Papers by Keyword: Biofilm

Abstract: This investigation assessed how incorporating vanillin into a dental adhesive influences bacterial adhesion and biofilm formation. Experimental adhesives, formulated with 0.5% and 1.0% vanillin, were compared against a vanillin-free adhesive and a control using a multi-species biofilm model. No significant differences in bacterial adhesion were observed across the groups, with optical density (OD) values ranging from 0.28 to 0.40. In contrast, adhesives containing vanillin demonstrated a marked reduction in biofilm formation. After 48 hours, the strongest inhibition was recorded for the 1.0% vanillin adhesive (0.63 ± 0.07), followed by the 0.5% vanillin adhesive (0.77 ± 0.04), which were significantly lower than the vanillin-free (1.19 ± 0.08) and control (1.68 ± 0.05) groups. While biofilm accumulation increased in all groups by 72 hours, the vanillin-containing adhesives consistently maintained lower values than the controls. A concentration-dependent effect was confirmed by percent reduction analysis, which showed the 1.0% vanillin adhesive lowered biofilm by 62.5% at 48 hours and 45.0% at 72 hours. Vanillin also suppressed acidogenicity, as reflected by significantly higher culture medium pH values that remained near neutral at both 48 and 72 h, while the vanillin-free adhesive and control dropped below the critical pH 5.5 threshold. These findings suggest that vanillin effectively suppresses biofilm development without impacting bacterial adhesion, potentially serving as a natural additive to enhance the antibacterial properties of dental adhesives and minimize the risk of secondary caries.

Abstract: Secondary caries is a type of carious lesion found at the margins of or next to an existing restoration after the filling has been used for a period of time. It generally arises from the formation of defects or cracks in the filling material after restoration. This can create gaps between the material and the tooth tissue, which will allow bacteria in the biofilm to enter the interface. Dental adhesives are commonly used to provide retention for composite cement or filling materials. A good adhesive should be able to prevent leakage along the restoration margin as well as resist the mechanical load of chewing pressure. Recently, the inclusion of calcium in the adhesive monomer has been produced as Bio-Coat Ca, and its antimicrobial property against some oral bacteria has been studied. No information was found on anaerobes. The aim of this study was to evaluate the antimicrobial potential of dental adhesive on the biofilm formation of anaerobic bacteria involved in secondary caries. An adhesive containing CMET (calcium salt of 4-methacryloxyethyl trimellitic acid) and 10-methacryloyloxydecyl dihydrogen calcium phosphate (MDCP) (Bio-Coat Ca, Sun Medical, Moriyama, Shiga, Japan) was applied to the flat-bottom surface of the saliva-coated 96-well plate. Then it was polymerized with LED light at 460 nm and sterile with UV light. Porphyromonas gingivalis ATCC 33277, Prevotella intermedia ATCC 25611, and Fusobacterium nucleatum ATCC 25586 were prepared as a suspension of approximately 1 × 10⁸ CFU/mL and added to the well. The plate was left for 120 min at 37°C in a shaking incubator (120 r/min) to allow bacterial adhesion. After removing non-adherent cells, Schaedler broth was added and further incubated for 48-72 h to grow the biofilm. The culture medium was changed every 24 h. A biofilm formed on a 96-well plate surface without the adhesive was set up as a control. The amount of vital biofilm was assessed by the WST Microbial Cell Counting Kit (Dojindo Molecular Technologies, USA). All tests were triplicated performed and repeated three times. As a statistical analysis, the Mann-Whitney U test was applied. The results showed that dental adhesive exhibited significant anti-biofilm formation of P. gingivalis and F. nucleatum at a percent inhibition of 56% and 46%, respectively. On the other hand, no significant effect was found on P. intermedia. This was similar to our previous report on bacteria associated with primary caries, which revealed that the anti-biofilm effect of Bio-Coat Ca adhesive on Streptococcus mutans was 65% while no significant suppressive action was observed Lactobacillus casei and Actinomyces viscosus. The inhibitory effect of the adhesive was proposed to be the acidic characteristic of the monomers. This newly developed adhesive could be a promising material for the prevention of secondary caries. However, this study was done on the single-species biofilm formation in vitro and conducted in a short time. Long-term clinical studies are needed to evaluate the effect on the patients.

Abstract: Dentures are prosthetic device made of polymethyl methacrylate acrylic (PMMA) resin generally used by elderly people to replace their real teeth. However, the porosity and surface irregularities of acrylic resin are undesirable aspects of the denture base that can support colonization of oral microorganisms on the denture surface, particularly those associated with malodor. Oral malodor is a common and upsetting situation frequently found in denture wearers due to inadequate denture cleaning. A large number of strategies had been employed to minimize the potential bacterial pathogens, including mechanical brushing and using chemical agents. Therefore, we have developed denture PMMA resin containing vanillin and evaluated antimicrobial potential against four species of bacteria related to oral malodor. Denture base resin samples were prepared with dimensions of 10 mm x 10 mm x 1 mm. according to the amount of incorporating vanillin concentrations (0.1%, 0.5% and 1.0% w/w) and coated with saliva. Then, the biofilm formation of four species of bacteria (Porphyromonas gingivalis ATCC 33277, Aggregatibacter actinomycetemcomitans ATCC 33384, Fusobacterium nucleatum ATCC 25586, and Actinomyces viscosus ATCC 19246) was performed on resin samples in anaerobically condition for 48-72 h. Denture resin sample without vanillin was prepared in the same way and used as a control. The quantity of vital biofilm was assessed by WST Microbial Cell Counting Kit (Dojindo Molecular Technologies, USA). Kruskal-Wallis test followed by Dunn’s method were applied for statistical analysis. The results showed that in the groups of 0.5% and 1.0% w/w vanillin, significant decreases of biofilm mass were observed in all tested bacteria (P. gingivalis, A. actinomycetemcomitans, and F. nucleatum) compared with controls, except for A. viscosus. Approximately 30% to 38% bacterial biofilm reduction was demonstrated at 1.0% w/w vanillin on three species of bacteria, whereas 7% to 27% biofilm decreases were observed at 0.5% vanillin. No significant biofilm reduction was observed in A. viscosus. This conforms to our previous studies which revealed the antimicrobial properties of denture PMMA resin containing vanillin on respiratory pathogens and oral yeast. Denture base resin consisting of vanillin may be considered as a promising dental appliance for the use to prevent or solve the problem of oral malodor in patients who wear dentures.

Abstract: Orthodontic appliance made of resin is one of the main factors involving the shift of oral microbial community towards the source of pathogens by providing an extra site for adhesion and attachment of microorganisms to form biofilm associated with infections of multiple organs including respiratory tract. As a result, there is a demand for seeking of oral appliances with antimicrobial properties to reduce the risk of these infections. Here, vanillin (a principal flavoring agent with its antimicrobial property) has been incorporated to orthodontic resin and investigated the effect on biofilm formation of respiratory pathogens. The self-curing PMMA orthodontic resin samples (Siam Cement Group, Thailand) were fabricated depending on the percentage of vanillin adding (0.1% and 0.5% w/w vanillin). Resin without vanillin was used as a control. All samples were coated with sterile saliva collected from healthy volunteers. The bacterial biofilm formation assay was done by adding suspension (10⁷ colony forming unit/mL) of Staphylococcus aureus ATCC 5638, Staphylococcus epidermidis ATCC 14990, Streptococcus pneumoniae ATCC 49619, and Pseudomonas aeruginosa ATCC 27853 to the samples and incubated in shaking incubator (120 r/min) for 120 min at 37 °C to allow the adhesion of the bacteria. Afterwards, non-adherent bacterial cells were washed out and Brain Heart Infusion broth was added and further incubated for 24 h to enable the biofilm formation. The amount of vital biofilm was quantified by Cell Counting Kit WST-8 (Dojindo Molecular Technologies, USA). It was found that the inclusion of vanillin to resin decreased the biofilm formation of S. aureus, S. epidermidis, and P. aeruginosa whereas no effect was observed on S. pneumoniae. Both the 0.1% and 0.5% vanillin concentrations could significantly inhibit the biofilm of S. aureus and P. aeruginosa whereas only 0.5% vanillin showed the inhibitory effect on S. epidermidis. Up to 40% biofilm reduction was noticed with S. epidermidis and P. aeruginosa while that of S. aureus was about 30%. In conclusion, the present data demonstrate that the development of orthodontic PMMA resin incorporated with vanillin may be a possible candidate for fabricating of oral appliance with antimicrobial property against respiratory pathogens. This suggests that it could be useful for the prevention or management of respiratory infection related to bacterial biofilm formed on oral appliances.

Abstract: Glycerol is a by-product produced from biodiesel production through the transesterification process. The excessive amount of glycerol generated during this process may become an environmental problem since it cannot be disposed on the environment. One of the possible applications is its use in biofilm production as a plasticizer. This research aims to characterize the physicochemical properties of biofilm produced from Dioscorea hispida (Ubi Gadong) starch with a different formulation of glycerol extracted from recycling cooking oil. Dioscorea hispida starch has shown great potential as a major component in bioplastic production due to its faster degradation rate, while glycerol acted as a plasticizer. The contact angle, water absorption, water content, and degradation rate of biofilm were also studied. Biofilm with the highest ratio of glycerol content showed the highest water absorption, which achieved 75.75%, and the biodegradable rate of biofilm was 97.99% on day 3. The lowest glycerol content in biofilm showed the lowest contact angle and completely biodegradation after five days buried in the soil.

Abstract: Saliva plays a crucial role in oral health. Reduced salivary flow (hyposalivation) and xerostomia (dry mouth) are commonly found in many groups of patients, such as cancer patients with head and neck radiotherapy, patients taking medications with antisialogogic effects, diabetic patients, autoimmune patients, and the elderly. When salivary function is diminished, there is more risk of patients to develop opportunistic infections. One of therapeutic methods to restore lost or improve existing functions of natural saliva is the use of saliva substitutes. Several types of salivary substitutes have been developed to simulate human saliva as well as having antimicrobial property. Vanillin, an extensive flavor and aromatic component used worldwide, has been revealed to have antimicrobial potential against many types of bacteria yeasts and molds. The objective of this study was to evaluate the antimicrobial potential of vanillin-incorporated artificial saliva against biofilm formation of Candida. Artificial saliva containing different concentrations of vanillin (8, 4, 2, 1, 0.5 and 0.25 mg/mL) was prepared. Candida albicans ATCC 10231 and a clinical strain isolated from oral lesion of patient were used in this study. The Candida biofilm formation assay was performed on a 96-well plate by adding 150 μl 10⁷colony forming unit (CFU)/ml of each Candida suspension and 50 μl of artificial saliva and incubated at 37°C in shaking incubator for 24 h. The vital Candida in biofilm formed on the bottom surface of microwell was determined by WST-8 microbial viability assay kit (Dojindo, USA). Artificial saliva without vanillin was used as a control. All tests were done in triplicate and repeated three times. Kruskal-Wallis and Dunn’ tests were used as statistical analysis. It was found that in the presence of vanillin (at concentrations > 0.5 mg/mL) the amount of vital biofilm formation of both strains of C. albicans was significantly reduced (31-56%) compared with a control. The suppressive activity was not different between both strains of Candida. Similarly, dose dependent effects were found except at 2 mg/mL and 1 mg/mL. The result was conformed to the previous study of our group that the incorporation of vanillin to surgical obturator resin could reduce the Candida biofilm formation to nearly 46-54%. The mechanisms of antifungal property of vanillin had been demonstrated to be the induction of oxidative stress, and an interference of metabolic pathways. In conclusion, artificial saliva containing vanillin exhibited a distinct inhibitory effect on biofilm formation of C. albicans as demonstrated in the present study. Attention is required for further studies to implement vanillin added artificial saliva to be used as a productive saliva substitute for the prevention or treatment of Candida infection.

Abstract: The proportion of older people within the general population is expanding due to the decline of birth rate and the increase of life expectancy. Increasing elderly populations, which is retaining its teeth longer, leads to oral health problems, for example, root caries. The main etiological factor for the initiation and development of root caries is the appearance of cariogenic biofilm. The acid-producing and acid-tolerant bacteria and oral yeast, such as Candida albicans are the major contributors in root caries formation. A new generation of adhesives is developed with several modifications for the prevention of dental caries by incorporating antimicrobial components. The aim of this study was to investigate the antimicrobial effect of adhesive containing calcium salt of acidic monomers on the biofilm formation of oral Candida related to root caries. In the experiments, the flat-bottom surfaces of 96-well plate were painted with an adhesive containing calcium salts of 4-methacryloxyethyl trimellitic acid (CMET) and 10-methacryloyloxydecyl dihydrogen calcium phosphate (MDCP) (Bio-Coat, CA, Sun Medical, Japan). Then they were LED light-cured and coated with sterile saliva at 37 °C for 60 min. The biofilm formation was made by adding of Candida albicans (ATCC 10238 and two clinical strains) suspensions (10⁷ colony forming unit/mL) and incubated at 37°C for 24 h. The amount of vital biofilm was determined by WST-8 Microbial Cell Counting Kit (Dojindo Molecular Technologies, USA). All experiments were done in triplicate and repeated three times. Statistical analysis was performed using Student’s t-test. The results clearly showed that adhesive could significantly inhibit biofilm formation of all tested Candida compared with a control. This suppressive effect was not different among the strains of Candida. The percentages of vital biofilm reduction were 25% to 40%. The ability of this adhesive to suppress biofilm of oral yeast may be its antimicrobial property of acidic monomer or the effect of calcium ion within the adhesive, which can alter Candida cell morphology, and influence their structures or process of biofilm formation. In conclusion, an adhesive containing calcium salts of acidic monomers could significantly inhibit biofilm formation of C. albicans. This adhesive could be effectively applied to exposed root surfaces to prevent or inhibit the progression of root caries. Further studies are necessary to clarify the effect on multispecies biofilm, on long-term activity, and in vivo conditions.

Abstract: Early childhood caries (ECC), or baby bottle tooth decay is one of the most common chronic diseases that affect young children at 6 years of age or younger. The disease is resulted from complex interactions between tooth structures and cariogenic microorganisms in dental biofilm after exposure to fermentable carbohydrates through improper feeding practices. This leads to a rapid development of demineralization of teeth shortly after they erupt in the oral cavity. Candida albicans have been reported recently to be associated with the pathogenesis of ECC. One of the general approaches currently used for the prevention of ECC is the application of dental sealant to occlusal (chewing surface) pits and fissures of caries-susceptible teeth. Many types of dental sealants have been extensively used and various antimicrobial agents were introduced to dental sealants to increase their caries preventive effect. The aim of the present study was to evaluate the antimicrobial effect of vanillin incorporated resin-based dental sealant on C. albicans biofilm. Resin-based dental sealants (Clinpro; 3M ESPE, USA) containing different concentrations of vanillin (0%, 0.5%, 1% and 5% w/v) were prepared in 96-well plate. The suspension (10⁷ colony forming unit/mL) of each strain of Candida albicans (ATCC 10231, ATCC 18804 and 2 clinical isolates) was added to saliva-coated sealant resin samples and incubated at 37°C in 5% CO₂ atmosphere for 24 h to allow the Candida biofilm formation. The quantity of vital biofilm was determined by WST Microbial Cell Counting Kit (Dojindo Molecular Technologies, USA) at 460 nm. All tests were done in triplicate and repeated three times. Kruskal-Wallis test followed by Dunn’s method were used to account for multiple comparisons. Statistical significance was set at p-value less than 0.05. The results showed that 0.5-5% vanillin-incorporated dental sealant resins showed a significant suppressive effect against all strains of Candida. No significant differences of anti-biofilm effects were found among Candida strains. Approximately 30-50% Candida biofilm reduction was noticed in sealant resins containing vanillin compared with controls without vanillin. In conclusion, the adding of vanillin (0.5-5% w/v) to dental sealant resins is effective in inhibiting biofilm formation of fungus related to early childhood caries, C. albicans. Therefore, the use of dental sealants containing vanillin has the potential to prevent this type of dental caries in young children and biofilm formation by oral Candida. Further clinical studies are required to investigate physical properties of this sealant and clinical evaluation in the patients.

Abstract: Tooth decay or dental caries is an important oral health problem involving people of all age groups. The disease is the outcome of the demineralize process in which aciduric and acidogenic bacteria in a biofilm decompose tooth structure. Dental sealant, a resin material, which is applied on the occlusal pit and fissure surfaces of the teeth as a protective layer has been commonly used to prevent dental caries. However, the microbial effect on food residue is found to be a major cause of microleakage of sealant and secondary caries. Several types of antimicrobial agents were introduced to increase the caries preventive effect of dental sealants. Vanillin, the main component of flavoring agent vanilla, has been found to have antimicrobial property against Gram-positive and Gram-negative bacteria. The objective of this study was to investigate the antimicrobial effect of vanillin-incorporated dental sealant against biofilm formation of cariogenic bacteria. Dental sealant resin samples (Clinpro; 3M ESPE, USA) were prepared in 96-well plate in accordance with the amount of vanillin adding (0%, 0.5%, 1% and 5% vanillin). The cariogenic bacterial suspensions of Streptococcus mutans ATCC 25175 and Lactobacillus casei ATCC 334 were added to saliva-coated samples and incubated at 37°C in 5% CO2 atmosphere for 48 h to allow the biofilm formation. The quantity of vital biofilm was determined by WST Microbial Cell Counting Kit (Dojindo Molecular Technologies, USA) at 460 nm. One-way ANOVA and Tukey’s test were applied to the statistical analysis. A significant inhibitory effect against L. casei biofilm was observed in all vanillin incorporated samples (0.5%, 1% and 5% vanillin) compared with samples without vanillin. The percentage of biofilm reduction was 32-39%. For S. mutans, the suppressive effect was noticed only at >1% vanillin with 18-25% biofilm reduction. In conclusion, the incorporation of vanillin to dental sealants could decrease biofilm formation of cariogenic bacteria (S. mutans and L. casei). The use of dental sealants containing vanillin could be a promising measure to prevent dental caries due to their antibacterial biofilm formation property.

Abstract: The use of natural polymeric materials has been growing notably in order to replace packaging from non-renewable sources. In this sense, cassava starch is a very promising natural polymer for this purpose due to its ease of production, the low cost, besides being biodegradable. However, cassava starch biofilms when dried have a brittle character requiring the addition of a plasticizer. Thus, biofilms were synthesized based on cassava starch (3%) with different percentages of glycerol (5%, 10%, 20%, 40% and 50%) to evaluate changes in physic-chemical and mechanical properties. The results indicate that the increase in percentage of glycerol contributed directly to the increase of water vapor permeability while decreases the contact angle and modulus of elasticity of cassava starch films.