-
Microbiology Spectrum Aug 2022The number of bacterial species recognized to utilize purposeful amyloid aggregation within biofilms continues to grow. The oral pathogen Streptococcus mutans produces...
The number of bacterial species recognized to utilize purposeful amyloid aggregation within biofilms continues to grow. The oral pathogen Streptococcus mutans produces several amyloidogenic proteins, including adhesins P1 (also known as AgI/II, PAc) and WapA, whose truncation products, namely, AgII and AgA, respectively, represent the amyloidogenic moieties. Amyloids demonstrate common biophysical properties, including recognition by Thioflavin T (ThT) and Congo red (CR) dyes that bind to the cross β-sheet quaternary structure of amyloid aggregates. Previously, we observed amyloid formation to occur only after 60 h or more of S. mutans biofilm growth. Here, we extend those findings to investigate where amyloid is detected within 1- and 5-day-old biofilms, including within tightly adherent compared with those in nonadherent fractions. CR birefringence and ThT uptake demonstrated amyloid within nonadherent material removed from 5-day-old cultures but not within 1-day-old or adherent samples. These experiments were done in conjunction with confocal microscopy and immunofluorescence staining with AgII- and AgA-reactive antibodies, including monoclonal reagents shown to discriminate between monomeric protein and amyloid aggregates. These results also localized amyloid primarily to the nonadherent fraction of biofilms. Lastly, we show that the C-terminal region of P1 loses adhesive function following amyloidogenesis and is no longer able to competitively inhibit binding of S. mutans to its physiologic substrate, salivary agglutinin. Taken together, our results provide new evidence that amyloid aggregation negatively impacts the functional activity of a widely studied S. mutans adhesin and are consistent with a model in which amyloidogenesis of adhesive proteins facilitates the detachment of aging biofilms. Streptococcus mutans is a keystone pathogen and causative agent of human dental caries, commonly known as tooth decay, the most prevalent infectious disease in the world. Like many pathogens, S. mutans causes disease in biofilms, which for dental decay begins with bacterial attachment to the salivary pellicle coating the tooth surface. Some strains of S. mutans are also associated with bacterial endocarditis. Amyloid aggregation was initially thought to represent only a consequence of protein mal-folding, but now, many microorganisms are known to produce functional amyloids with biofilm environments. In this study, we learned that amyloid formation diminishes the activity of a known S. mutans adhesin and that amyloid is found within the nonadherent fraction of older biofilms. This finding suggests that the transition from adhesin monomer to amyloid facilitates biofilm detachment. Knowing where and when S. mutans produces amyloid will help in developing therapeutic strategies to control tooth decay and other biofilm-related diseases.
Topics: Adhesins, Bacterial; Aging; Amyloid; Amyloidogenic Proteins; Biofilms; Dental Caries; Humans; Streptococcus mutans
PubMed: 35950854
DOI: 10.1128/spectrum.01661-22 -
Scientific Reports Feb 2018The aim of this study was to test the hypothesis that duplication/hybridization of functional domains of naturally occurring pellicle peptides amplified the inhibitory...
The aim of this study was to test the hypothesis that duplication/hybridization of functional domains of naturally occurring pellicle peptides amplified the inhibitory effect of hydroxyapatite crystal growth, which is related to enamel remineralization and dental calculus formation. Histatin 3, statherin, their functional domains (RR14 and DR9), and engineered peptides (DR9-DR9 and DR9-RR14) were tested at seven different concentrations to evaluate the effect on hydroxyapatite crystal growth inhibition. A microplate colorimetric assay was used to quantify hydroxyapatite crystal growth. The half-maximal inhibitory concentration (IC) was determined for each group. ANOVA and Student-Newman-Keuls pairwise comparisons were used to compare the groups. DR9-DR9 increased the inhibitory effect of hydroxyapatite crystal growth compared to single DR9 (p < 0.05), indicating that functional domain multiplication represented a strong protein evolution pathway. Interestingly, the hybrid peptide DR9-RR14 had an intermediate inhibitory effect compared to DR9 and DR9-DR9. This study used an engineered peptide approach to investigate a potential evolution protein pathway related to duplication/hybridization of acquired enamel pellicle's natural peptide constituents, contributing to the development of synthetic peptides for therapeutic use against dental caries and periodontal disease.
Topics: Crystallization; Dental Enamel; Dental Pellicle; Durapatite; Engineering; Peptides; Protein Domains
PubMed: 29491390
DOI: 10.1038/s41598-018-21854-4 -
Materials (Basel, Switzerland) Apr 2024Scanning force microscopy (SFM) is one of the most widely used techniques in biomaterials research. In addition to imaging the materials of interest, SFM enables the... (Review)
Review
Scanning force microscopy (SFM) is one of the most widely used techniques in biomaterials research. In addition to imaging the materials of interest, SFM enables the mapping of mechanical properties and biological responses with sub-nanometer resolution and piconewton sensitivity. This review aims to give an overview of using the scanning force microscope (SFM) for investigations on dental materials. In particular, SFM-derived methods such as force-distance curves (scanning force spectroscopy), lateral force spectroscopy, and applications of the FluidFM will be presented. In addition to the properties of dental materials, this paper reports the development of the pellicle by the interaction of biopolymers such as proteins and polysaccharides, as well as the interaction of bacteria with dental materials.
PubMed: 38730904
DOI: 10.3390/ma17092100 -
Caries Research 2015The effectiveness of fluoride in caries prevention has been convincingly proven. In recent years, researchers have investigated the preventive effects of different... (Review)
Review
The effectiveness of fluoride in caries prevention has been convincingly proven. In recent years, researchers have investigated the preventive effects of different fluoride formulations on erosive tooth wear with positive results, but their action on caries and erosion prevention must be based on different requirements, because there is no sheltered area in the erosive process as there is in the subsurface carious lesions. Thus, any protective mechanism from fluoride concerning erosion is limited to the surface or the near surface layer of enamel. However, reports on other protective agents show superior preventive results. The mechanism of action of tin-containing products is related to tin deposition onto the tooth surface, as well as the incorporation of tin into the near-surface layer of enamel. These tin-rich deposits are less susceptible to dissolution and may result in enhanced protection of the underlying tooth. Titanium tetrafluoride forms a protective layer on the tooth surface. It is believed that this layer is made up of hydrated hydrogen titanium phosphate. Products containing phosphates and/or proteins may adsorb either to the pellicle, rendering it more protective against demineralization, or directly to the dental hard tissue, probably competing with H(+) at specific sites on the tooth surface. Other substances may further enhance precipitation of calcium phosphates on the enamel surface, protecting it from additional acid impacts. Hence, the future of fluoride alone in erosion prevention looks grim, but the combination of fluoride with protective agents, such as polyvalent metal ions and some polymers, has much brighter prospects.
Topics: Cariostatic Agents; Dental Enamel Solubility; Fluorides; Humans; Phosphates; Protective Agents; Protons; Tin Compounds; Tooth Erosion
PubMed: 25871415
DOI: 10.1159/000380886 -
Brazilian Oral Research 2023The objective of this study was to compare the protein profile of the acquired enamel pellicle (AEP) formed in vivo in patients with or without gastroesophageal reflux...
The objective of this study was to compare the protein profile of the acquired enamel pellicle (AEP) formed in vivo in patients with or without gastroesophageal reflux disease (GERD), and with or without erosive tooth wear (ETW). Twenty-four volunteers were divided into 3 groups: 1) GERD and ETW; 2) GERD without ETW; and 3) control (without GERD). The AEP formed 120 min after prophylaxis was collected from the lingual/palatal surfaces. The samples were subjected to mass spectrometry (nLC-ESI-MS/MS) and label-free quantification by Protein Lynx Global Service software. A total of 213 proteins were identified, or 119, 92 and 106 from each group, respectively. Group 2 showed a high number of phosphorylated and calcium-binding proteins. Twenty-three proteins were found in all the groups, including 14-3-3 protein zeta/delta and 1-phosphatidylinositol. Several intracellular proteins that join saliva after the exfoliation of oral mucosa cells might have the potential to bind hydroxyapatite, or participate in forming supramolecular aggregates that bind to precursor proteins in the AEP. Proteins might play a central role in protecting the dental surface against acid dissolution.
Topics: Humans; Dental Pellicle; Tandem Mass Spectrometry; Tooth Wear; Durapatite; Gastroesophageal Reflux
PubMed: 37729290
DOI: 10.1590/1807-3107bor-2023.vol37.0085 -
Journal of Applied Oral Science :... 2020The acquired pellicle formation is the first step in dental biofilm formation. It distinguishes dental biofilms from other biofilm types.
INTRODUCTION
The acquired pellicle formation is the first step in dental biofilm formation. It distinguishes dental biofilms from other biofilm types.
OBJECTIVE
To explore the influence of salivary pellicle formation before biofilm formation on enamel demineralization.
METHODOLOGY
Saliva collection was approved by Indiana University IRB. Three donors provided wax-stimulated saliva as the microcosm bacterial inoculum source. Acquired pellicle was formed on bovine enamel samples. Two groups (0.5% and 1% sucrose-supplemented growth media) with three subgroups (surface conditioning using filtered/pasteurized saliva; filtered saliva; and deionized water (DIW)) were included (n=9/subgroup). Biofilm was then allowed to grow for 48 h using Brain Heart Infusion media supplemented with 5 g/l yeast extract, 1 mM CaCl2.2H2O, 5% vitamin K and hemin (v/v), and sucrose. Enamel samples were analyzed for Vickers surface microhardness change (VHNchange), and transverse microradiography measuring lesion depth (L) and mineral loss (∆Z). Data were analyzed using two-way ANOVA.
RESULTS
The two-way interaction of sucrose concentration × surface conditioning was not significant for VHNchange (p=0.872), ∆Z (p=0.662) or L (p=0.436). Surface conditioning affected VHNchange (p=0.0079), while sucrose concentration impacted ∆Z (p<0.0001) and L (p<0.0001). Surface conditioning with filtered/pasteurized saliva resulted in the lowest VHNchange values for both sucrose concentrations. The differences between filtered/pasteurized subgroups and the two other surface conditionings were significant (filtered saliva p=0.006; DIW p=0.0075). Growing the biofilm in 1% sucrose resulted in lesions with higher ∆Z and L values when compared with 0.5% sucrose. The differences in ∆Z and L between sucrose concentration subgroups was significant, regardless of surface conditioning (both p<0.0001).
CONCLUSION
Within the study limitations, surface conditioning using human saliva does not influence biofilm-mediated enamel caries lesion formation as measured by transverse microradiography, while differences were observed using surface microhardness, indicating a complex interaction between pellicle proteins and biofilm-mediated demineralization of the enamel surface.
Topics: Animals; Biofilms; Cattle; Dental Enamel; Dental Pellicle; Hardness; Microradiography; Pasteurization; Reference Values; Saliva; Sucrose; Surface Properties; Tooth Demineralization
PubMed: 32236356
DOI: 10.1590/1678-7757-2019-0501 -
Scientific Reports May 2023In contrast to pellicles formed in vivo, pellicles formed in vitro provide little to no erosion protection for enamel, possibly due to protein degradation from proteases...
In contrast to pellicles formed in vivo, pellicles formed in vitro provide little to no erosion protection for enamel, possibly due to protein degradation from proteases during pellicle formation. With the objective to achieve a more similar effect as observed for in vivo pellicles, the effects of adding protease inhibitors (PI) to saliva in vitro, and/or exchanging saliva repeatedly during pellicle formation were investigated in a cyclic model of pellicle formation and erosion with human enamel specimens. We repeatedly assessed surface microhardness (SMH), measured initial and final surface reflection intensity (SRI), and determined calcium released during erosion. For all the parameters tested, we observed a clear positive effect on erosion protection when adding PI to saliva for pellicle formation: SMH remained harder, SRI remained higher, and less calcium was released. Additionally, exchanging saliva with fresh one during pellicle formation led to a protective effect, but not as strong as the addition of PI. We conclude that adding protease inhibitors to saliva in vitro for pellicle formation leads to an erosion protective effect, which was further increased by repeatedly exchanging the saliva. Whether the pellicle itself more closely resembles in vivo pellicles remains to be investigated.
Topics: Humans; Saliva; Tooth Erosion; Calcium; Dental Pellicle; Calcium, Dietary; Protease Inhibitors
PubMed: 37244955
DOI: 10.1038/s41598-023-35334-x -
Archives of Oral Biology Jun 2016Numerous environmental factors influence the pathogenesis of Candida biofilms and an understanding of these is necessary for appropriate clinical management.
INTRODUCTION
Numerous environmental factors influence the pathogenesis of Candida biofilms and an understanding of these is necessary for appropriate clinical management.
AIMS
To investigate the role of material type, pellicle and stage of biofilm development on the viability, bioactivity, virulence and structure of C. albicans biofilms.
METHODS
The surface roughness (SR) and surface free energy (SFE) of acrylic and titanium discs was measured. Pellicles of saliva, or saliva supplemented with plasma, were formed on acrylic and titanium discs. Candida albicans biofilms were then generated for 1.5 h, 24h, 48 h and 72 h. The cell viability in biofilms was analysed by culture, whilst DNA concentration and the expression of Candida virulence genes (ALS1, ALS3 and HWP1) were evaluated using qPCR. Biofilm metabolic activity was determined using XTT reduction assay, and biofilm structure analysed by Scanning Electron Microscopy (SEM).
RESULTS
Whilst the SR of acrylic and titanium did not significantly differ, the saliva with plasma pellicle increased significantly the total SFE of both surface. The number of viable microorganisms and DNA concentration increased with biofilm development, not differing within materials and pellicles. Biofilms developed on saliva with plasma pellicle surfaces had significantly higher activity after 24h and this was accompanied with higher expression of virulence genes at all periods.
CONCLUSION
Induction of C. albicans virulence occurs with the presence of plasma proteins in pellicles, throughout biofilm growth. To mitigate such effects, reduction of increased plasmatic exudate, related to chronic inflammatory response, could aid the management of candidal biofilm-related infections.
Topics: Acrylic Resins; Biofilms; Candida albicans; Dental Pellicle; Fungal Proteins; Humans; Imaging, Three-Dimensional; In Vitro Techniques; Microbial Viability; Microscopy, Electron, Scanning; Polymethyl Methacrylate; Random Allocation; Salivary Proteins and Peptides; Surface Properties; Titanium; Virulence
PubMed: 26945171
DOI: 10.1016/j.archoralbio.2016.02.016 -
Regenerative Biomaterials 2024Eradicating biofouling from implant surfaces is essential in treating peri-implant infections, as it directly addresses the microbial source for infection and...
Eradicating biofouling from implant surfaces is essential in treating peri-implant infections, as it directly addresses the microbial source for infection and inflammation around dental implants. This controlled laboratory study examines the effectiveness of the four commercially available debridement solutions '(EDTA (Prefgel), NaOCl (Perisolv), HO (Sigma-Aldrich) and Chlorhexidine (GUM Paroex))' in removing the acquired pellicle, preventing pellicle re-formation and removing of a multi-species oral biofilm growing on a titanium implant surface, and compare the results with the effect of a novel formulation of a peroxide-activated 'Poloxamer gel (Nubone Clean)'. Evaluation of pellicle removal and re-formation was conducted using scanning electron microscope (SEM), energy-dispersive X-ray spectroscopy and X-ray photoelectron spectroscopy to assess the surface morphology, elemental composition and chemical surface composition. Hydrophilicity was assessed through contact angle measurements. The multi-species biofilm model included , and , reflecting the natural oral microbiome's complexity. Biofilm biomass was quantified using safranin staining, biofilm viability was evaluated using confocal laser scanning microscopy, and SEM was used for morphological analyses of the biofilm. Results indicated that while no single agent completely eradicated the biofilm, the 'Poloxamer gel' activated with 'HO' exhibited promising results. It minimized re-contamination of the pellicle by significantly lowering the contact angle, indicating enhanced hydrophilicity. This combination also showed a notable reduction in carbon contaminants, suggesting the effective removal of organic residues from the titanium surface, in addition to effectively reducing viable bacterial counts. In conclusion, the 'Poloxamer gel + HO' combination emerged as a promising chemical decontamination strategy for peri-implant diseases. It underlines the importance of tailoring treatment methods to the unique microbial challenges in peri-implant diseases and the necessity of combining chemical decontaminating strategies with established mechanical cleaning procedures for optimal management of peri-implant diseases.
PubMed: 38435376
DOI: 10.1093/rb/rbae014 -
PloS One 2019Vibrio cholerae is an important human pathogen causing intestinal disease with a high incidence in developing countries. V. cholerae can switch between planktonic and...
Vibrio cholerae is an important human pathogen causing intestinal disease with a high incidence in developing countries. V. cholerae can switch between planktonic and biofilm lifestyles. Biofilm formation is determinant for transmission, virulence and antibiotic resistance. Due to the enhanced antibiotic resistance observed by bacterial pathogens, antimicrobial nanomaterials have been used to combat infections by stopping bacterial growth and preventing biofilm formation. In this study, the effect of the nanocomposites zeolite-embedded silver (Ag), copper (Cu), or zinc (Zn) nanoparticles (NPs) was evaluated in V. cholerae planktonic cells, and in two biofilm states: pellicle biofilm (PB), formed between air-liquid interphase, and surface-attached biofilm (SB), formed at solid-liquid interfaces. Each nanocomposite type had a distinctive antimicrobial effect altering each V. cholerae lifestyles differently. The ZEO-AgNPs nanocomposite inhibited PB formation at 4 μg/ml, and prevented SB formation and eliminated planktonic cells at 8 μg/ml. In contrast, the nanocomposites ZEO-CuNPs and ZEO-ZnNPs affect V. cholerae viability but did not completely avoid bacterial growth. At transcriptional level, depending on the nanoparticles and biofilm type, nanocomposites modified the relative expression of the vpsL, rbmA and bap1, genes involved in biofilm formation. Furthermore, the relative abundance of the outer membrane proteins OmpT, OmpU, OmpA and OmpW also differs among treatments in PB and SB. This work provides a basis for further study of the nanomaterials effect at structural, genetic and proteomic levels to understand the response mechanisms of V. cholerae against metallic nanoparticles.
Topics: Anti-Bacterial Agents; Bacterial Outer Membrane Proteins; Biofilms; Copper; Dental Pellicle; Gene Expression Regulation, Bacterial; Humans; Metal Nanoparticles; Microbial Sensitivity Tests; Nanocomposites; Plankton; Silver; Transcription, Genetic; Vibrio cholerae; Zeolites; Zinc
PubMed: 31188854
DOI: 10.1371/journal.pone.0217869