Oral Biofilm Growth Research Articles

Correlation Analyses of the Oral Biofilm Growth Inhibition towards Hydrophobicity Reduction of Oral Pathogenic Bacteria

Correlation Analyses of the Oral Biofilm Growth Inhibition towards Hydrophobicity Reduction of Oral Pathogenic Bacteria

Crystal structure of the pilus-specific sortase from early colonizing oral Streptococcus sanguinis captures an active open-lid conformation

Dental plaque is a complex microbial biofilm community of many species and a major cause of oral infections and infectious endocarditis. Plaque development begins when primary colonizers attach to oral tissues and undergo coaggregation. Primary colonizers facilitate cellular attachment and inter-bacterial interactions through sortase-dependent pili (or fimbriae) extending out from their cell surface. Consequently, the sortase enzyme is viewed as a potential drug target for controlling biofilm formation and avoiding infection. Streptococcus sanguinis is a primary colonizing bacterium whose pili consist of three different pilin subunits that are assembled together by the pilus-specific (C-type) SsaSrtC sortase. Here, we report on the crystal structure determination of the recombinant wild-type and active-site mutant forms of SsaSrtC. Interestingly, the SsaSrtC structure exhibits an open-lid conformation, although a conserved DPX motif is lacking in the lid. Based on molecular docking and structural analysis, we identified the substrate-binding residues essential for pilin recognition and pilus assembly. We also demonstrated that while recombinant SsaSrtC is enzymatically active toward the five-residue LPNTG sorting motif peptide of the pilins, this activity is significantly reduced by the presence of zinc. We further showed that rutin and α-crocin are potential candidate inhibitors of the SsaSrtC sortase via structure-based virtual screening and inhibition assays. The structural knowledge gained from our study will provide the means to develop new approaches that target pilus-mediated attachment, thereby preventing oral biofilm growth and infection.

Impact of different oral treatments on the composition of the supragingival plaque microbiome

ABSTRACT Background Antiseptics are used to inhibit oral biofilm growth. However, they affect not only pathogenic but also commensal bacteria, which are a natural barrier against oral diseases. Objective Using a metaproteome approach combined with a standard plaque-regrowth study, this pilot study examined the impact of different concentrations of lactoperoxidase (LPO)-system containing lozenges on early plaque formation, and active biological processes. Design Sixteen orally healthy subjects received four local treatments as a randomized single-blind study based on a cross-over design. Two lozenges containing components of the LPO-system in different concentrations were compared to a placebo and Listerine®. The newly formed dental plaque was analyzed by mass spectrometry (nLC-MS/MS). Results On average 1,916 metaproteins per sample were identified, which could be assigned to 116 genera and 1,316 protein functions. Listerine® reduced the number of metaprotein groups and their relative abundance, confirming the plaque inhibiting effect. The LPO-lozenges triggered mainly higher metaprotein abundances of early and secondary colonizers as well as bacteria associated with dental health but also periodontitis. Functional information indicated plaque biofilm growth. Conclusion The effects of Listerine® and LPO-system containing lozenges used for plaque inhibition are different. In contrast to Listerine®, the lozenges allowed maintenance of a higher bacterial diversity.

OZONE THERAPY AS AN ADJUNCT THERAPY IN DENTISTRY: A REVIEW

The oral cavity gives the impression of being an open ecosystem, with a dynamic equilibrium between the entry of microbes, colonisation methods, and host defences designed to remove them: the oral mucosa, the gingiva, the gingival crevices, the gingival crevices, the ging Bacteria need to attach themselves to either hard tooth surfaces or epithelial surfaces in order to prevent eradication. Oral caries, periodontal disease, and peri-implantitis are among of the most frequent oral diseases, and the production and growth of oral biofilms, as well as the inside selection of certain microbes, have all been found to be connected with these conditions. The eradication of the biofilm by mechanical means and the use of antibiotic disinfectants or other types of antibiotics as an adjuvant treatment have been the traditional approaches to periodontal care. Ozone, which has the chemical formula O3, is a triatomic molecule that is composed of three atoms of oxygen. The use of ozone in dentistry and medicine has been advised for the treatment of 260 various diseases and conditions. The ozone therapy has proven to be more helpful than the currently available conventional therapeutic modalities. These modalities typically involve a conservative and minimally intrusive approach to dental care. The demonstration of the molecular processes behind ozone's practical functions is of great help to the field of dentistry.

Bacterial Biofilm Growth on Various Dental Stabilization Systems for Avulsed and Luxated Teeth

With the increased incidence of traumatic injuries and the advanced understanding of the periodontal and alveolar healing process, teeth splinting has become a common practice for stabilizing traumatized teeth. Consequently, several splinting materials and techniques have been introduced in the past few years. Despite the detrimental role of bacterial biofilm on healing, the level of biofilm development on these material surfaces has not been well investigated. Bacterial biofilms are severely detrimental for periodontal healing of avulsed and luxated teeth. Thus, biofilm growth becomes a critical factor in selecting the material of choice for dental splints. In this study, we aim to assess the level of oral biofilm growth on four different splinting systems: Ribbond©, orthodontic NiTi wire, monofilament fishing line, and Titanium Trauma Splint. A total of 72 extracted anterior teeth were divided into four groups. We splinted six rows of three teeth each per group. The teeth selected were caries-free and periodontitis-free at the time of extraction. To assess biofilm growth, a supragingival dental plaque sample was cultured and directly inoculated into all groups. After 7 days, bacterial growth was quantified by live/dead fluorescent microscopy assay and colony forming unit counts (CFU). Using one-way ANOVA and Bonferroni’s post hoc tests, we demonstrated that all splint systems allowed for bacterial growth. However, the Titanium Trauma Splint (TTS) allowed for the least amount of biofilm growth compared to other splint systems.

Long-term antibacterial activity and cytocompatibility of novel low-shrinkage-stress, remineralizing composites

A low-shrinkage-stress (LSS), antibacterial and remineralizing nanocomposite was recently developed; however, validation of its long-term antibacterial potency in modulating human salivary-derived biofilm is an unmet need. This study aimed to evaluate the antibacterial effect of the bioactive LSS composite before and after aging in acidic solution for 90 days using a multi-species biofilm model, and to evaluate its cytotoxicity. The LSS composite consisted of urethane dimethacrylate (UDMA) and triethylene glycol divinylbenzyl ether (TEG-DVBE), 3% dimethylaminohexadecyl methacrylate (DMAHDM) and 20% nanoparticles of amorphous calcium phosphate (NACP). Biofilm colony-forming units (CFU), lactic acid production, and confocal laser scanning microscopy (3D biofilm) were evaluated before and after three months of aging. Cytotoxicity was assessed against human gingival fibroblasts (HGF). The new LSS composite presented the lowest biofilm CFU, lactic acid and biofilm biomass, compared to controls (n = 6, p < 0.05). Importantly, the new composite exhibited no significant difference in antibacterial performance before and after 90-day-aging, demonstrating long-term antibacterial activity (p > 0.1). The LSS antibacterial and remineralizing composite presented a low cell viability at original extract that has increased with further dilutions. In conclusion, this study spotlighted that the new bioactive composite not only had a low shrinkage stress, but also down-regulated the growth of oral biofilms, reduced acid production, maintained antibacterial activity after the 90-day-aging, and did not compromise the cytocompatibility.

Evidence for Proline Utilization by Oral Bacterial Biofilms Grown in Saliva.

Within the mouth bacteria are starved of saccharides as their main nutrient source between meals and it is unclear what drives their metabolism. Previously oral in vitro biofilms grown in saliva have shown proteolytic degradation of salivary proteins and increased extracellular proline. Although arginine and glucose have been shown before to have an effect on oral biofilm growth and activity, there is limited evidence for proline. Nuclear magnetic resonance (NMR) spectroscopy was used to identify extracellular metabolites produced by bacteria in oral biofilms grown on hydroxyapatite discs. Biofilms were inoculated with stimulated whole mouth saliva and then grown for 7 days using sterilized stimulated whole mouth saliva supplemented with proline, arginine or glucose as a growth-medium. Overall proline had a beneficial effect on biofilm growth—with significantly fewer dead bacteria present by biomass and surface area of the biofilms (p < 0.05). Where arginine and glucose significantly increased and decreased pH, respectively, the pH of proline supplemented biofilms remained neutral at pH 7.3–7.5. SDS-polyacrylamide gel electrophoresis of the spent saliva from proline and arginine supplemented biofilms showed inhibition of salivary protein degradation of immature biofilms. NMR analysis of the spent saliva revealed that proline supplemented biofilms were metabolically similar to unsupplemented biofilms, but these biofilms actively metabolized proline to 5-aminopentanoate, butyrate and propionate, and actively utilized glycine. This study shows that in a nutrient limited environment, proline has a beneficial effect on in vitro oral biofilms grown from a saliva inoculum.

Novel Crown Cement Containing Antibacterial Monomer and Calcium Phosphate Nanoparticles.

Oral biofilm accumulation at the tooth–restoration interface often leads to recurrent dental caries and restoration failure. The objectives of this study were to: (1) develop a novel bioactive crown cement containing dimethylaminohexadecyl methacrylate (DMAHDM) and nano-sized amorphous calcium phosphate (NACP), and (2) investigate the mechanical properties, anti-biofilm activity, and calcium (Ca2+) and phosphate (PO43−) ion release of the crown cement for the first time. The cement matrix consisted of pyromellitic glycerol dimethacrylate and ethoxylated bisphenol-A dimethacrylate monomers and was denoted PEHB resin matrix. The following cements were tested: (1) RelyX luting cement (commercial control); (2) 55% PEHB + 45% glass fillers (experimental control); (3) 55% PEHB + 20% glass + 25% NACP + 0% DMAHDM; (4) 52% PEHB + 20% glass + 25% NACP + 3% DMAHDM; (5) 51% PEHB + 20% glass + 25% NACP + 4% DMAHDM; (6) 50% PEHB + 20% glass + 25% NACP + 5% DMAHDM. Mechanical properties and ion release were measured. Streptococcus mutans (S. mutans) biofilms were grown on cements, and colony-forming units (CFUs) and other biofilm properties were measured. The novel bioactive cement demonstrated strong antibacterial properties and high levels of Ca2+ and PO43− ion release to remineralize tooth lesions. Adding NACP and DMAHDM into the cement did not adversely affect the mechanical properties and dentin bonding strength. In conclusion, the novel NACP + DMAHDM crown cement has excellent potential for restoration cementation to inhibit caries by suppressing oral biofilm growth and increasing remineralization via Ca2+ and PO43− ions. The NACP + DMAHDM composition may have wide applicability to other biomaterials to promote hard-tissue formation and combat bacterial infection.

Titanium particles and ions favor dysbiosis in oral biofilms.

To evaluate the effect of titanium (Ti) particles and ions on oral biofilm growth and composition. Particles and ions of Ti released from dental implants can trigger unfavorable biological responses in human cells. However, their effect on oral biofilms composition has not been tested. In this blind in situ study, volunteers wore a palatal appliance containing Ti disks for 7days to allow biofilm formation. Disks were then collected and biofilms were treated, in vitro, with Ti particles (0.75% and 1%), ions (10 and 20ppm), or a combination of both (1% particles+20ppm ions). Biofilms exposed only to medium was used as control group. After 24hours, biofilms were collected and analyzed by checkerboard DNA-DNA hybridization. Direct effects of Ti particles and ions on biofilm/cellular morphology were evaluated by transmission electron microscopy (TEM). Ti particles affected biofilm composition, increasing population of four bacterial species (P<.05), while Ti ions showed higher levels of putative pathogens from the orange complex with reduction in species from the yellow complex (P<.05), compared with control. The combination of particles+ions increased green complex and reduced yellow complex proportions (P<.05). TEM showed clusters of particles agglomerated in extracellular environment, while Ti ions were precipitated in both extracellular and intracellular sites. Ti products, especially Ti ions, have the potential to change the microbiological composition of biofilms formed on Ti surfaces. Therefore, the presence of Ti products around dental implants may contribute to microbial dysbiosis and peri-implantitis.

The application of natural antibacterial coating for the surface modification of dental implants and abutments

Background Nowadays, peri-implantitis, a destructive inflammatory process around implants contributing to bone loss, has become the main reason for implant failure. Bacterial colonization and subsequent plaque formation on the implant surface is one of the main factors causing peri-implantitis. Totarol is a kind of natural biocompatible antibacterial agent, with bactericidal effect towards a wide range of gram-positive, even antibiotic-resistant bacteria, including the methicillin-resistant S. aureus. Aim/Hypothesis This in vitro study aims at applying totarol coatings on experimental dental implant and abutment surfaces and investigating the antibacterial capability of totarol coatings in a simulated oral environment. Material and Methods After ultrasonic cleaning with 70% ethanol, highly polished Ti disks (D = 10 mm) are coated with totarol by spin coating. Column chromatography-purified totarol dissolved in ethyl acetate solutions (20 mg/ml) is coated on Ti disks. Ti disks (n = 6 group) with and without totarol coating are incubated in an S. gordonii-suspension in Schaedler medium or in a mixed oral bacteria suspension in Schaedler medium (start OD λ=620 nm = 0.54) in a stirring system (each cycle including pause for 60 min and stirring for 15 min in, 37°C incubator) simulating the eating and resting period in human oral environment. The incubated Ti disks were harvested after 4 h, 8 h, 24 h, 48 h of incubation, respectively, then went through Live Dead staining for testing the vitality and through crystal violet staining for quantifying the amount of surface-bound bacteria. One-way ANOVA was used to analyze differences of the quantity of surface-bound bacteria between totarol coatings and control group. Results Approximately all S. gordonii and all mixed oral bacteria bound onto the totarol surface are red stained compared with the green stained live bacteria in the Ti control group after 4 h, 8 h, 24 h incubation (Fig. 1). This implied a contact killing effect towards S. gordonii and the mixed oral bacteria at a different stage of biofilm formation. Furthermore, crystal violet staining shows a significantly lower density of bacteria (P < 0.05) on totarol compared with the control, indicating an inhibition effect of totarol coatings towards the S. gordonii and multi-species oral biofilm growth (Fig. 2). Conclusion and Clinical Implications The natural antibacterial totarol coating showed promising bactericidal effects towards the early oral colonizer S. gordonii and the multispecies oral bacterial culture, and simultaneously significantly inhibits their biofilm growth. This might favor the undisturbed formation of a tight epithelial seal and prevent bacterial infiltration. Therefore, totarol coatings on dental implant and abutment surfaces showed great potential for the reduction or prevention of bacterial caused peri-implantitis.

Description of Streptococcus mutans, Streptococcus sanguinis, and Candida albicans biofilms after exposure to propolis dentifrice by using OpenCFU method

ContextDental caries is a major and chronic dental public health problem, which can usually be prevented by regular oral hygiene. The most common oral hygiene practice is brushing teeth with a dentifrice. Propolis has emerged as a promising anti-cariogenic agent, which is considered to be a good oral antiseptic for prevention of caries. Several studies have shown that the use of C has an influence in the growth of oral biofilms. There are several standard methods used to count bacterial colonies, such as crystal violet and CFU Count assays. OpenCFU method is a technique that can be used to calculate biofilm colonies more faster, precisely, and accurately. AimTo compare several methods for evaluating the number of biofilm colonies formed with exposure to a standard dentifrice and propolis. Methods and materialsBiofilm assays were carried out on 96-well microplates. Reference strains of oral Streptococcus species (S. mutans ATCC 25175T and S. sanguinis ATCC 10566T) and yeast (Candida albicans ATCC 10231T) were inoculated into wells, and 200 µL of standard and propolis dentifrice solution were added to each well and incubated for 18 h at 37 °C. Bacteria and yeast were then sub-cultured on respective media and the colony-forming units (CFU) were counted manually. The other wells were stained by crystal violet and incubated for 15 min, followed by observation using an inverted microscope and evaluated using crystal violet analysis and the OpenCFU automated method. ResultsThe numbers of CFUs determined for all strains were similar in the standard-dentifrice group and propolis-dentifrice group, and were similar among the three methods: crystal violet staining, manual CFU count, and OpenCFU analysis. ConclusionOpenCFU analysis can be reliably used as a rapid and a more practical method to analyse the growth of oral microorganism biofilms. However, high digital image quality is required to provide an accurate analysis for colony counting.

Novel Protein-Repellent and Antibacterial Resins and Cements to Inhibit Lesions and Protect Teeth

Orthodontic treatment is increasingly popular as people worldwide seek esthetics and better quality of life. In orthodontic treatment, complex appliances and retainers are placed in the patients’ mouths for at least one year, which often lead to biofilm plaque accumulation. This in turn increases the caries-inducing bacteria, decreases the pH of the retained plaque on an enamel surface, and causes white spot lesions (WSLs) in enamel. This article reviews the cutting-edge research on a new class of bioactive and therapeutic dental resins, cements, and adhesives that can inhibit biofilms and protect tooth structures. The novel approaches include the use of protein-repellent and anticaries polymeric dental cements containing 2-methacryloyloxyethyl phosphorylcholine (MPC) and dimethylaminododecyl methacrylate (DMAHDM); multifunctional resins that can inhibit enamel demineralization; protein-repellent and self-etching adhesives to greatly reduce oral biofilm growth; and novel polymethyl methacrylate resins to suppress oral biofilms and acid production. These new materials could reduce biofilm attachment, raise local biofilm pH, and facilitate the remineralization to protect the teeth. This novel class of dental resin with dual benefits of antibacterial and protein-repellent capabilities has the potential for a wide range of dental and biomedical applications to inhibit bacterial infection and protect the tissues.

Consistent and reproducible long-term in vitro growth of health and disease-associated oral subgingival biofilms

BackgroundSeveral in vitro oral biofilm growth systems can reliably construct oral microbiome communities in culture, yet their stability and reproducibility through time has not been well characterized. Long-term in vitro growth of natural biofilms would enable use of these biofilms in both in vitro and in vivo studies that require complex microbial communities with minimal variation over a period of time. Understanding biofilm community dynamics in continuous culture, and whether they maintain distinct signatures of health and disease, is necessary to determine the reliability and applicability of such models to broader studies. To this end, we performed next-generation sequencing on biofilms grown from healthy and disease-site subgingival plaque for 80 days to assess stability and reliability of continuous oral biofilm growth.ResultsBiofilms were grown from subgingival plaque collected from periodontitis-affected sites and healthy individuals for ten eight-day long generations, using hydroxyapatite disks. The bacterial community in each generation was determined using Human Oral Microbe Identification by Next-Generation Sequencing (HOMINGS) technology, and analyzed in QIIME. Profiles were steady through the ten generations, as determined by species abundance and prevalence, Spearman’s correlation coefficient, and Faith’s phylogenetic distance, with slight variation predominantly in low abundance species. Community profiles were distinct between healthy and disease site-derived biofilms as demonstrated by weighted UniFrac distance throughout the ten generations. Differentially abundant species between healthy and disease site-derived biofilms were consistent throughout the generations.ConclusionsHealthy and disease site-derived biofilms can reliably maintain consistent communities through ten generations of in vitro growth. These communities maintain signatures of health and disease and of individual donors despite culture in identical environments. This subgingival oral biofilm growth and perpetuation model may prove useful to studies involving oral infection or cell stimulation, or those measuring microbial interactions, which require the same biofilms over a period of time.

Soft, adhesive (+) alpha tocopherol phosphate planar bilayers that control oral biofilm growth through a substantive antimicrobial effect

‘Soft’ nanomaterials have the potential to produce substantive antibiofilm effects. The aim of this study was to understand the oral antimicrobial activity of soft nanomaterials generated from alpha-tocopherol (α-T) and alpha-tocopherol phosphate (α-TP). (+) α-TP formed planar bilayer islands (175 ± 21 nm, −14.9 ± 3.5 mV) in a Trizma® buffer, whereas (+) α-T formed spherical liposomes (563 ± 1 nm, −10.5 ± 0.2 mV). The (+) α-TP bilayers displayed superior Streptococcus oralis biofilm growth retardation, a more substantive action, generated a superior adsorption to hydroxyapatite and showed an enhanced inhibition of multi-species bacterial saliva biofilm growth (38 ± 7μm vs 58 ± 18 μm, P ˂ 0.05) compared to (+) α-T. Atomic force microscopy data indicated that the ability of the ‘soft’ α-TP nanomaterials to transition into planar bilayer structures upon contact with interfaces facilitated their adhesive properties and substantive antimicrobial effects.

Do Dental Resin Composites Accumulate More Oral Biofilms and Plaque than Amalgam and Glass Ionomer Materials?

A long-time drawback of dental composites is that they accumulate more biofilms and plaques than amalgam and glass ionomer restorative materials. It would be highly desirable to develop a new composite with reduced biofilm growth, while avoiding the non-esthetics of amalgam and low strength of glass ionomer. The objectives of this study were to: (1) develop a protein-repellent composite with reduced biofilms matching amalgam and glass ionomer for the first time; and (2) investigate their protein adsorption, biofilms, and mechanical properties. Five materials were tested: A new composite containing 3% of protein-repellent 2-methacryloyloxyethyl phosphorylcholine (MPC); the composite with 0% MPC as control; commercial composite control; dental amalgam; resin-modified glass ionomer (RMGI). A dental plaque microcosm biofilm model with human saliva as inoculum was used to investigate metabolic activity, colony-forming units (CFU), and lactic acid production. Composite with 3% MPC had flexural strength similar to those with 0% MPC and commercial composite control (p > 0.1), and much greater than RMGI (p < 0.05). Composite with 3% MPC had protein adsorption that was only 1/10 that of control composites (p < 0.05). Composite with 3% MPC had biofilm CFU and lactic acid much lower than control composites (p < 0.05). Biofilm growth, metabolic activity and lactic acid on the new composite with 3% MPC were reduced to the low level of amalgam and RMGI (p > 0.1). In conclusion, a new protein-repellent dental resin composite reduced oral biofilm growth and acid production to the low levels of non-esthetic amalgam and RMGI for the first time. The long-held conclusion that dental composites accumulate more biofilms than amalgam and glass ionomer is no longer true. The novel composite is promising to finally overcome the major biofilm-accumulation drawback of dental composites in order to reduce biofilm acids and secondary caries.

From Mouth to Model: Combining in vivo and in vitro Oral Biofilm Growth.

Background: Oral biofilm studies based on simplified experimental setups are difficult to interpret. Models are limited mostly by the number of bacterial species observed and the insufficiency of artificial media. Few studies have attempted to overcome these limitations and to cultivate native oral biofilm.Aims: This study aimed to grow oral biofilm in vivo before transfer to a biofilm reactor for ex situ incubation. The in vitro survival of this oral biofilm and the changes in bacterial composition over time were observed.Methods: Six human enamel-dentin slabs embedded buccally in dental splints were used as biofilm carriers. Fitted individually to the upper jaw of 25 non-smoking male volunteers, the splints were worn continuously for 48 h. During this time, tooth-brushing and alcohol-consumption were not permitted. The biofilm was then transferred on slabs into a biofilm reactor and incubated there for 48 h while being nourished in BHI medium. Live/dead staining and confocal laser scanning microscopy were used to observe bacterial survival over four points in time: directly after removal (T0) and after 1 (T1), 24 (T2), and 48 h (T3) of incubation. Bacterial diversity at T0 and T3 was compared with 454-pyrosequencing. Fluorescence in situ hybridization (FISH) was performed to show specific taxa. Survival curves were calculated with a specially designed MATLAB script. Acacia and QIIME 1.9.1 were used to process pyrosequencing data. SPSS 21.0 and R 3.3.1 were used for statistical analysis.Results: After initial fluctuations at T1, survival curves mostly showed approximation of the bacterial numbers to the initial level at T3. Pyrosequencing analysis resulted in 117 OTUs common to all samples. The genera Streptococcus and Veillonella (both Firmicutes) dominated at T0 and T3. They make up two thirds of the biofilm. Genera with lower relative abundance had grown significantly at T3. FISH analysis confirmed the pyrosequencing results, i.e., the predominant staining of Firmicutes.Conclusion: We demonstrate the in vitro survival of native primary oral biofilm in its natural complexity over 48 h. Our results offer a baseline for cultivation studies of native oral biofilms in (phyto-) pharmacological and dental materials research. Further investigations and validation of culturing conditions could also facilitate the study of biofilm-induced diseases.

Peptide KSL-W-Loaded Mucoadhesive Liquid Crystalline Vehicle as an Alternative Treatment for Multispecies Oral Biofilm.

Decapeptide KSL-W shows antibacterial activities and can be used in the oral cavity, however, it is easily degraded in aqueous solution and eliminated. Therefore, we aimed to develop liquid crystalline systems (F1 and F2) for KSL-W buccal administration to treat multispecies oral biofilms. The systems were prepared with oleic acid, polyoxypropylene (5) polyoxyethylene (20) cetyl alcohol (PPG-5-CETETH-20), and a 1% poloxamer 407 dispersion as the oil phase (OP), surfactant (S), and aqueous phase (AP), respectively. We characterized them using polarized light microscopy (PLM), small-angle X-ray scattering (SAXS), rheology, and in vitro bioadhesion, and performed in vitro biological analysis. PLM showed isotropy (F1) or anisotropy with lamellar mesophases (F2), confirmed by peak ratio quantification using SAXS. Rheological tests demonstrated that F1 exhibited Newtonian behavior but not F2, which showed a structured AP concentration-dependent system. Bioadhesion studies revealed an AP concentration-dependent increase in the system’s bioadhesiveness (F2 = 15.50 ± 1.00 mN·s) to bovine teeth blocks. Antimicrobial testing revealed 100% inhibition of multispecies oral biofilm growth after KSL-W administration, which was incorporated in the F2 aqueous phase at a concentration of 1 mg/mL. Our results suggest that this system could serve as a potential vehicle for buccal administration of antibiofilm peptides.

Antiplaque effect of essential oils and 0.2% chlorhexidine on an in situ model of oral biofilm growth: a randomised clinical trial.

ObjectiveTo evaluate the in situ antiplaque effect after 4 days of using of 2 commercial antimicrobial agents in short term on undisturbed plaque-like biofilm.Trial Design and ParticipantsAn observer-masked, crossover randomised clinical trial on 15 oral and systemically healthy volunteers between 20–30 years who were randomly and sequentially allocated in the same group which performed 3 interventions in different randomised sequences.InterventionThe participants wore an appliance in 3 different rinsing periods doing mouthwashes twice a day (1/0/1) with essential oils, 0.2% chlorhexidine or sterile water (negative control). At the end of each 4-day mouthwash period, samples were removed from the appliance. Posteriorly, after bacterial vital staining, samples were analysed using a Confocal Laser Scanning Microscope.Main Outcome MeasuresBacterial vitality, thickness and covering grade by the biofilm after 4 days of applying each of the mouthwashes.ResultsThe essential oils and the 0.2% chlorhexidine were significantly more effective than the sterile water at reducing bacterial vitality, thickness and covering grade by the biofilm. No significant differences were found between the 0.2% chlorhexidine and the essential oils at reducing the bacterial vitality (13.2% vs. 14.7%). However, the 0.2% chlorhexidine showed more reduction than the essential oils in thickness (6.5 μm vs. 10.0 μm; p<0.05) and covering grade by the biofilm (20.0% vs. 54.3%; p<0.001).ConclusionThe essential oils and 0.2% chlorhexidine showed a high antiplaque effect. Although the 0.2% chlorhexidine showed better results with regard to reducing the thickness and covering grade by the biofilm, both antiseptics showed a high and similar antibacterial activity.Clinical RelevanceDaily essential oils or 0.2% chlorhexidine mouthwashes are effective when reducing dental plaque formation in the short term. Although 0.2% chlorhexidine continues to be the “gold standard” in terms of antiplaque effect, essential oils could be considered a reliable alternative.Trial RegistrationClinicalTrials.gov NCT02124655

Plant Extracts of Psidium guajava, Mangifera and Mentha sp. inhibit the Growth of the Population of Single-species Oral Biofilm

The use of oral gargle has gained popularity recently. Despite being either alcohol-based or plant-based, the main purpose for the usage of oral gargle is to prevent the accumulation of heavy plaque or oral malodour, that usually caused by the growth of oral biofilm. In this study, the antimicrobial effect of Psidium guajava, Mangifera sp. and Mentha sp. was tested against the single-species biofilm consisting of Streptococcus sanguinis and Streptococcus mitis. The biofilms were allowed to grow on the saliva-coated glass beads in Nordini’s Artificial Mouth (NAM) model to represent the oral cavity for 24 hours. The saliva would form the experimental pellicle on the glass beads. The results obtained showed that biofilm of Streptococcus mitis displayed a maximum adherence (11.53%) compared to Streptococcus sanguinis (1.83%) on the untreated experimental pellicle. When the aqueous extraction of plant extract was applied on the experimental pellicle, the bacterial adherence was significantly reduced to 1.54% (Streptococcus mitis) and 0.21% (Streptococcus sanguinis). This result indicates that the selected plant extracts can be used to inhibit the heavy growth of oral biofilm.

Oral multispecies biofilm development and the key role of cell–cell distance

Growth of oral bacteria in situ requires adhesion to a surface because the constant flow of host secretions thwarts the ability of planktonic cells to grow before they are swallowed. Therefore, oral bacteria evolved to form biofilms on hard tooth surfaces and on soft epithelial tissues, which often contain multiple bacterial species. Because these biofilms are easy to study, they have become the paradigm of multispecies biofilms. In this Review we describe the factors involved in the formation of these biofilms, including the initial adherence to the oral tissues and teeth, cooperation between bacterial species in the biofilm, signalling between the bacteria and its role in pathogenesis, and the transfer of DNA between bacteria. In all these aspects distance between cells of different species is integral for oral biofilm growth.