Differential Modulation of Saliva-Derived Microcosm Biofilms by Antimicrobial Peptide LL-31 and D-LL-31.

Microbiome modulation, aiming to restore a health-compatible microbiota, is a novel strategy to treat periodontitis. This study evaluated the modulation effects of antimicrobial peptide LL-31 and its D-enantiomer (D-LL-31) on saliva-derived microcosm biofilms, spiked with or without Porphyromonas gingivalis. To this end, one-day-old biofilms were incubated for 24 h with biofilm medium alone, or medium containing 40 µM LL-31 or D-LL-31, after which biofilms were grown for 5 days. Biofilms were assessed at 1 day and 5 days after intervention for the total viable cell counts, dipeptidyl peptidase IV (DPP4) activity, P. gingivalis amount (by qPCR) and microbial composition (by sequencing). The results showed that D-LL-31, not LL-31, significantly reduced the total viable cell counts, the P. gingivalis amount, and the DPP4 activity of the biofilms spiked with P. gingivalis, but only at 1 day after intervention. In the biofilms spiked with P. gingivalis, D-LL-31 tended to reduce the α-diversity and the compositional shift of the biofilms in time as compared to the control and LL-31 groups. In conclusion, D-LL-31 showed a better performance than LL-31 in biofilm modulation. The biofilm modulation function of the peptides could be impaired when the biofilms were in a severely dysbiotic state.

Are the counts of Streptococcus mutans and Staphylococcus aureus changed in complete denture wearers carrying denture stomatitis? A systematic review with meta-analyses.

STATEMENT OF PROBLEM: Despite the importance of Candida spp. on the etiology of denture stomatitis (DS), information on the role of the bacterial component is still scarce. PURPOSE: The purpose of this systematic review was to evaluate whether the counts of Staphylococcus aureus and Streptococcus mutans were changed in complete denture wearers diagnosed with Candida-associated DS. MATERIAL AND METHODS: The literature search was performed in 8 databases and by hand searching. The risk of bias was assessed according to the Newcastle-Ottawa qualifier. Meta-analyses were performed considering the microorganism evaluated (S. aureus or S. mutans) and the collection area (mucosa or dentures). The certainty of evidence was assessed according to the grading of recommendations assessment, development and evaluations (GRADE) criteria. RESULTS: Participants with DS presented higher counts of S. aureus in the mucosa compared with those from the control group (OR, 3.16 [1.62, 6.15]; P<.001). No significant difference between the groups was observed for samples collected from dentures (OR, 0.73 [0.50, 1.07]; P=.110). Conversely, participants without DS presented higher counts of S. mutans both in the mucosa (OR, 0.19 [0.06, 0.63]; P=.006) and dentures (OR, 0.64 [0.41, 1.0]; P=.050). CONCLUSIONS: Microbial counts in participants with DS changed as a function of the type of microorganism and collection site. The certainty of evidence ranged from very low to low. The findings reinforce the fact that bacteria also play a relevant role in DS and should be more extensively studied. Such information may be useful to guide further therapies to prevent or control DS.

Cytotoxicity and anti-biofilm properties of novel hybrid-glass-based caries infiltrant.

OBJECTIVES: To assess the cytotoxicity of an experimental hybrid-glass-based infiltrant and its effect on biofilm attachment, growth and metabolic activity, and to compare it to the resin-based infiltrant Icon. METHODS: Cytotoxicity of hybrid-glass-based material (EXP) and resin-based infiltrant Icon (Icon) was tested in direct contact tests on freshly cured (direct_mat) and on materials kept for 24 h in cell culture medium (direct_exmat), and extract test with materials 24-h extracts (extract). Cell viability of L929 mouse fibroblast cell line was measured with MTT assay, according to ISO10993-5:2009. Biofilm attachment (5 h), growth (24 h and 48 h) and lactic-acid production (24 h and 48 h) on glass-disk specimens coated with EXP or Icon, or uncoated (control), were assessed using a microcosm biofilm model and Amsterdam Active Attachment system. At indicated time points, biofilms were harvested, plated, and CFU counts were determined, while lactic-acid production was measured colorimetrically. RESULTS: Cell viability reduction by EXP was below 30%-threshold in direct contact tests, while in extract test an increased cell viability was observed. Icon reduced cell viability substantially in all three tests. Significantly less bacteria attached to the surface of EXP after 5 h compared to Icon and control. Biofilm growth was significantly lower on EXP than on Icon and control after 24 h, but this difference was smaller and statistically insignificant after 48 h. There was no difference in lactic-acid production among groups. SIGNIFICANCE: Novel hybrid-glass-based infiltrant seems to have a better biocompatibility and accumulates on its surface less bacteria than resin-based infiltrant, which makes it an attractive resin-free alternative.

Effects of Surface Prereacted Glass on Saliva-Derived Polymicrobial Biofilms in an Active Attachment Biofilm Model.

Bioactive restorative materials are being developed to either influence the de/remineralization balance of the dental hard tissues locally or to release components that interact with the oral microbiota. Surface prereacted glass (S-PRG, Shofu, Japan) is a material that may influence both processes. S-PRG releases fluoride, which can interact with the de/remineralization process, and a range of other compounds that may influence the oral microbiota. In the current study, several experiments were performed to investigate the potential of S-PRG to influence both the growth and lactic acid production of saliva-derived polymicrobial biofilms. Biofilm formation was studied using the Amsterdam Active Attachment model. An eluate of the S-PRG particles was tested by adding it to the growth medium or by exposing the biofilms to it for 1 h. The effect of S-PRG particles was tested by adding the particles to the growth medium. The current experiments showed that the presence of S-PRG eluate in the medium influenced biofilm growth and lactic acid production even at low concentrations. The composition of the biofilms changed in the presence of S-PRG eluate, even at concentrations of S-PRG eluate at which biofilm viability was not affected. Treatment of developing biofilms with S-PRG eluate did neither show an effect on biofilm viability nor on lactic acid production. The addition of S-PRG particles to the growth medium resulted in both a lower biofilm viability and lower lactic acid production, indicating that the release of ions from the particles was fast enough to influence biofilm formation. From the current experiments, it can be concluded that S-PRG has the potential to influence biofilm growth, but the presence of the released ions during biofilm formation is required to show an effect.

Effects of sodium hexametaphosphate microparticles or nanoparticles on the growth of saliva-derived microcosm biofilms.

OBJECTIVES: This study evaluated the effects of sodium hexametaphosphate microparticles (HMPmicro) or nanoparticles (HMPnano) on the growth of saliva-derived microcosm biofilms MATERIALS AND METHODS: Saliva-derived biofilms were formed on glass coverslips for 24 h. Thereafter, Streptococcus mutans (C180-2) was incorporated or not into the biofilms. From that time point onwards, solutions containing 0.2% HMPmicro or HMPnano, combined or not with 220 ppm F, were constantly present in the culture medium. In addition, 220 ppm F alone (220F) and McBain medium without any compound were also tested as positive and negative controls (CTL), respectively. After 96 h, the biofilms were plated on anaerobic blood agar or sucrose agar bacitracin for total and S. mutans CFU-counting, respectively. Biofilms' lactic acid production was analysed spectrophotometrically. Data were submitted to ANOVA or Kruskal-Wallis' tests, followed by Student-Newman-Keuls' test (p<0.05; n=12). RESULTS: HMPmicro or HMPnano led to significantly lower lactic acid production, and significant reductions in total CFU-counting in microcosm biofilms, supplemented or not with S. mutans, in comparison to both controls, with significant differences between 220F and CTL. No significant differences were observed among the groups treated with HMPmicro or HMPnano (with or without F). The same trend was seen for S. mutans CFU-counting, in biofilms supplemented with S. mutans. CONCLUSIONS: HMP significantly reduced total and S. mutans CFU counts, as well as lactic acid production by saliva-derived microcosm biofilms. CLINICAL RELEVANCE: These findings in saliva-derived microcosm biofilms suggest that HMP stands as a promising alternative for the control of cariogenic biofilms.

Effect of high-fluoride dentifrice on root dentine and bacterial composition in a multispecies biofilm model.

The present study evaluated the effect of high-fluoride dentifrice on dentine demineralization and bacterial composition in a multispecies biofilm model in vitro. A seven-organism bacterial consortium was grown on bovine dentine discs in a high-throughput active attachment model. The biofilms were submitted twice per day to the following dentifrices treatments: 5,000 ppm F, 1,100 ppm F, with placebo as a negative control. After 5 days of biofilm growth, dentine samples were assessed by transversal microradiography, the biofilm was collected for bacterial counts and the pH of the media was determined. Lower integrated mineral loss values were observed when 5,000 ppm F-treatment was used compared to the other treatments. Overall microbiological counts decreased with increasing F-concentration as well the pH of the media throughout the experiment. The 5,000 ppm F-treatment caused a shift in microbial composition and reduced dentine demineralization in the in-vitro experimental model.

The Effect of Various Fluoride Products on Dentine Lesions during pH-Cycling.

This study compared the effect of topically applied fluoride products on dentine lesions in an in vitro experiment. Demineralized bovine dentine specimens were treated once with either SDF solution (35,400 ppm F), NaF varnish (22,600 ppm F), TiF4 solution (9,200 ppm F), SnF2 gel (1,000 ppm F), no treatment (control), or preserved as baseline lesions. After the application and subsequent removal of the fluoride products, the specimens were subjected to pH-cycling. Calcium loss and uptake in the de- and remineralization buffers were assessed daily. Fluoride release into the buffers was analyzed on days 1, 2, 3, 5, 8, and 13. After the pH-cycling period, mineral distribution throughout the lesion depth was analyzed using transversal microradiography (TMR). X-ray energy-dispersive spectroscopy (EDS) examined the deposition of silver, titanium, and tin after application of SDF, TiF4, and SnF2, respectively. Overall, calcium loss and uptake analysis in the de- and remineralization buffers revealed that the SDF product was the most effective in inhibiting lesion progression, followed by the TiF4, NaF, and SnF2 products. Fluoride analysis disclosed a steep reduction of the amount of fluoride released into de- and remineralization buffers with time. The fluoride effects on de- and remineralization continued beyond the days that fluoride was released into the buffers. TMR analysis showed significant remineralization in the outer zone of the dentine lesions for all fluoride products, with SDF giving hypermineralization in this zone. In the inner zone, lesions developed in all fluoride groups, with the smallest in the SDF group. EDS showed silver and titanium deposition in depth up to 85 µm and 8 µm, respectively, while no tin deposition was observed. The silver in the dentine lesions did not contribute significantly to the density of the TMR profiles in the SDF group. In conclusion, all topical fluoride products protected the dentine lesions against lesion progression, but at different degrees. SDF showed a superior effect in protection against further demineralization and enhancement of remineralization. This was probably attributed to its fluoride concentration that was the highest among the fluoride products.

Comparison of Red-Complex Bacteria Between Saliva and Subgingival Plaque of Periodontitis Patients: A Systematic Review and Meta-Analysis.

The development of periodontitis is associated with an imbalanced subgingival microbial community enriched with species such as the traditionally classified red-complex bacteria (Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola). Saliva has been suggested as an alternative to subgingival plaque for the microbial analysis due to its easy and non-invasive collection. This systematic review aims to determine whether the levels of red-complex bacteria assessed using saliva reflect those in subgingival plaque from periodontitis patients. The MEDLINE, EMBASE, and Cochrane Library databases were searched up to April 30, 2021. Studies were considered eligible if microbial data of at least one of the red-complex species were reported in both saliva and subgingival plaque from periodontitis patients, based on DNA-based methods. Of the 17 included studies, 4 studies used 16S rRNA gene sequencing techniques, and the rest used PCR-based approaches. The detection frequency of each red-complex species in periodontitis patients was reported to be > 60% in most studies, irrespective of samples types. Meta-analyses revealed that both detection frequencies and relative abundances of red-complex bacteria in saliva were significantly lower than those in subgingival plaque. Moreover, the relative abundances of all 3 bacterial species in saliva showed significantly positive correlation with those in subgingival plaque. In conclusion, current evidence suggests that one-time saliva sampling cannot replace subgingival plaque for microbial analysis of the red-complex bacteria in periodontitis patients. Given the positive microbial associations between saliva and subgingival plaque, a thorough review of longitudinal clinical studies is needed to further assess the role of saliva.

The effect of equal fluoride concentrations in silver diamine fluoride and potassium fluoride on demineralized dentin during pH-cycling: chemical data.

The actual contribution of silver in silver diamine fluoride (SDF) towards the anti-demineralizing effect is unclear. This study compared the effects of single applications of three concentrations of fluoride (4.1%, 1.025%, 0.26% F- ) in the form of SDF and potassium fluoride (KF) on demineralized dentin in a 15-day non-microbial pH-cycling model. Calcium loss and uptake in de- and remineralization buffers were analyzed daily. Fluoride release in both buffers was analyzed on days 1, 2, 3, and 8. The net calcium results of de- and remineralization cycles revealed dose-response protection without significant differences between equal fluoride concentrations of SDF and KF. In the demineralization cycles, all fluoride treatments, except KF 0.26% F- , significantly inhibited demineralization, with KF 4.1% F- being the most effective. In the remineralization cycles, remineralization was enhanced in all fluoride concentration groups in a dose-response manner with no difference between similar fluoride concentrations of SDF and KF. Daily fluoride effects were constant throughout the experiment. Fluoride analysis revealed statistically significant differences in fluoride release between the treatments on day 1 that diminished on days 2 and 3. The non-microbial model showed no differences between SDF and KF in inhibiting demineralization and enhancing remineralization of dentin lesions.

Manipulation of Saliva-Derived Microcosm Biofilms To Resemble Dysbiotic Subgingival Microbiota.

Periodontitis is a highly prevalent oral inflammatory disease triggered by dysbiotic subgingival microbiota. For the development of microbiome modulators that can reverse the dysbiotic state and reestablish a health-associated microbiota, a high-throughput in vitro multispecies biofilm model is needed. Our aim is to establish a model that resembles a dysbiotic subgingival microbial biofilm by incorporating the major periodontal pathogen Porphyromonas gingivalis into microcosm biofilms cultured from pooled saliva of healthy volunteers. The biofilms were grown for 3, 7, and 10 days and analyzed for their microbial composition by 16S rRNA gene amplicon sequencing as well as measurement of dipeptidyl peptidase IV (DPP4) activity and butyric acid production. The addition of P. gingivalis increased its abundance in saliva-derived microcosm biofilms from 2.7% on day 3 to >50% on day 10, which significantly reduced the Shannon diversity but did not affect the total number of operational taxonomic units (OTUs). The P. gingivalis-enriched biofilms displayed altered microbial composition as revealed by principal-component analysis and reduced interactions among microbial species. Moreover, these biofilms exhibited enhanced DPP4 activity and butyric acid production. In conclusion, by adding P. gingivalis to saliva-derived microcosm biofilms, we established an in vitro pathogen-enriched dysbiotic microbiota which resembles periodontitis-associated subgingival microbiota in terms of increased P. gingivalis abundance and higher DPP4 activity and butyric acid production. This model may allow for investigating factors that accelerate or hinder a microbial shift from symbiosis to dysbiosis and for developing microbiome modulation strategies.IMPORTANCE In line with the new paradigm of the etiology of periodontitis, an inflammatory disorder initiated by dysbiotic subgingival microbiota, novel therapeutic strategies have been proposed targeting reversing dysbiosis and restoring host-compatible microbiota rather than eliminating the biofilms unselectively. Thus, appropriate laboratory models are required to evaluate the efficacy of potential microbiome modulators. In the present study, we used the easily obtainable saliva as an inoculum, spiked the microcosm biofilms with the periodontal pathogen Porphyromonas gingivalis, and obtained a P. gingivalis-enriched microbiota, which resembles the in vivo pathogen-enriched subgingival microbiota in severe periodontitis. This biofilm model circumvents the difficulties encountered when using subgingival plaque as the inoculum and achieves microbiota in a dysbiotic state in a controlled and reproducible manner, which is required for high-throughput and large-scale evaluation of strategies that can potentially modulate microbial ecology.

Short-Chain N-Acylhomoserine Lactone Quorum-Sensing Molecules Promote Periodontal Pathogens in In Vitro Oral Biofilms.

Acylhomoserine lactones (AHLs), the quorum-sensing (QS) signals produced by a range of Gram-negative bacteria, are involved in biofilm formation in many pathogenic and environmental bacteria. Nevertheless, the current paradigm excludes a role of AHLs in dental plaque formation, while other QS signals, such as AI-2 and autoinducer peptides, have been demonstrated to play an important role in biofilm formation and virulence-related gene expression in oral pathogens. In the present work, we have explored the effect of externally added AHLs on in vitro oral biofilm models for commensal, cariogenic, and periodontal dental plaque. While little effect on bacterial growth was observed, some AHLs specifically affected the lactic acid production and protease activity of the biofilms. Most importantly, the analysis of bacterial diversity in the biofilms showed that the addition of C6-homoserine lactone (C6-HSL) results in a shift toward a periodontal bacterial composition profile by increasing the relative presence of the orange-complex bacteria Peptostreptococcus and Prevotella These results point to a relevant role of AHL-mediated QS in dental plaque formation and might be involved in the development of dysbiosis, the mechanism of which should be further investigated. This finding potentially opens new opportunities for the prevention or treatment of the periodontal disease.IMPORTANCE Dental plaque is omnipresent in healthy oral cavities and part of our commensal microbial colonization. At the same time, dental plaque is the cause of the most common human diseases, caries and gum disease. Dental plaque consists of billions of microbes attached to the surface of your teeth. Communication among these microbes is pivotal for development of these complex communities yet poorly studied in dental plaque. In the present study, we show that a specific communication molecule induces changes within the community related to the development of gum disease. This finding suggests that interfering with microbial communication may represent an interesting novel strategy to prevent gum disease that should be further investigated.

Microcosm biofilms cultured from different oral niches in periodontitis patients.

Objective: Periodontal diseases are triggered by dysbiotic microbial biofilms. Therefore, it is essential to develop appropriate biofilm models. Aim of the present study was to culture microcosm biofilms inoculated from different niches in periodontitis patients and compare their microbial composition to those inoculated from subgingival plaque. Methods: Saliva, subgingival plaque, tongue and tonsils were sampled in five periodontitis patients to serve as inocula for culturing biofilms in vitro in an active attachment model. Biofilms were grown for 14 or 28 d and analyzed for their microbial composition by 16S rDNA sequencing. Results: As classified by HOMD, all biofilms were dominated by periodontitis-associated taxa, irrespective which niche had been used for inoculation. There was a low similarity between 14 d biofilms and their respective inocula (Bray-Curtis similarity 0.26), while biofilms cultured for 14 and 28 d shared high similarity (0.69). Principal components analysis showed much stronger clustering per patient than per niche indicating that the choice of patients may be more crucial than choice of the respective niches in these patients. Conclusion: Saliva, tongue scrapings or tonsil swabs may represent sufficient alternative inocula for growing microcosm biofilms resembling periodontitis-associated microbial communities in cases when sampling subgingival plaque is not possible.

Effect of mouthwashes on the composition and metabolic activity of oral biofilms grown in vitro.

OBJECTIVE: The aim of this study was to determine the effect of an oxygenating mouthwash compared to two other established mouthwash products on bacterial composition and metabolic activity of oral biofilms in vitro. MATERIAL AND METHODS: Twelve healthy subjects participated as donors. Plaque-saliva mixture inoculated biofilms were grown and treated with 3 different chemotherapeutic mouthwashes [amine fluoride/stannous fluoride (MD), oxygenating agent (AX), chlorhexidine 0.12 % (PA), and water (W)]. Effects of treatments were assessed on biofilm composition (16S rRNA gene amplicon sequencing), production of organic acids (formate, acetate, lactate, propionate, butyrate using capillary electrophoresis), and viability of the remaining biofilm (CFUs). RESULTS: Microbial profiles of biofilms clustered per inoculum donor and were dominated by the genera Veillonella, Streptococcus, and Prevotella. Microbial diversity was only reduced after PA treatment. Significant changes in composition occurred after treatment with AX, resulting in lower proportions of Veillonella and higher proportions of non-mutans streptococci. Production of all organic acids after PA and lactate after MD was significantly lower as compared to W. AX resulted in reduction of acetate, butyrate, and propionate and increase in lactate production (p < 0.05). Viable counts were significantly lower after PA and AX treatments compared to W, while no significant reduction was observed after MD. CONCLUSIONS: All studied mouthwashes affected the in vitro biofilms differently. The effects of the AX treatment were the most prominent which resulted in changes of the bacterial composition and metabolism. CLINICAL IMPLICATIONS: Awareness by the dental team that mouthwashes can change the bacterial composition and metabolism is important when advising its use.

In vitro phenotypic differentiation towards commensal and pathogenic oral biofilms.

Commensal oral biofilms, defined by the absence of pathology-related phenotypes, are ubiquitously present. In contrast to pathological biofilms commensal biofilms are rarely studied. Here, the effect of the initial inoculum and subsequent growth conditions on in vitro oral biofilms was studied. Biofilms were inoculated with saliva and grown anaerobically for up to 21 days in McBain medium with or without fetal calf serum (FCS) or sucrose. Pathology-related phenotypes were quantified and the community composition was determined. Biofilms inoculated with pooled saliva or individual inocula were similar. Denaturing gradient gel electrophoresis (DGGE) analysis allowed differentiation of biofilms grown with sucrose, but not with FCS. Lactate production by biofilms was significantly increased by sucrose and protease activity by FCS. McBain grown biofilms showed low activity for both phenotypes. Three clinically relevant in vitro biofilm models were developed and could be differentiated based on pathology-related phenotypes but not DGGE analysis. These models allow analysis of health-to-disease shifts and the effectiveness of prevention measures.

Application of an active attachment model as a high-throughput demineralization biofilm model.

OBJECTIVES: To investigate the potential of an active attachment biofilm model as a high-throughput demineralization biofilm model for the evaluation of caries-preventive agents. METHODS: Streptococcus mutans UA159 biofilms were grown on bovine dentine discs in a high-throughput active attachment model. Biofilms were first formed in a medium with high buffer capacity for 24h and then subjected to various photodynamic therapies (PACT) using the combination of Light Emitting Diodes (LEDs, Biotable(®)) and Photogem(®). Viability of the biofilms was evaluated by plate counts. To investigate treatment effects on dentine lesion formation, the treated biofilms were grown in a medium with low buffer capacity for an additional 24h. Integrated mineral loss (IML) and lesion depth (LD) were assessed by transversal microradiography. Calcium release in the biofilm medium was measured by atomic absorption spectroscopy. RESULTS: Compared to the water treated control group, significant reduction in viability of S. mutans biofilms was observed when the combination of LEDs and Photogem(®) was applied. LEDs or Photogem(®) only did not result in biofilm viability changes. Similar outcomes were also found for dentine lesion formation. Significant lower IML and LD values were only found in the group subjected to the combined treatment of LEDs and Photogem(®). There was a good correlation between the calcium release data and the IML or LD values. CONCLUSIONS: The high-throughput active attachment biofilm model is applicable for evaluating novel caries-preventive agents on both biofilm and demineralization inhibition. PACT had a killing effect on 24h S. mutans biofilms and could inhibit the demineralization process.

Effects of Lactobacillus rhamnosus GG on saliva-derived microcosms.

OBJECTIVE: The probiotic strain Lactobacillus rhamnosus GG (LGG) is shown to hamper the presence of mutans streptococci in saliva and may have positive effects on oral health. We investigated the effects of LGG on the cariogenic potential and microbial composition of saliva-derived microcosms. DESIGN: Single and dual species biofilms of LGG and Streptococcus mutans, and saliva-derived microcosms with or without LGG were grown in an Active Attachment Biofilm model. The microcosms were grown on bovine dentin/enamel discs in the presence or absence of sucrose (suc+/suc-). The presence of LGG was determined by multiplex ligation-dependent probe amplification (MLPA) and real-time PCR. Mutans streptococci (MS) and total viable counts, pH of the spent medium, capacity of lactate formation and integrated mineral loss in dentin was assessed. MLPA was used for identification and relative quantification of 20 oral microorganisms in the microcosms. Principal Component Analysis was applied to MLPA data. RESULTS: LGG inhibited the growth of S. mutans in dual species biofilms and did not affect the pH. LGG established in saliva-derived microcosms and reduced MS counts significantly, but did not affect pH or dentin demineralization. Simultaneous growth of the microcosms with LGG under heavy cariogenic conditions (suc+) introduced a compositional shift in the microbial community. The CFU, real-time PCR and MLPA data correlated significantly. CONCLUSION: We conclude that LGG established into and inhibited the growth of MS in complex saliva-derived biofilms, but this had no significant effect on cariogenic potential of the microcosms. This suggests that other microorganisms besides MS were responsible for increased cariogenicity of sucrose-exposed biofilms.

Influence of Streptococcus mutans on Enterococcus faecalis biofilm formation.

INTRODUCTION: An important virulence factor of Enterococcus faecalis is its ability to form biofilms. Most studies on biofilm formation have been carried out by using E. faecalis monocultures. Given the polymicrobial nature of root canal infections, it is important to understand biofilm formation of E. faecalis in the presence of other microorganisms. METHODS: Eight clinical strains of E. faecalis were tested for biofilm formation on hydroxyapatite disks in the presence and absence of a Streptococcus mutans biofilm. RESULTS: Significantly more E. faecalis viable cells were found in biofilms in the presence of S. mutans. This phenomenon was, however, strain-dependent. Of the 8 strains tested, biofilm formation of strains AA-OR34, ER5/1, and V583 was not influenced by S. mutans biofilms. CONCLUSIONS: The results from this study, especially the strain difference, underline the importance of studying biofilm formation in a more realistic multispecies setting.

Effects of Isomalt on enamel de- and remineralization, a combined in vitro pH-cycling model and in situ study.

Isomalt is a non-cariogenic sweetener, which is widely used in sugar-free candy and chewing gum. Little is known about the effects of Isomalt on de- and remineralization. Binding between calcium and Isomalt has been reported, which could affect the mineral balance. The objective of this study was to examine the effects of Isomalt on de- and remineralization of bovine enamel lesions, both in vitro and in situ. In in vitro study, subsurface enamel lesions were subjected to 3-weeks pH-cycling. Treatments were 5-min rinses with 10% Isomalt solutions daily and 10% Isomalt additions to re- or demineralizing solutions. Standard pH-cycling conditions were used with a 0.2 ppm fluoride background during the remineralization phase. In in situ study, subsurface lesions were exposed 2 months in vivo and brushed three times daily with 10% Isomalt containing toothpaste. Treatment effects were assessed by chemical analysis of the solutions (in vitro) and transversal microradiography (in vitro and in situ). In in vitro study, while 5-min rinses with 10% Isomalt gave slightly increased remineralization, continuous presence of 10% Isomalt (in re- or demineralizing solutions) inhibited both de- and/or remineralization. This lead to significantly smaller overall mineral loss when Isomalt was added during demineralization. In in situ study, remineralization enhancement during short Isomalt treatments was confirmed. Isomalt had a positive effect on the de/remineralization balance when given under conditions relevant to practical use.

Effects of a new titanium fluoride derivative on enamel de- and remineralization.

The potential of a new titanium fluoride (TiF) derivative for caries prevention was tested in a pH-cycling model. Daily treatments with various concentrations (100, 250, and 500 p.p.m.) of TiF were compared with similar sodium fluoride (NaF) treatments given at the same pH. Bovine enamel lesions were subjected to 3 wk of pH cycling. The effects were assessed by analyzing calcium uptake and loss in the re- and demineralizing solutions, respectively, and by post pH cycling microradiographic analysis of the lesions. Treatments with NaF reduced calcium loss, enhanced calcium uptake, and induced overall lesion remineralization. Treatments with TiF derivative gradually caused almost complete inhibition of calcium loss and uptake (lesion 'arrestment'), irrespective of the concentration of the TiF derivative. To test the permanence of protection, sound enamel was pretreated with either TiF derivative or NaF, and demineralized for 14 d at pH 4.4 and 4.6. Calcium loss data show that up to 80% inhibition of demineralization could be achieved for the TiF derivative, which was not possible for NaF treatments. This inhibition was obtained through a combination of concentration and number of treatments. The TiF derivative is a promising agent for the prevention of dental caries, especially when aimed at preventing the onset of caries.