Jacob M. ten Cate

Development of Candida-associated denture stomatitis: new insights

Despite therapeutic progress, opportunistic oral fungal infectious diseases have increased in prevalence, especially in denture wearers. The combination of entrapment of yeast cells in irregularities in denture-base and denture-relining materials, poor oral hygiene and several systemic factors is the most probable cause for the onset of this infectious disease. Hence colonization and growth on prostheses by Candida species are of clinical importance. The purpose of this review is to critically discuss several key factors controlling the adhesion of Candida species which are relevant to denture-associated stomatitis. Although there is some consensus on the role of surface properties, studies on several other factors, as the use of denture liners, salivary properties and yeast-bacterial interactions, have shown contradictory findings. A comprehensive fundamental understanding is hampered by conflicting findings due to the large variations in experimental protocols, while other factors have never been thoroughly studied. Surface free energy and surface roughness control the initial adherence, but temporal changes have not been reported. Neither have in vivo studies shown if the substratum type is critical in dictating biofilm accumulation during longer periods in the oral environment. The contribution of saliva is unclear due to factors like variations in its collection and handling. Initial findings have disclosed that also bacteria are crucial for the successful establishment of Candida in biofilms, but the clinical significance of this observation is yet to be confirmed. In conclusion, there is a need to standardize experimental procedures, to bridge the gap between laboratory and in vivo methodologies and findings and – in general – to thoroughly investigate the factors that modulate the initial attachment and subsequent colonization of denture-base materials and the oral mucosa of patients subjected to Candida infections. Information on how these factors can be controlled is required and this may help to prevent the disease. The societal impact of such information is significant given the magnitude of the candidosis problem worldwide.

The effect of Streptococcus mutans and Candida glabrata on Candida albicans biofilms formed on different surfaces

Although Candida containing biofilms contribute to the development of oral candidosis, the characteristics of multi-species Candida biofilms and how oral bacteria modulate these biofilms is poorly understood. The aim of this study was to investigate interactions between Candida albicans and either Candida glabrata or Streptococcus mutans in biofilms grown on various surfaces, with or without saliva. Hydroxyapatite (HA), polymethylmetacrylate (PMMA) and soft denture liner (SL) discs were used as substratum. Counts of viable micro-organisms in the accumulating biofilm layer were determined and converted to colony forming units per unit surface area. Confocal laser scanning microscopy was used to characterize biofilms and to quantitate the number of hyphae in each condition tested. Viable counts of C. albicans and C. glabrata per mm(2) decreased in the order HA>PMMA>SL (p<0.05). Biofilms grown on saliva-coated specimens harboured fewer C. glabrata than uncoated specimens (p<0.05). Glucose and the presence of S. mutans suppressed C. albicans hyphal formation. Dual Candida species biofilms did not show competitive interaction between the two species. We conclude that Candida biofilms are significantly affected by saliva, substratum type and by the presence of other micro-organisms.

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.

Effect of denture base‐resin with prereacted glass‐ionomer filler on dentin demineralization

The demineralization of dentin was studied when placed adjacent to one of four experimental denture base-resins. These experimental resins contained polymethylmethacrylate (PMMA) and 0, 5, 10, 20 or 30 wt% surface reaction-type prereacted glass-ionomer (S-PRG) filler, respectively. A dentin thin-section was sandwiched between these resins and subjected to demineralization for 1 wk. Microradiographic analyses showed that the mineral vol% of the surface was increased, and lesion formation was inhibited, in a dose-response relationship with the amount of S-PRG filler. Moreover, the mineral loss decreased, by up to 60%, with an increasing amount of filler. These results indicate that denture base-resins containing S-PRG filler possess a substantial inhibition capability against demineralization, and this may thereby assist in preventing root caries of abutment teeth.

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.

Pre- and postnatal determinants of deciduous molar hypomineralisation in 6-year-old children. The generation R study

Background Deciduous Molar Hypomineralisation (DMH) and Molar Incisor Hypomineralisation (MIH) are common developmental disturbances in pediatric dentistry. Their occurrence is related. The same determinants as suggested for MIH are expected for DMH, though somewhat earlier in life. Perinatal medical problems may influence the prevalence of DMH but this has not been studied sufficiently. Objective This study aimed to identify possible determinants of DMH in a prospective cohort study among 6-year-old children. Study Design This study was embedded in the Generation R Study, a population-based prospective cohort study from fetal life until young adulthood. The the data were used to identify the determinants of DMH. Clinical photographs of clean, moist teeth were taken with an intra-oral camera in 6690 children (mean age 6.2 years; 49.9% girls). Data on possible determinants that had occurred during pregnancy and/or the childs first year of life were on the basis of manual standardized measurements (like length and weight) and questionnaires. Multivariate analyse with backward and forward selection was performed. Results A number of factors in the pre-, peri- and postnatal phase were found to be associated with DMH. After multivariate logistic regression analyses, Dutch ethnic background, low birth weight, maternal alcohol consumption during pregnancy, and fever episodes in the first year of the childs life were found to play a role in the development of DMH in 6-year-old children. Conclusion This study shows that Dutch ethnicity, low birth weight, alcohol consumption by the mother during pregnancy and any fever in the first year of the childs life are associated with DMH. Not only childhood factors but also prenatal lifestyle factors need to be taken into account when studying determinants for DMH.

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.

Changes in the oral ecosystem induced by the use of 8% arginine toothpaste

OBJECTIVE Bacterial metabolism of arginine in the oral cavity has a pH-raising and thus, potential anti-caries effect. However, the influence of arginine on the oral microbial ecosystem remains largely unresolved. DESIGN In this pilot study, nine healthy individuals used toothpaste containing 8% arginine for eight weeks. Saliva was collected to determine arginolytic potential and sucrose metabolic activity at the Baseline, Week 4, Week 8 and after a two weeks Wash-out period. To follow the effects on microbial ecology, 16S rDNA sequencing on saliva and plaque samples at Baseline and Week 8 and metagenome sequencing on selected saliva samples of the same time-points was performed. RESULTS During the study period, the arginolytic potential of saliva increased, while the sucrose metabolism in saliva decreased. These effects were reversed during the Wash-out period. Although a few operational taxonomic units (OTUs) in plaque changed in abundance during the study period, there was no real shift in the plaque microbiome. In the saliva microbiome there was a significant compositional shift, specifically the genus Veillonella had increased significantly in abundance at Week 8. CONCLUSION Indeed, the presence of arginine in toothpaste affects the arginolytic capacity of saliva and reduces its sucrose metabolic activity. Additionally, it leads to a shift in the salivary microbiome composition towards a healthy ecology from a caries point of view. Therefore, arginine can be regarded as a genuine oral prebiotic.

Dynamics of red fluorescent dental plaque during experimental gingivitis-A cohort study

OBJECTIVES The dynamics of red fluorescent plaque (RFP) in comparison to clinical plaque and bleeding scores were studied during an experimental gingivitis protocol in a cohort of healthy participants. METHODS Forty-one participants were monitored for RFP before (24h plaque), during 14 days plaque accumulation (days 2, 5, 9, 14) and after 7 days recovery (24h plaque). RFP was assessed on fluorescence photographs of the vestibular aspect of the anterior teeth (cuspid to cuspid) in the upper and lower jaw. Clinical plaque and bleeding were assessed at days -14, 0, 14 and 21. RESULTS RFP of 24h plaque was reproducible (days -14, 0), then increased during 14 days plaque accumulation and returned to baseline after 7 days recovery. Groups of low, moderate and high RFP formers were statistically significantly different at all times even already at baseline. The individual RFP response during 14 days plaque accumulation correlated well with RFP of 24h plaque (days -14, 0). RFP correlated moderate to well with clinical plaque at days -14, 0, 14 and 21. From day 2 of the gingivitis challenge RFP correlated with bleeding at day 14. CONCLUSIONS RFP provided an objective measure of oral hygiene status. Given the correlation with clinical parameters found, the amount of RFP after 24h plaque accumulation was indicatory for the inflammatory response during a prolonged period of no oral hygiene. This trial was registered at the public trial register ​of the Central Committee on Research Involving Human Subjects (CCMO) under number NL51111.029.14 CLINICAL SIGNIFICANCE: This paper shows the association between RFP after 24h plaque accumulation and inflammatory response after a prolonged period of no oral hygiene. Red plaque fluorescence can be used to identify subjects at risk for developing gingival inflammation.

The effect of fixed orthodontic appliances and fluoride mouthwash on the oral microbiome of adolescents: a randomized controlled clinical trial

While the aesthetic effect of orthodontic treatment is clear, the knowledge on how it influences the oral microbiota and the consequential effects on oral health are limited. In this randomized controlled clinical trial we investigated the changes introduced in the oral ecosystem, during and after orthodontic treatment with fixed appliances in combination with or without a fluoride mouthwash, of 10–16.8 year old individuals (N = 91). We followed several clinical parameters in time, in combination with microbiome changes using next-generation sequencing of the bacterial 16S rRNA gene. During the course of our study, the oral microbial community displayed remarkable resilience towards the disturbances it was presented with. The effects of the fluoride mouthwash on the microbial composition were trivial. More pronounced microbial changes were related to gingival health status, orthodontic treatment and time. Periodontal pathogens (e.g. Selenomonas and Porphyromonas) were highest in abundance during the orthodontic treatment, while the health associated Streptococcus, Rothia and Haemophilus gained abundance towards the end and after the orthodontic treatment. Only minor compositional changes remained in the oral microbiome after the end of treatment. We conclude that, provided proper oral hygiene is maintained, changes in the oral microbiome composition resulting from orthodontic treatment are minimal and do not negatively affect oral health.