Synthetic brominated furanone F202 prevents biofilm formation by potentially human pathogenic Escherichia coli O103:H2 and Salmonella ser. Agona on abiotic surfaces.

AIMS: Investigate the use of a synthetic brominated furanone (F202) against the establishment of biofilm by Salmonella ser. Agona and E. coli O103:H2 under temperature conditions relevant for the food and feed industry as well as under temperature conditions optimum for growth. METHODS AND RESULTS: Effect of F202 on biofilm formation by Salmonella ser. Agona and E. coli O103:H2 was evaluated using a microtiter plate assay and confocal microscopy. Effect of F202 on bacterial motility was investigated using swimming and swarming assays. Influence on flagellar synthesis by F202 was examined by flagellar staining. Results showed that F202 inhibited biofilm formation without being bactericidal. F202 was found to affect both swimming and swarming motility without, however, affecting the expression of flagella. CONCLUSIONS: F202 showed its potential as a biofilm inhibitor of Salmonella ser. Agona and E. coli O103:H2 under temperature conditions relevant for the feed and food industry as well as temperatures optimum for growth. One potential mode of action of F202 was found to be by targeting flagellar function. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study gives valuable new knowledge to the potential use of furanones as a tool in biofilm management in the food and feed industry.

Inter-individual variation in the plaque formation rate of young individuals.

OBJECTIVES: The aim of the present study was to describe the inter-individual variation in the plaque formation rate of 38 young adults. METHODS: The plaque formation rate was estimated by employing a quantitative plaque percent index (P% index). A substantial inter-individual variation in the plaque formation rate was observed. The possible contributions of stimulated salivary flow rate, buffer capacity, diet and smoking to the variation in plaque formation were estimated by regression analysis. RESULTS: The control variables explained only 2.5% of the variance in the plaque formation rate. Neither associations nor the total model were significant. The present method of measuring plaque presented as a simple and time-effective procedure. CONCLUSIONS: It is suggested that the observed variation in the plague formation rate between the young individuals can be regarded as a biological function which is possibly an inherent individual characteristic. Studies with larger sample sizes are required to confirm the findings of the present study.

Involvement of antigen I/II surface proteins in Streptococcus mutans and Streptococcus intermedius biofilm formation.

BACKGROUND/AIM: Dental diseases are caused by microorganisms organized in biofilms. Streptococcus mutans and Streptococcus intermedius are commensals of the human oral cavity. S. mutans is associated with caries, whereas S. intermedius is associated with purulent infections. Oral streptococci including S. mutants and S. intermedius express a family of surface proteins termed antigen I/II (Ag I/II). Ag I/II is implicated in adhesion; however, its role in biofilm formation has not yet been investigated. METHODS: By using isogenic Ag I/II-deficient mutants of S. mutans and S. intermedius we studied the influence of Ag I/II on in vitro biofilm formation. Biofilm was quantified in polystyrene microtiter plates and visualized by scanning electron microscopy. Ag I/II expression in planktonic and biofilm cells, as well as in the presence or absence of saliva was investigated by immunoblotting. RESULTS: In the presence of saliva, the Ag I/II-deficient mutants formed 65% less biofilm than the wild-types. In the absence of saliva, no difference was observed in S. mutans, whereas the S. intermedius Ag I/II mutant formed 41% less biofilm. Ag I/II expression was reduced in the presence of saliva. No differences in expression were observed between biofilm and planktonic cells. CONCLUSION: The results indicated that Ag I/II may be important during biofilm formation particularly in the presence of saliva. These findings may provide useful information regarding the importance of Ag I/II in biofilm formation and in the search of new strategies to control biofilm-mediated infections.

Functional variation of the antigen I/II surface protein in Streptococcus mutans and Streptococcus intermedius.

Although Streptococcus intermedius and Streptococcus mutans are regarded as members of the commensal microflora of the body, S. intermedius is often associated with deep-seated purulent infections, whereas S. mutans is frequently associated with dental caries. In this study, we investigated the roles of the S. mutans and S. intermedius antigen I/II proteins in adhesion and modulation of cell surface characteristics. By using isogenic mutants, we show that the antigen I/II in S. mutans, but not in S. intermedius, was involved in adhesion to a salivary film under flowing conditions, as well as in binding to rat collagen type I. Binding to human fibronectin was a common function associated with the S. mutans and S. intermedius antigen I/II. Adhesion of S. mutans or S. intermedius to human collagen types I or IV was negligible. Hydrophobicity, as measured by water contact angles, and zeta potentials were unaltered in the S. intermedius mutant. The S. mutans isogenic mutants, on the other hand, exhibited more positive zeta potentials at physiological pH values than did the wild type. The results indicate common and species-specific roles for the antigen I/II in mediating the attachment of S. mutans and S. intermedius to host components and in determining cell surface properties.

Expression and functional properties of the Streptococcus intermedius surface protein antigen I/II.

Streptococcus intermedius is associated with deep-seated purulent infections. In this study, we investigated expression and functional activities of antigen I/II in S. intermedius. The S. intermedius antigen I/II appeared to be cell surface associated, with a molecular mass of approximately 160 kDa. Northern blotting indicated that the S. intermedius NCTC 11324 antigen I/II gene was transcribed as a monocistronic message. Maximum expression was seen during the early exponential phase. Insertional inactivation of the antigen I/II gene resulted in reduced hydrophobicity during early exponential phase, whereas no effect was detected during mid- and late exponential phases. Binding to human fibronectin and laminin was reduced in the isogenic mutant, whereas binding to human collagen types I and IV and to rat collagen type I was not significant for either the wild type or the mutant. Compared to the wild type, the capacity of the isogenic mutant to induce interleukin 8 (IL-8) release by THP-1 monocytic cells was significantly reduced. The results indicate that the S. intermedius antigen I/II is involved in adhesion to human receptors and in IL-8 induction.

Genetic transformation in Streptococcus mutans requires a peptide secretion-like apparatus.

Competence for genetic transformation in Streptococcus pneumoniae and Streptococcus gordonii involves the ComAB secretion apparatus, which is thought to export the competence-stimulating peptide. Homologous secretory systems are also used for the export of certain bacteriocins and bacteriocin-like peptides. In this study, a similar secretory apparatus was found in the Streptococcus mutans genome, and its role in transformation was investigated. Gene inactivation resulted in a mutant deficient in transformability. We suggest that secretion of a peptide, possibly the competence-stimulating peptide itself, is involved in competence induction also in S. mutans.

Effects of mouth rinses with xylitol and fluoride on dental plaque and saliva.

The aim of this study was to test the hypothesis that xylitol, alone and in combination with fluoride, affects the salivary flow rate and micro-biota, dental plaque accumulation, gingivitis development, and the acidogenic potential of plaque. Three groups, each of 10 subjects, rinsed for 1 min 3 times daily over two 4-week periods, first with 10 ml water (control), and thereafter with either 0.05% NaF, 40% xylitol, or with 0.025% NaF plus 20% xylitol according to a double-blind controlled design. They performed habitual mechanical tooth cleaning during the first 2 weeks of each period but abstained from interdental cleaning during the final 2 weeks. While mouth rinsing was continued, all mechanical oral hygiene was discontinued the last 2 days of each period to permit plaque accumulation. The last mouth rinse was administered in the clinic before the final examination. The following parameters were assessed: (1) unstimulated and paraffin-stimulated salivary secretion rates; (2) salivary micro-biota; (3) plaque index; (4) papillar bleeding; (5) plaque pH response to sucrose, and (6) lactate formation by dental plaque. No statistically significant differences in any of the parameters were found. In conclusion, three daily mouth rinses with fluoride and xylitol, separately or in combination, did not affect the salivary flow rate or micro-biota, dental plaque accumulation, gingivitis development, or the acidogenic potential of plaque.

The effects of xylitol-containing chewing gums on dental plaque and acidogenic potential.

The aim of this study was to test the hypothesis that the chewing of xylitol- or xylitol/sorbitol-containing chewing gum reduces plaque formation and the acidogenic potential of dental plaque. Thirty healthy volunteers aged from 19 to 28 yrs were randomly allocated to one of three test groups, chewing either xylitol-, xylitol/sorbitol-, or sucrose-sweetened gums. A three-day plaque accumulation period of no oral hygiene was instituted prior to and at the termination of the chewing gum program, which lasted 33 days. Plaque quantity was assessed on the basis of protein content of individual plaque samples collected by a standardized technique. Acidogenic potential of individual baseline and test plaque samples was assessed by the quantity of various organic acids formed from D-(U-14C)glucose. Identification of extracellular and intracellular metabolites was performed by HPLC. Statistical evaluation of data was performed according to paired comparisons of individual baseline and post-chewing data. Plaque formation, acidogenic potential, and glycolytic profiles were similar at baseline and after the gum-chewing periods. Also, there was no intracellular accumulation of glycolytic metabolites within the plaque bacteria to indicate the inhibition of glycolysis. The study thus leads to the conclusion that, in young adults with low caries experience, exposure of the oral cavity to acceptable doses of xylitol or xylitol and sorbitol has no effect on the microbial deposits on the teeth.

Xylitol in caries prevention: what is the evidence for clinical efficacy?

Xylitol has attracted much attention as an alternative sweetener. Essentially all clinical studies concerning the effect of xylitol on caries development consent to its non-cariogenicity and to the beneficial effect of substituting sucrose with xylitol in chewing gums and sweets. However, claims of anti-caries or therapeutic effects, and superiority of xylitol over other polyols are still to be confirmed by well designed and conducted studies from independent research groups.

Detection of microbial-derived fatty acids in carious dentin by gas chromatography and gas chromatography-mass spectrometry.

The aim of the present study was to analyze the fatty acid content of carious and sound human dentin. Gas chromatography and gas chromatography-mass spectrometry revealed the presence of fatty acids of C10-C18 size in the carious dentin, whereas fatty acids of C16 size were present in minute amounts in three samples of the corresponding sound dentine controls. No fatty acids were detected in the other sound dentin control samples. The source of fatty acids was considered to be microorganisms invading the dentin during the progression of the caries lesion. The presence of bacterial fatty acids in carious dentin may serve as a marker for the pathological process and thus contribute to the understanding of the mechanisms involved.

The effects of a mouthrinse containing sodium benzoate and alcohol on the quantity and metabolism of dental plaque.

The aim of this study was to investigate the efficacy of a pre-brushing rinse containing sodium benzoate and alcohol, on 2 week old dental plaque. In a double-blind, placebo-controlled crossover study 20 individuals abstained from oral hygiene for periods of 2 weeks. Ten individuals were then assigned to rinse once for 30 seconds with either the test mouthrinse or a placebo mouthrinse. Following this, all individuals had their teeth cleaned and maintained proper oral hygiene for four weeks before the experiment was repeated. This time, the control individuals were assigned to the test rinse and vice versa. Using the Plaque Index on three buccal sites on all teeth, plaque was recorded before and after rinsing, as well as following a subsequent toothbrushing. For metabolic studies plaque from the lingual surfaces on the teeth in one quadrant in the upper and the lower jaw was sampled before rinsing and from the two remaining quadrants following the rinse, and analysed. There was no difference in Plaque Index between the two experimental periods. An oral rinse with the test mouthrinse had no effect on the amount of plaque, nor did it enhance the subsequent plaque removal after brushing. The independent biochemical analysis showed a significant effect of rinsing on the glycolytic potential, but with no effect on the glycolytic profile, which was similar for test and control plaque samples. Thus, a single oral rinse with a mouthrinse containing sodium benzoate and alcohol does not affect removal of plaque nor does it seems to influence the glycolytic potential of the plaque.

Effects of mouthrinses with chlorhexidine and zinc ions combined with fluoride on the viability and glycolytic activity of dental plaque.

Inhibition of plaque acidogenicity by a mouthrinse with chlorhexidine (CHX) or zinc ions has been ascribed to a prolonged bacteriostasis due to substantive properties of the agents. The present aim was to study the effects of mouthrinses with CHX and Zn ions combined with fluoride on the viability and glycolytic activity of dental plaque in order to assess the bacteriostatic versus possible bactericidal effects. Following 2 d of plaque accumulation, 4 groups of 10 students rinsed with either 12 mM NaF (F), 0.55 mM CHX diacetate+F (F-CHX), 10 mM Zn acetate+F (F-Zn), or with the three agents in combination (F-CHX-Zn). Plaque samples were collected before and 90 min after mouthrinsing. Thereafter, the in vivo plaque pH response to sucrose was monitored in each student using touch microelectrodes. F-CHX and F-CHX-Zn reduced the in vivo pH fall significantly as compared with F, whereas F-Zn exerted a non-significant inhibition. Pooled pre- and post-rinse plaque samples were used to measure the pH fall during fermentation of [14C]-glucose, and the glycolytic profiles were analyzed by HPLC. Bacterial viability was assessed by counting the colony-forming units (CFU). All mouthrinses except F reduced glucose consumption and acid formation and thus the pH fall. F-CHX reduced the CFU equal to the reduction of glucose consumption, indicating that inhibition of plaque acidogenicity was due to a bactericidal rather than a bacteriostatic effect. F and F-Zn did not reduce the CFU, thus F-Zn decreased glucose metabolism without affecting plaque viability. F-CHX-Zn reduced both the CFU and glucose metabolism of surviving plaque microorganisms.

Effects of chlorhexidine-fluoride mouthrinses on viability, acidogenic potential, and glycolytic profile of established dental plaque.

Inhibition of dental plaque acidogenicity by chlorhexidine (CHX) mouthrinses has been ascribed to a long-lasting bacteriostatic effect due to binding of CHX to oral surface structures combined with a slow release rate from the binding sites. The present aims were to study the effects of CHX-containing mouthrinses on the viability and glycolytic activity of established plaque in order to assess the bactericidal versus the bacteriostatic effects. Following 2 days of plaque accumulation, three groups of 10 students rinsed with either 12.0 mM NaF, 0.55 mM CHX plus NaF, or with 2.2 mM CHX plus NaF. Plaque samples were collected before and 90 min after mouthrinsing. The pH in pooled pre- and post-rinse plaque samples was recorded before and up to 10 min after the addition of D-[U-14C]glucose. Total colony-forming units in each sample were determined. High-performance liquid chromatography analyses showed lactate to be the major extracellular glycolytic metabolite in all samples. CHX-NaF markedly reduced the colony-forming units, the pH fall from fermentation of glucose, as well as glucose consumption and lactate formation, whereas NaF alone exhibited no such effects. The reduction of glucose consumption by the CHX-NaF mouthrinses corresponded to the reduction of colony-forming units, indicating no bacteriostatic effect. The plaque pH in vivo was monitored in each student 90 min after mouthrinsing with the test solutions prior to and up to 1 h after a sucrose mouthrinse using touch microelectrodes. The CHX-NaF mouthrinses reduced the fall in pH significantly (p < 0.05) as compared with the NaF mouthrinse.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanisms of dental plaque formation.

Much effort has been placed on elucidating the diverse mechanisms of microbial adhesion to tooth surfaces. Both specific and non-specific types of adhesion have been envisaged. Pioneer colonizers represent a selected part of the oral microflora, and it has been assumed that specific adhesin-receptor interactions between the microbial surface and the pellicle account for this specificity. Whereas microbial adhesion to tooth surfaces is a general prerequisite for initiation of plaque formation, microbial multiplication is probably the dominant feature in the build-up of dental plaque. Local environmental factors which influence the establishment and composition of the ultimate plaque community are therefore of greater importance than initial adhesion per se. The highly individual and site-related characteristics of the plaque flora illustrate the selective power of the environment. Environmental conditions are not uniform. Thus, each site represents its own conditions are not uniform. Thus, each site represents its own distinct ecosystem, and the microbial composition at the site depends on the outcome of a variety of host-microbial and microbial-microbial interactions. The relative in vivo significance of these interactions is difficult to assess.

The effect of mineral-derived zinc ions on in vitro glucose metabolism of Streptococcus mutans NCTC 10449.

Mouthrinses containing zinc ions inhibit plaque acidogenicity, but the effect is transient. Zinc-containing apatite or zinc phosphate precipitated within dental plaque might serve as a reservoir for zinc ions, thus providing prolonged inhibition of acid formation. Zinc-containing fluorhydroxyapatite was prepared from solutions containing CaCl2, KH2PO4, NaF and increasing amounts of ZnCl2 (0.0, 0.005, 0.02, 0.1, 0.2 or 1 mM; minerals No. 1-6, respectively) by raising the pH with ammonia. Zinc phosphate tetrahydrate (mineral No. 7) was prepared in a similar manner from a solution containing ZnCl2 and KH2PO4 only. Dense cell suspensions of Streptococcus mutans NCTC 10449 were incubated with 14C-glucose and one of the test minerals (No. 1-7). Glycolysis was allowed to proceed, with or without pH control, in a pH-stat. Samples were withdrawn at 1, 2, and 3 min, and extracellular glycolytic metabolites were identified by HPLC. Mineral No. 7 inhibited glycolysis and any pH fall almost completely. With the pH fixed at 5.5, reduction of glucose consumption and lactate formation was 83 and 93%, respectively, compared to the no-zinc control mineral (No. 1). No changes in glucose consumption or lactate formation were evident in the presence of minerals No. 2-6. All apatitic minerals had a buffering effect and, in the absence of pH control, glycolysis was increased due to the higher pH. Detectable levels of fluoride were not released by any mineral into the incubation mixture, and zinc only by minerals No. 6 and 7 in greater than trace amounts.(ABSTRACT TRUNCATED AT 250 WORDS)

In vivo effects of fluoride, chlorhexidine and zinc ions on acid formation by dental plaque and salivary mutans streptococcus counts in patients with irradiation-induced xerostomia.

Irradiation therapy including major salivary glands may result in xerostomia and enhanced susceptibility to dental caries. The present aim was to assess the ability of mouthrinses with F-, Zn2+, and chlorhexidine (CH), in various combinations, to reduce acidogenic potential of dental plaque and salivary mutans streptococcus counts (SMSC) in 7 patients with xerostomia secondary to irradiation. The patients rinsed twice daily for 3 weeks with the following test solutions: (1) 12 mmol/l NaF (F; control), (2) NaF + 20 mmol/l ZnCl2 (F-Zn), and (3) NaF + 1.1 mmol/l CH (F-CH). Resting periods (F) of varying lengths were incorporated. Acid formation by dental plaque was monitored as plaque pH response to a sucrose mouthrinse, at the end of each test period, 4 h after mouthrinsing with test solution. Plaque pH was measured repeatedly at 2-8 sites in each patient before, and up to 60 min after the sucrose mouthrinse using touch microelectrodes. SMSC were determined using Dentocult SM-Strip mutans. Compared with F, F-CH significantly (P < or = 0.02) reduced acid formation by plaque and SMSC, whereas F-Zn did not affect acid formation or SMSC significantly. Pilot experiments in 4 patients showed mouthrinses with NaF + 0.55 mmol/l CH + 10 mmol/l Zn2+ to be ineffective, whereas NaF + 2.2 mmol/l CH was highly effective, but no better than F-CH. Twice daily mouthrinses with 12 mmol/l NaF in combination with 1.1 mmol/l CH may be an effective regimen to prevent post-irradiation caries.

Effects of food preservatives on growth and metabolism of plaque bacteria in vitro and in vivo.

The aims of the present study were to assess the consumption of food preservatives during the last decades, and to study the effect of the preservatives, sorbic and benzoic acid, on growth and glycolysis of oral bacteria in vitro, and on acid formation by dental plaque in vivo. Five consumption reports from the Central Bureau of Statistics of Norway were used to estimate alterations in consumption of staple food containing the two preservatives. A modified broth dilution method was used to determine the MIC values of the preservatives against Streptococcus sobrinus and Streptococcus sanguis. Extracellular 14C-glycolytic metabolites were studied by HPLC analyses. Plaque-pH measurements were used to assess possible effects on acid production. The consumption reports indicated increased consumption of preservatives. The in vitro testing suggested that legal concentrations of preservatives may inhibit the growth of oral streptococci. However, the preservatives did not inhibit in vitro glycolysis at tested concentrations. In vivo testing with similar concentrations (0.4% w/v) showed a significant effect. A higher concentration (2% w/v potassium sorbate) had a tendency to inhibit acid-formation by dental plaque even more.

Use of palladium touch microelectrodes under field conditions for in vivo assessment of dental plaque pH in children.

The aim of this study was to assess the applicability of palladium touch microelectrodes, connected to battery-run pH meters, for in vivo plaque pH measurements in children. The pH was assessed in 20 7-year-old and in 19 14-year-old caries-active and caries-inactive rural Kenyan children. The resting pH was measured at non-carious interproximal and occlusal sites and in open dentine cavities. Independent repeated measurements were performed at given sites at intervals of 15 s and 5 min and on different days. The resting plaque pH varied widely among the children, and there was no significant difference between caries-active and caries-inactive groups. The most striking feature was the considerable erratic fluctuations of pH at a given site with time, both in resting and in sucrose-challenged plaque. These fluctuations were sensitively recorded by palladium touch microelectrodes. After a sucrose rinse, not all sites in the same mouth behaved in a similar fashion, and thus the classical 'Stephan curve' was not always apparent. In conclusion, the palladium touch microelectrodes are highly applicable for plaque pH measurements in children, even under extreme field conditions.

Effect of sugarcane chewing on plaque pH in rural Kenyan children.

In 5 rural Kenyan children, the effect of sugarcane chewing on plaque pH was compared with the effect of a mouthrinse with 10% sucrose at various intraoral sites. They all had poor oral hygiene and at least two carious cavities in occlusal surfaces of molars. pH measurements were conducted under field conditions using paladium touch microelectrodes connected to a battery-operated pH meter. There was a marked difference in pH response of non-carious approximal sites between maxilla and mandible, with the lowest values in the maxilla. However, the pH recovery following the instantaneous drop occurred in parallel even if most pH values had not returned to baseline values 30 min after the sucrose rinse. Following the sugarcane chewing, the pH fall was less pronounced on all sites, and within 5-10 min the values had returned to resting pH and even exceeded this. In carious cavities, a similar pattern was observed, although the acidity in these sites was more pronounced, also reflected in a lower resting mean pH. The main conclusion from this study is that sugarcane chewing yields a less pronounced pH drop and a quicker pH recovery in dental plaque than is seen following a mouthrinse with 10% sucrose. This difference probably results from stimulation of salivary flow associated with the chewing.

The effect of sucrose on plaque pH in the primary and permanent dentition of caries-inactive and -active Kenyan children.

The hypothesis that the Stephan pH responses of dental plaque would be different in caries-active and -inactive individuals was tested in 20 seven-year-old and 19 14-year-old Kenyan children. In each age group, half the children had greater than or equal to 2 dentin cavities; the other half had no such lesions. With a palladium-touch microelectrode, interdental plaque pH was monitored between m1/m2 in each quadrant in the primary dentition and in the four molar/premolar regions in the permanent dentition. pH was also monitored in caries cavities in the occlusal surfaces of lower first molars and on the tongue. pH was measured before and up to 60 min after the children rinsed with 10 mL of 10% sucrose. Caries status of the individual was unrelated to plaque pH in comparable non-carious sites in both of the age groups. The pH minimum in the maxilla was about 0.5 pH units lower than that in the mandible. Active occlusal caries lesions had a resting pH value of about 5.5, about 1 pH unit lower than that of sound surfaces. The pH dropped to about 4.5 in caries lesions and recovered slowly. In sound occlusal sites, a pH drop to about 6.0 was followed by a relatively rapid return to the resting value. Thus, when the mean values were considered, the classic Stephan curve response was evident. However, when the pH changes at single sites were considered at various time intervals, a substantial, erratic fluctuation was observed.(ABSTRACT TRUNCATED AT 250 WORDS)