level. In the population of the clinical isolates, 14 unambiguously different S. mutans and three different S. sobrinus phenotypes were detected. ICM proved to be a powerful tool for a differentiation of mutans streptococci down to the subspecies level.
Child↗
HLA-DR4 and salivary immunoglobulin A reactions to oral streptococci.
The aim of this study was to describe and compare salivary immunoglobulin A (IgA) antibody reactions to extracts of strains of three oral streptococci in human leukocyte antigen (HLA)-DR4-positive and -DR4-negative subjects. Whole paraffin-stimulated saliva samples were collected from 27 apparently healthy subjects. Previous HLA typing showed that 20 subjects were DR4 positive and 7 were DR4 negative. HLA-DRB1*04 subtyping was performed among the DR4-positive subjects. Whole-cell antigen extracts from Streptococcus mutans (KPSK 2), Streptococcus sobrinus (OMZ 65) and Streptococcus parasanguis (Nt 62) were separated in SDS-PAGE. The antigens were immunoblotted with diluted saliva (Western blot), scanned and analyzed in a computer system. All immunoblot bands were recorded in DR4-positive and DR4-negative saliva pools, and bands with an optical density >or=0.1 were selected for analysis in individual salivas. The DR4-negative subjects in general had more immunoblot bands and more distinct bands than did the DR4-positive subjects. A higher concentration of total IgA in saliva was correlated with more bands, especially to antigens separated from S. mutans. When the number of bands was calculated per IgA unit, significant differences were observed between DR4-positive and DR4-negative salivas. This was particularly seen for S. mutans and S. parasanguis. As the number of bands was analyzed in relation to DR4 subgroups, DRB1*04, there was a lower salivary IgA activity to S. mutans in the DRB1*0401 and *0404. The variable level of correlation previously demonstrated for S. mutans colonisation and serologically defined DR4 positive subjects might be explained by the heterogeneity in this group, and the relation should be sought on a subgroup level.
Antibodies, Bacterial↗
Antimicrobial activity of Croton cajucara Benth linalool-rich essential oil on artificial biofilms and planktonic microorganisms.
We have previously demonstrated that a linalool-rich essential oil from Croton cajucara Benth presents leishmanicidal activity. In the present study, we demonstrate that this essential oil inhibits the growth of reference samples of Candida albicans, Lactobacillus casei, Staphylococcus aureus, Streptococcus sobrinus, Porphyromonas gingivalis and Streptococcus mutans cell suspensions, all of them associated with oral cavity disease. The purified linalool fraction was only inhibitory for C. albicans. Microbes of saliva specimens from human individuals with fixed orthodontic appliances, as well as the reference strains, were used to construct an artificial biofilm which was exposed to linalool or to the essential oil. As in microbial suspensions, the essential oil was toxic for all the microorganisms, while the purified linalool fraction mainly inhibited the growth of C. albicans. The compounds of the essential oil were separated by thin layer chromatography and exposed to the above-cited microorganisms. In this analysis, the proliferation of the bacterial cells was inhibited by still uncharacterized molecules, and linalool was confirmed as the antifungal component of the essential oil. The effects of linalool on the cell biology of C. albicans were evaluated by electron microscopy, which showed that linalool induced a reduction in cell size and abnormal germination. Neither the crude essential oil nor the purified linalool fraction is toxic to mammalian cells, which suggests that the essential oil or its purified components may be useful to control the microbial population in patients with fixed orthodontic appliances.
Acyclic Monoterpenes↗
Effects of Weight Watchers' diet on salivary secretion rate, buffer effect and numbers of mutans streptococci and lactobacilli.
The aim of this investigation was to study how a change to the dietary regimen of the Weight Watchers' (WW) affected salivary secretion rate, buffer effect and number of cariogenic bacteria. Thirty-three women joining the WW were saliva sampled at the evening prior the day of the start of the dietary regimen and after 4 and 8 wk on the dietary regimen. No significant changes were found regarding saliva secretion rate and buffer effect. A significant decrease of salivary numbers of mutans streptococci and lactobacilli occurred. Between the 4th and 8th wk of dietary regimen an increase of cariogenic bacteria occurred but not back to the baseline values. At baseline 15 of the WW carried Streptococcus sobrinus in addition to Streptococcus mutans. The total number of salivary mutans streptococci and lactobacilli was significantly higher in those subjects. In conclusion the dietary regimen of the WW, based on food choice and distributions of meals in agreement with a non-cariogenic diet, reduced the number of cariogenic bacteria in most subjects, but not in all, while the secretion rate and buffer effect remained the same.
Adult↗
Anti-cariogenic properties of a water-soluble extract from cacao.
The addition of a water-soluble extract from cacao-extracted powder (CEPWS) to a cariogenic model food, a white chocolate-like diet that contains 35% sucrose, significantly reduced caries scores in SPF rats infected with Streptococcus sobrinus 6715, compared to control rats fed a white chocolate-like diet. CEPWS markedly inhibited water-insoluble glucan (WIG) synthesis through crude glucosyltransferases (GTFs) from Streptococcus sobrinus B13N in vitro. GTF-inhibitor(s) in CEPWS was prepared through three-step fractionation, and was termed CEPWS-BT, which is a high molecular weight (>10 kDa) heat-stable matrix of sugar, protein, and polyphenol. When the inhibitory effect of CEPWS-BT on glucan synthesis was examined using the purified GTF-I, GTF-T, and GTF-U enzymes from S. sobrinus B13N, significant reduction in GTF-I and GTF-T activity as a result of adding CEPWS-BT at low concentrations was observed. These results suggest that the addition of CEPWS to cariogenic food could be useful in controlling dental caries.
Animals↗
In vitro and in vivo effects of isolated fractions of Brazilian propolis on caries development.
Recently, two chemically different types of Brazilian propolis (type-3 and -12) were shown to have cariostatic properties. This study aimed to evaluate the influence of their isolated fractions on mutans streptococci viability, glucosyltransferases (GTFs) activity and caries development in rats. The ethanolic extracts of propolis (EEPs) were serially fractionated into hexane (H-fr), chloroform, ethyl acetate, and ethanol. The ability of the four fractions and EEP to inhibit Streptococcus mutans and Streptococcus sobrinus growth and adherence to a glass surface was examined. The effect on GTFs B and C activity was also determined. For the caries study, 60 Wistar rats infected with Streptococcus sobrinus were treated topically twice daily as follows: (1) EEP type-3, (2) H-fr type-3, (3) EEP type-12, (4) H-fr type-12, and (5) control. In general, the H-fr from both types of propolis showed the highest antibacterial activity and GTFs inhibition. Furthermore, the EEP and H-fr type-3 and -12 were equally effective in reducing dental caries in rats. The data suggest that the putative cariostatic compounds of propolis type-3 and -12 are mostly non-polar; and H-fr should be the fraction of choice for identifying further potentially novel anti-caries agents.
Animals↗
Kinetic properties of glucosyltransferase adsorbed onto saliva-coated hydroxyapatite.
Results from previous studies have shown that several properties of glucosyltransferase (GTF) adsorbed onto saliva-coated hydroxyapatite beads differ from those of GTF in solution. For example: thermostability, pH-activity dependency, sensitivity to inhibitors. The aim of this study was to compare the kinetics of the adsorbed GTF with its kinetic properties in solution. Hydroxyapatite beads were coated with human parotid saliva (sHA). Following washes, cell-free GTF enzyme from Streptococcus sobrinus 6715 (S. sobrinus 6715) or Streptococcus mutans GS-5 (S. mutans GS-5) was adsorbed onto sHA. The GTF-coated sHA were then incubated with radiolabeled sucrose for intervals of 5-360 minutes and the amount of glucans synthesized in situ by the adsorbed GTF was determined and compared with that produced in solution. The adsorbed GTF (from S. sobrinus 6715) exhibited a sharp increase in glucan production within the first 5 minutes of incubation while surface-bound GTF of S. mutans GS-5 displayed an initial burst of activity within the first 15 minutes of incubation. During the next 6 hours (duration of experiment) the amount of glucan on the beads did not increase with either enzyme. In contrast, the kinetic profile of the two GTFs in solution demonstrated a linear increase in the amount of glucans formed, with no initial burst effect. The results indicate that the rapid formation of glucans by GTF adsorbed onto sHA could have implications for colonization by oral microorganisms on tooth surfaces. The accelerated synthesis of glucan on tooth surfaces may affect the microbiology of the dental plaque, and might also influence the movement of substances, such as acids and antiplaque agents, across the acquired pellicle and dental plaque.
Adsorption↗
Effect of acidulated phosphate fluoride (APF) gel on the adherence of cariogenic bacteria to resin composites.
PURPOSE: To evaluate the effect of 1.23% acidulated phosphate fluoride (APF) and the subsequent polishing on the in vitro adherence of mutans streptococci to resin composites in a model that simulated the oral cavity conditions. METHODS: Samples of light-curing resin-based composites (Z100 and Z250) were treated with 1.23% APF gel (Sultan) for 1 minute and polished subsequently with Sof-Lex disks. The bacterial adherence of Streptococcus mutans and Streptococcus sobrinus to the APF gel treated resin samples and the subsequent polished samples were analyzed and compared with the adherence to the non-treated controls. RESULTS: The 1.23% APF gel caused a significant increase in the adherence of Streptococcus mutans and Streptococcus sobrinus. The increasing rate of the bacterial adherence to resin composites after APF treatment ranged from 73.5% to 167% and differed according to the bacterial strains. A decrease in the adherence of the bacteria was found after subsequent polishing with Sof-Lex disks.
Acidulated Phosphate Fluoride↗
In vitro screening of Chinese medicinal toothpastes: their effects on growth and plaque formation of mutans streptococci.
Many plant extracts or derivatives have been incorporated into commercial toothpastes to treat oral diseases related to caries or periodontal diseases in China. However, no information is available concerning their in vitro effects on oral bacteria. Thirty-one Chinese medicinal toothpastes were selected for this study. Their ability to inhibit growth, in vitro plaque formation and glucosyltransferase (GTF) activity of Streptococcus sobrinus B13 and Streptococcus mutans 3209 were examined. Eighty-seven percent of the tested toothpastes inhibited the growth of the mutans streptococci, with zones ranging from 0.8 to 2.5 cm. At 10 mg/ml, 74% of the toothpastes inhibited in vitro plaque formation by S. mutans. Among these, 60% completely suppressed water-insoluble glucan synthesis from sucrose by GTF. Based on data obtained from our study, the incorporation of natural plant products or their derivatives into dentifrices seems a feasible means of promoting oral health and controlling dental diseases.
Dental Plaque↗
The influence of sucralose on bacterial metabolism.
Sucralose (1',4',6' trideoxy-trichloro-galactosucrose) is a nontoxic, intensely sweet sucrose derivative that has been shown to be non-cariogenic in experimental animals. The purpose of this study was to determine whether certain oral bacteria could utilize sucralose. Sucralose, as a sole carbon source, was unable to support growth of ten strains of oral bacteria and dental plaque. When sucrolose was incorporated into a liquid medium containing glucose or sucrose, all organisms tested displayed similar pH falls, compared with controls. The incorporation of 126 mmol/L sucralose into glucose agar medium caused total inhibition of growth of Streptococcus sobrinus 6715-17, Streptococcus sanguis 10904, Streptococcus sanguis Challis, Streptococcus salivarius, and Actinomyces viscosus WVU627. Sucralose had no effect on IPS production. Sucralose was not bound to, nor taken up by, cells. Sucralose inhibited the formation of glucan and fructan polymers in proportion to the sucralose-to-enzyme ratio, but independent of the sucrose concentration in the assay mixture. No radioactive polymer was formed from 14C-U-sucralose added to mixtures containing glucosyltransferase (GTF) or fructosyl-transferase (FTF). Inhibition of GTF and FTF by sucralose was removed following dialysis of the enzyme/sucralose mixture. These results show that sucralose was not utilized by the oral bacteria tested and that the inhibitory effect of sucralose on GTF and FTF was non-competitive and reversible. The results further support the concept that sucralose is non-cariogenic.
Actinomyces↗
Adsorption of oral bacteria to porous type calcium carbonate.
The purpose of this study was to investigate the adsorption of [3H]-thymidine labeled oral microorganisms to porous type calcium carbonate (PCC) beads in a buffer containing human parotid saliva and to PCC combined chewing gum sheets. Adsorption rates of Streptococcus sobrinus B13 and 6715, Streptococcus mutans MT8148R and Actinomyces naeslundii T14V with PCC were significantly higher than those with calcium carbonate (CC) beads (p < 0.01). Adsorption rates of S. sobrinus, S. mutans and A. naeslundii with PCC combined chewing gum were significantly higher than those with CC combined chewing gum (p < 0.01). The present results suggested that the chewing gum containing PCC may be able to exclude oral bacteria, including cariogenic and periodontopathic bacteria, for prevention of dental caries and periodontal disease.
Actinomyces↗
Chemical compositions and antimicrobial activities of four different Anatolian propolis samples.
Propolis means a gum that is gathered by bees from various plants. It is known for its biological properties, having antibacterial, antifungal and healing properties. The aims of this study were to evaluate the antimicrobial activity of four different Anatolian propolis samples on different groups of microorganisms including some oral pathogens and comparison between their chemical compositions. Ethanol extracts of propolis (EEP) were prepared from four different Anatolian propolis samples and examined whether EEP inhibit the growth of the test microorganisms or not. For the antimicrobial activity assays, minimum inhibitory concentrations (MIC) were determined by using macrodilution method. The MIC values of the most effective propolis (TB) were 2 microg/ml for Streptococcus sobrinus and Enterococcus faecalis, 4 microg/ml for Micrococcus luteus, Candida albicans and C. krusei, 8 microg/ml for Streptococcus mutans, Staphylococcus aureus, Staphylococcus epidermidis and Enterobacter aerogenes, 16 microg/ml for Escherichia coli and C. tropicalis and 32 microg/ml for Salmonella typhimurium and Pseudomonas aeruginosa. The chemical compositions of EEP's were determined by high-temperature high-resolution gas chromatography coupled to mass spectrometry. The main compounds of four Anatolian propolis samples were flavonoids such as pinocembrin, pinostropin, isalpinin, pinobanksin, quercetin, naringenin, galangine and chrysin. Although propolis samples were collected from different regions of Anatolia all showed significant antimicrobial activity against the Gram positive bacteria and yeasts. Propolis can prevent dental caries since it demonstrated significant antimicrobial activity against the microorganisms such as Streptococcus mutans, Streptococcus sobrinus and C. albicans, which involves in oral diseases.
Anti-Infective Agents↗
[Experimental study of bacteriostatic activity of Chinese herbal medicines on primary cariogenic bacteria in vitro].
OBJECTIVE: To screen some Chinese herbal medicines for their inhibitory activity on cariogenic bacteria, and investigate their active ingredients, and measure their minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC). METHODS: Active components were isolated from every tested Chinese herbal medicine by means of aqueous extraction and ethanolic extraction. Berberine was purified from Coptis chinensis Fra. Disk agar diffusion method was employed in screening herbs with inhibiting effect on cariogenic bacteria. MIC and MBC were determined by broth dilution method. RESULTS: Against Streptococcus mutans Ingbritt, MBCs of Magnolia officinalis ethanolic extract, Berberine, Coptis chinensis Fra aqueous extract and Coptis chinensis Fra ethanolic extract were 0.488, 0.625, 7.800 and 1.950 g/L respectively. Against Streptococcus sobrinus 6715, MBCs of Magnolia extract, Coptis chinensis Fra ethanolic extract, Rhus chinensis Mill ethanolic extract and Phellodendron chinen ethanolic extract were 0.488, 0.625, 1.950, 3.900, 3.900 and 3.900 g/L respectively. Against Actinomyces viscosus ATCC 19246, MBCs of Berberine, Coptis chinensis Fra aqueous extract, Coptis chinensis Fra ethanolic extract, Rheum palmatum L aqueous extract and Rheum palmatum L ethanolic extract were 1.250, 3.900, 3.900, 15.600 and 31.250 g/L respectively. CONCLUSIONS: Magnolia officinalis, Coptis chinensis Fran, Rheum palmatum L aqueous extracts exhibit strong inhibition on cariogenic bacteria. Magnolia officinalis ethanolic extract has the strongest bactericidal effects on Streptococcus mutans and Streptococcus sobrinus.
Cariogenic Agents↗
Glucosyltransferase production by Streptococcus sanguis Challis and comparison with other oral streptococci.
Glucosyltransferase (GTF) activity in batch cultures of Streptococcus sanguis strain Challis in defined medium was maximum at peak growth and declined rapidly on further incubation into stationary phase. Activity was present in spent culture medium and was associated with 2 polypeptides of approximate Mrs 170 kDa (90% of activity) and 155 kDa (about 10% of activity). Other S. sanguis strains produced similar antigenically-related polypeptides but with varying activities, and antibodies to the S. sanguis enzyme did not react with enzymes produced by other species of oral streptococci. Increasing the [Na+] reduced GTF production by S. sanguis strains Challis and NCTC 7865, and by Streptococcus sobrinus, but not by Streptococcus salivarius. Addition of Tween 80 to cultures caused a rapid inhibition of GTF synthesis by strain Challis, but not by strain NCTC 7865. Benzyl alcohol had a slower and less inhibitory effect on GTF production by strain Challis. Tween 80 generally inhibited to varying degrees GTF production by other oral streptococci through in S. sobrinus it was stimulated. The results suggest controls operating on GTF synthesis and secretion by different strains of oral streptococci appear to be diverse.
Animals↗
Specificity of the glucan-binding lectin of Streptococcus cricetus.
The specificity of the glucan-binding lectin (GBL) of Streptococcus cricetus AHT was determined. Examination of the kinetics of aggregation of cell suspensions with glucans containing various percentages of alpha-1,6, alpha-1,4, alpha-1,3, and alpha-1,2 anomeric linkages revealed that only glucans with at least 80% alpha-1,6 linkages promoted strong aggregation. Moreover, only linear glucans with molecular weights greater than 5 X 10(5) were capable of causing rapid aggregation of the bacteria. The lectin was observed to be present on S. cricetus strains, on Streptococcus sobrinus, and on several Streptococcus mutants strains. Preincubation of suspensions of S. cricetus AHT with glucan T10 (molecular weight of 10,000) before the addition of high-molecular-weight glucan resulted in competitive inhibition in a concentration-dependent manner. Inhibition was achieved also with isomaltopentaose, isomaltohexaose, and isomaltooctaose, but at higher concentrations than glucan T10. In contrast, no inhibition was observed with maltoheptaose, providing additional evidence for the specificity of GBL. Treatment of suspensions of S. cricetus AHT with trypsin before and after aggregation with high-molecular-weight glucan revealed a substantial level of protection of GBL when in a bound state. Collectively, these results indicated that GBL has an absolute affinity for glucans rich in alpha-1,6 linkages and possesses an active site which recognizes internal sequences and accommodates isomaltosaccharides of at least nine residues. This unusual specificity may contribute to the colonization of S. cricetus, S. sobrinus, and S. mutans in glucan-containing plaque in the oral cavity.
Agglutination↗
Microbial induction of dentinal caries in human teeth in vitro.
The ability of Lactobacillus casei, Streptococcus sobrinus, Actinomyces viscosus, and Streptococcus salivarius to induce dental caries was determined in vitro. A class I cavity (depth: 2mm) was prepared in extracted human caries-free premolars to make dentin blocks. The blocks were inoculated with these four bacterial strains in a monoinfective fashion and were incubated under anaerobic conditions. In addition to the monoinfection groups, mixed-infection groups of L. Casei with S. sobrinus or A. viscosus were also prepared. Half of the culture medium was renewed every 3 days, and the pH of the medium was measured. After 4 or 12 wk, these dentin blocks were prepared by Brown-Brenn staining and by contact microradiography for light microscopic observation and for immunohistochemical staining. The final pH of the S. salivarius group was the highest among the experimental groups, at approximately 5.1; that of the others was approximately 4.3. Bacterial invasion into the dentinal tubules was observed in all but the S. salivarius group. Among the monoinfection groups, the S. sobrinus group showed the highest invasion rate, followed by the A. Viscosus group and the L. casei group. The invasion rate was also high in the mixed-infection groups. Immunohistochemical staining revealed invasion only by L. casei, and not by S. sobrinus and A. viscosus. The invasion rate by L. casei was higher in the mixed-infection group with either S. sobrinus or A. viscosus than in the monoinfection groups. These findings suggest that lactobacillus might play an important role in the initiation and progress of dentinal caries, and that this bacterial species might exhibit a cooperative cariogenicity when it coexists with other bacterial species that surpasses its individual cariogenicity.
Actinomyces viscosus↗