Cariogenic Streptococcus Mutans Research Articles

Antibacterial activities of Miang extracts against selected pathogens and the potential of the tannin-free extracts in the growth inhibition of Streptococcus mutans.

Bacterial pathogens have remained a major public health concern for several decades. This study investigated the antibacterial activities of Miang extracts (at non-neutral and neutral pH) against Bacillus cereus TISTR 747, Escherichia coli ATCC 22595, Salmonella enterica serovar Typhimurium TISTR 292 and Streptococcus mutans DMST 18777. The potential of Polyvinylpolypyrrolidone (PVPP)-precipitated tannin-free Miang extracts in growth-inhibition of the cariogenic Streptococcus mutans DMST 18777 and its biofilms was also evaluated. The tannin-rich fermented extracts had the best bacterial growth inhibition against S. mutans DMST 18777 with an MIC of 0.29 and 0.72 mg/mL for nonfilamentous fungi (NFP) Miang and filamentous-fungi-processed (FFP) Miang respectively. This observed anti-streptococcal activity still remained after PVPP-mediated precipitation of bioactive tannins especially, in NFP and FFP Miang. Characterization of the PVPP-treated extracts using High performance liquid chromatography quadrupole-time of flight-mass spectrometry (HPLC-QToF-MS) analysis, also offered an insight into probable compound classes responsible for the activities. In addition, Crystal violet-staining also showed better IC50 values for NFP Miang (4.30 ± 0.66 mg/mL) and FFP Miang (12.73 ± 0.11 mg/mL) against S. mutans DMST 18777 biofilms in vitro. Homology modeling and molecular docking analysis using HPLC-MS identified ligands in tannin-free Miang supernatants, was performed against modelled S. mutans DMST 18777 sortase A enzyme. The in silico analysis suggested that the inhibition by NFP and FFP Miang might be attributed to the presence of ellagic acid, flavonoid aglycones, and glycosides. Thus, these Miang extracts could be optimized and explored as natural active pharmaceutical ingredients (NAPIs) for applications in oral hygienic products.

Epigallocatechin gallate reduces the virulence of cariogenic Streptococcus mutans biofilm by affecting the synthesis of biofilm matrix components

IntroductionThere have been reports on the effects of epigallocatechin gallate (EGCG) against Streptococcus mutans viability and acidogenesis. However, the effects of EGCG on the virulence of S. mutans biofilm development have yet to be fully investigated using validated cariogenic biofilm models. ObjectiveThus, this study aimed to evaluate the effects of EGCG on S. mutans biofilm virulence using a validated cariogenic model and clinically relevant treatment regimens, twice a day for 1.5 min. MethodsEffects of EGCG on bacterial viability, polyssacharide synthesis and biofilm acidogenesis were evaluated. The morphology and 3D structure of the biofilms were evaluated by scanning electron (SEM) and confocal laser scanning microscopy, respectively. ResultsNo significant change in S. mutans viability or culture medium pH were observed when comparing EGCG-treated and NaCl-treated biofilms. EGCG significantly reduced the accumulation of soluble and insoluble polysaccharides, resulting in the formation of a biofilm with interspaced exopolysaccharide-microcolony complexes unevenly distributed on enamel. The SEM images of the biofilm treated with EGCG depict multilayers of cells arranged in short chains of microorganisms adhered to an unstructured matrix, which is not continuous and does not enmesh or protect the microorganisms entirely. Importantly, confocal images demonstrated that treatment with EGCG affected the 3D structure and organization of S. mutans biofilm, which presented a biofilm matrix more confined to the location of the microcolonies. ConclusionIn conclusion, EGCG lowered the virulence of S. mutans matrix-rich biofilm by reducing the synthesis of biofilm matrix components, altering the biofilm matrix structure, organization, and distribution.

Antibiofilm and Antibacterial Properties of Herbal Extracts as Alternatives to Current Treatment Approaches: A Narrative Review

Bacterial biofilm formation poses significant challenges in the healthcare sector due to increased antibiotic resistance and persistent infections. This literature review explores the potential of some herbs and their extracts as alternative approaches to combat biofilm formation and multidrug-resistant bacteria. A detailed literature search was conducted across databases for published studies till 2023, to identify studies on medicinal plants' anti-biofilm and antibacterial properties. Key compounds within plant extracts showing anti-biofilm activity and their mechanisms of action were highlighted. A combination of keywords, MeSH terms, and Boolean operators were used to formulate the search strategy. Numerous studies demonstrated the efficacy of medicinal plants in inhibiting biofilm formation and combating multidrug-resistant bacteria. Active compounds such as benzyl (6Z,9Z,12Z)-6,9,12-octadecatrienoate, 3-benzyloxy-1-nitro-butan-2-ol, Pyridine, 3-(1-methyl-2-pyrrolidinyl)-(S), and others exhibited anti-biofilm and antibacterial potential. Extracts from Berginia ciliata, Clematis grata, and Clematis viticella showed over 80% inhibition of biofilm formation, while mango leaf extracts interfered with quorum sensing mechanisms in Pseudomonas aeruginosa PAO1. Salvadora persica extracts displayed significant biofilm inhibition against cariogenic Streptococcus mutans isolates. Medicinal plants and their extracts hold promise as alternative strategies to combat bacterial biofilms and multidrug-resistant bacteria. The identification of active compounds provides opportunities for further research and drug development. Molecular docking studies are crucial for understanding the molecular interactions between these compounds and bacterial targets, guiding the design of effective antibacterial agents based on natural compounds. Further research, including preclinical and clinical trials, is essential to validate the safety and efficacy of these extracts and their compounds for practical application in healthcare.

Dental composite biodeterioration in the presence of oral Streptococci and extracellular metabolic products

ObjectiveSecondary caries is a primary cause of early restoration failure. While primary dental caries has been extensively researched, our knowledge about the impact of secondary caries on dental restorations is relatively limited. In this study, we examined how different clinically relevant microbially-influenced environments impact the degradation of nano-filled (FIL) and micro-hybrid (AEL) dental composites. MethodsMaterial strength of two commercial dental composites was measured following incubation in aqueous media containing: i) cariogenic (Streptococcus mutans) and non-cariogenic bacteria (Streptococcus sanguinis) grown on sucrose or glucose, ii) abiotic mixtures of artificial saliva and sucrose and glucose fermentation products (volatile fatty acids and ethanol) in proportions known to be produced by these microorganisms, and iii) abiotic mixtures of artificial saliva and esterase, a common oral extracellular enzyme. ResultsNano-filled FIL composite strength decreased in all three types of incubations, while micro-hybrid AEL composite strength only decreased significantly in biotic incubations. The strength of both composites was statistically significantly decreased in all biotic incubations containing both cariogenic and non-cariogenic bacteria beyond that induced by either abiotic mixtures of fermentation products or esterase alone. Finally, there were no statistically significant differences in composite strength decrease among the tested biotic conditions. ConclusionsThe results show that conditions created during the growth of both cariogenic and non-cariogenic oral Streptococci substantially reduce commercial composite strength, and this effect warrants further study to identify the mechanism(s). Clinical SignificanceDental biofilms of oral Streptococci bacteria significantly affect the mechanical strength of dental restorations.

Combating Cariogenic Streptococcus mutans Biofilm Formation and Disruption with Coumaric Acid on Dentin Surface.

Streptococcus mutans, the primary cause of dental caries, relies on its ability to create and sustain a biofilm (dental plaque) for survival and pathogenicity in the oral cavity. This study was focused on the antimicrobial biofilm formation control and biofilm dispersal potential of Coumaric acid (CA) against Streptococcus mutans on the dentin surface. The biofilm was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) viability assay, microtiter plate assay, production of extracellular polymeric substances (EPSs), florescence microscopy (surface coverage and biomass μm2) and three-dimensional (3D) surface plots. It was observed that CA at 0.01 mg/mL reduced bacterial growth by 5.51%, whereases at 1 mg/mL, a significant (p < 0.05) reduction (98.37%) was observed. However, at 1 mg/mL of CA, a 95.48% biofilm formation reduction was achieved, while a 73.45% biofilm dispersal (after 24 h. treatment) was achieved against the preformed biofilm. The MTT assay showed that at 1 mg/mL of CA, the viability of bacteria in the biofilm was markedly (p < 0.05) reduced to 73.44%. Moreover, polysaccharide (EPS) was reduced to 24.80 μg/mL and protein (EPS) to 41.47 μg/mL. ImageJ software (version 1.54 g) was used to process florescence images, and it was observed that the biofilm mass was reduced to 213 (μm2); the surface coverage was reduced to 0.079%. Furthermore, the 3D surface plots showed that the untreated biofilm was highly dense, with more fibril-like projections. Additionally, molecular docking predicted a possible interaction pattern of CA (ligand) with the receptor Competence Stimulating Peptide (UA159sp, PDB ID: 2I2J). Our findings suggest that CA has antibacterial and biofilm control efficacy against S. mutans associated with dental plaque under tested conditions.

Measurement and analysis of microbial fluoride resistance in dental biofilm models.

The interactions between communities of microorganisms inhabiting the dental biofilm is a major determinant of oral health. These biofilms are periodically exposed to high concentrations of fluoride, which is present in almost all oral healthcare products. The microbes resist fluoride through the action of membrane export proteins. This chapter describes the culture, growth and harvest conditions of model three-species dental biofilm comprised of cariogenic pathogens Streptococcus mutans and Candida albicans and the commensal bacterium Streptococcus gordonii. In order to examine the role of fluoride export by S. mutans in model biofilms, procedures for generating a strain of S. mutans with a genetic knockout of the fluoride exporter are described. We present a case study examining the effects of this mutant strain on the biofilm mass, acid production and mineral dissolution under exposure to low levels of fluoride. These general approaches can be applied to study the effects of any gene of interest in physiologically realistic multispecies oral biofilms.

Chemical composition of essential oil from Lindera subumbelliflora Kosterm and its effect on the susceptibility and biofilm activities of Candida albicans and Streptococcus mutans

The chemical composition of the essential oil of Lindera subumbelliflora (Lauraceae) was investigated for the first time. The essential oil was obtained by hydrodistillation and fully characterised by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). The antifungal activity of L. subumbelliflora essential oil was tested against Candida albicans and Streptococcus mutans using the broth microdilution assay, whereas the microbial biofilms were determined using a semi-quantitative static biofilm. A total of 28 components (99.6%) were successfully identified, which were characterised by β-eudesmol (14.6%), cis-α-bergamotene (11.0%), α-copaene (8.5%), dodecen-1-ol (8.5%), and (E)-nerolidol (8.3%). The essential oil exhibited activity against Candida albicans and Streptococcus mutans with MIC values of 250 and 500 µg/mL, respectively. The essential oil increased the biofilm of Candida albicans by 38.25%, however, decreased the biofilm of Streptococcus mutans by 47.89% when treated with 500 µg/mL. Thus, the essential oil has a promising application in dentistry via inhibition of the growth of Candida albicans and Streptococcus mutans. However, the antibiofilm activity of the essential oil is only applicable for cariogenic Streptococcus mutans.

Novel Iron Chelators, Super-Polyphenols, Show Antimicrobial Effects against Cariogenic Streptococcus mutans.

Dental caries are an oral infectious disease that can affect human health both orally and systemically. It remains an urgent issue to establish a novel antibacterial method to prevent oral infection for a healthy life expectancy. The aim of this study was to evaluate the inhibitory effects of novel iron chelators, super-polyphenols (SPs), on the cariogenic bacterium Streptococcus mutans, in vitro. SPs were developed to reduce the side effects of iron chelation therapy and were either water-soluble or insoluble depending on their isoforms. We found that SP6 and SP10 inhibited bacterial growth equivalent to povidone-iodine, and viability tests indicated that their effects were bacteriostatic. These results suggest that SP6 and SP10 have the potential to control oral bacterial infections such as Streptococcus mutans.

Effects of Sulforaphene on the Cariogenic Properties of Streptococcus Mutans In Vitro and Dental Caries Development In Vivo.

Sulforaphene (SFE) is a common nutritional supplement with antibacterial, anti-cancer, and anti-inflammatory effects. However, the effects of SFE on the cariogenicity of Streptococcus mutans and dental caries have not been reported. The objectives of this study were to investigate the caries-controlling potential of SFE. The effects of SFE on S. mutans were investigated using the broth microdilution method, crystal violet staining, SEM observation, acid tolerance assays, lactic acid quantification, and polysaccharide measurements. A rat caries model was established to evaluate the caries-controlling effects and biocompatibility of SFE in vivo. SFE inhibited S. mutans growth and biofilm formation. Furthermore, SFE restrained the cariogenic properties of S. mutans, including its acid production, acid tolerance, and extracellular polysaccharide production, without affecting the bacterial viability at sub-inhibitory levels. In the rat caries model, SFE significantly arrested the onset and development of dental caries. Moreover, no visible hemolytic phenomenon or cytotoxicity was detected in the SFE groups. After four weeks of SFE treatment, all rats remained in apparent good health with no significant differences in weight gain; their hemogram and biochemical parameters were normal; no pathological changes were observed in the oral mucosa, liver, or kidneys. In conclusion, SFE was safe and inhibited the development of caries effectively.

Anticariogenic Activity of Celastrol and Its Enhancement of Streptococcal Antagonism in Multispecies Biofilm.

Dental caries is a chronic disease resulting from dysbiosis in the oral microbiome. Antagonism of commensal Streptococcus sanguinis and Streptococcus gordonii against cariogenic Streptococcus mutans is pivotal to keep the microecological balance. However, concerns are growing on antimicrobial agents in anticaries therapy, for broad spectrum antimicrobials may have a profound impact on the oral microbial community, especially on commensals. Here, we report celastrol, extracted from Traditional Chinese Medicine's Tripterygium wilfordii (TW) plant, as a promising anticaries candidate. Our results revealed that celastrol showed antibacterial and antibiofilm activity against cariogenic bacteria S. mutans while exhibiting low cytotoxicity. By using a multispecies biofilm formed by S. mutans UA159, S. sanguinis SK36, and S. gordonii DL1, we observed that even at relatively low concentrations, celastrol reduced S. mutans proportion and thereby inhibited lactic acid production as well as water-insoluble glucan formation. We found that celastrol thwarted S. mutans outgrowth through the activation of pyruvate oxidase (SpxB) and H2O2-dependent antagonism between commensal oral streptococci and S. mutans. Our data reveal new anticaries properties of celastrol that enhance oral streptococcal antagonism, which thwarts S. mutans outgrowth, indicating its potential to maintain oral microbial balance for prospective anticaries therapy.

Inhibition of biofilm formation and virulence factors of cariogenic oral pathogen Streptococcus mutans by natural flavonoid phloretin

ABSTRACT Objectives To evaluate the effect and mechanism of action of the flavonoid phloretin on the growth and sucrose-dependent biofilm formation of Streptococcus mutans. Methods Minimum inhibitory concentration, viability, and biofilm susceptibility assays were conducted to assess antimicrobial and antibiofilm effect of phloretin. Biofilm composition and structure were analysed with scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Water-soluble (WSG) and water-insoluble glucan (WIG) were determined using anthrone method. Lactic acid measurements and acid tolerance assay were performed to assess acidogenicity and aciduricity. Reverse transcription quantitative PCR (RT-qPCR) was used to measure the expression of virulence genes essential for surface attachment, biofilm formation, and quorum sensing. Results Phloretin inhibited S. mutans growth and viability in a dose-dependent manner. Furthermore, it reduced gtfB and gtfC gene expression, correlating with the reduction of extracellular polysaccharides (EPS)/bacteria and WIG/WSG ratio. Inhibition of comED and luxS gene expression, involved in stress tolerance, was associated with compromised acidogenicity and aciduricity of S. mutans. Conclusions Phloretin exhibits antibacterial properties against S. mutans, modulates acid production and tolerance, and reduces biofilm formation. Clinical significance Phloretin is a promising natural compound with pronounced inhibitory effect on key virulence factors of the cariogenic pathogen, S. mutans.

Assessment of microbial contamination in removable orthodontic appliances with and without the use of antimicrobial agents by checkerboard DNA-DNA hybridization analysis

This clinical, crossover, double-blind trial evaluated the microbial contamination of removable orthodontic appliances used by children and the efficacy of 0.12% chlorhexidine gluconate spray use for disinfection. Twenty children aged 7-11 years were instructed to wear removable orthodontic appliances for 1 week. They were instructed to use a placebo solution (control) or 0.12% chlorhexidine gluconate (experimental) to clean the appliances on days 4 and 7 after installation. After this period, the microbial contamination on the surfaces of the appliance was analyzed using checkerboard DNA-DNA hybridization for 40 bacterial species. Data were analyzed by Fisher exact, t, and Wilcoxon tests (α= 0.05). Removable orthodontic appliances were heavily contaminated by the target microorganisms. Streptococcus sanguinis, Streptococcus oralis, Streptococcus gordonii, and Eikenella corrodens were found in 100% of the appliances. Among cariogenic microorganisms, Streptococcus mutans and Streptococcus sobrinus were more abundant than Lactobacillus acidophilus and Lactobacillus casei. Red complex pathogens were more abundant than orange complex species. Purple complex bacteria were the most prevalent among bacterial complexes not associated with specific pathologies, detected in 34% of the samples. After the use of chlorhexidine, the number of cariogenic microorganisms (S. mutans, S. sobrinus, and L. casei) decreased significantly (P<0.05), and the numbers of periodontal pathogenic species from the orange and red complex also decreased significantly (P<0.05). There was no reduction for Treponema socranskii. Removable orthodontic appliances were densely contaminated by several bacterial species. Twice-a-week application of chlorhexidine spray effectively reduced cariogenic and orange and red complex periodontal pathogens.

Antagonistic activity of Lactobacillus spp. and Bifidobacterium spp. against cariogenic Streptococcus mutans in vitro and viability when added to chewing gum during storage.

The aim of the work was to evaluate antagonistic activity of Lactobacillus spp. and Bifidobacterium spp. in vitro against cariogenic Streptococcus mutans UA 159 and viability in chewing gum, during storage. Antagonistic activity was evaluated in vitro by the "spot on the lawn" test. Two bacteria were chosen and subjected to lyophilization and microencapsulation using the atomization method, containing polyvinylpyrrolidone polymer and lactose as encapsulating agents. For application in food matrices, four treatments were elaborated: chewing gum containing lyophilized B. lactis B94 (BLL), microencapsulated B. lactis B94 (BLE), lyophilized L. brevis (LBL), and microencapsulated L. brevis (LBE). Both microorganisms demonstrated a high capacity for inhibition against S. mutans, when compared to oral antiseptic chlorhexidine 0.2% in vitro, and according to the test of sensitivity profile to proteolytic enzymes, all the bacteria tested are producers of antimicrobial peptides, resulting in the inhibitory activity of the cariogenic bacterium. Furthermore, the viability of B. lactis B94 and L. brevis was maintained after microencapsulation, indicating that the process was efficient, with no significant difference (p < 0.05) between the results. And, in the chewing gum containing the bacteria during the storage period (33 days), it was found that cell immobilization did not significantly influence (p < 0.05) the counts of L. brevis but benefited the viability of B. lactis B94. Therefore, both probiotic bacteria are producers of antimicrobial substances with the ability to inhibit S. mutans, in vitro. The microencapsulation was considered efficient since it influenced the viability of B. lactis B94 (> 8 log CFU/g); however, the microencapsulation did not influence the viability of L. brevis since in both lyophilized and encapsulated form; the concentration of the bacteria remained above 8 log CFU/g during the storage period of the chewing gum.

Human-associated bacteria adopt an unusual route for synthesizing 3-acetylated tetramates for environmental adaptation

BackgroundTetramates or tetramic acid-containing compounds (TACs) are a group of bioactive natural products featuring a pyrrolidine-2,4-dione ring acknowledged being closed via Dieckmann cyclization. The cariogenic Streptococcus mutans strains bearing a muc biosynthetic gene cluster (BGC) can synthesize mutanocyclin (MUC), a 3-acetylated TAC that can inhibit both leukocyte chemotaxis and filamentous development in Candida albicans. Some strains can also accumulate reutericyclins (RTCs), the intermediates of MUC biosynthesis with antibacterial activities. However, the formation mechanism of the pyrrolidine-2,4-dione ring of MUC and the distribution of muc-like BGCs along with their ecological functions has not been explored extensively.ResultsWe demonstrated that a key intermediate of MUC biosynthesis, M-307, is installed by a hybrid nonribosomal peptide synthetase-polyketide synthase assembly line and its pyrrolidine-2,4-dione ring is closed via an unprecedented lactam bond formation style. Subsequent C-3 acetylation will convert M-307 to RTCs, which is then hydrolyzed by a deacylase, MucF, to remove the N-1 fatty acyl appendage to generate MUC. Distribution analysis showed that the muc-like BGCs distribute predominantly in human-associated bacteria. Interestingly, most of the muc-like BGCs possessing a mucF gene were isolated from human or livestock directly, indicating their involvement in alleviating the host’s immune attacks by synthesizing MUC; while those BGCs lacking mucF gene distribute mainly in bacteria from fermented products, suggesting that they tend to synthesize RTCs to compete with neighboring bacteria. It is noteworthy that many bacteria in the same habitats (e.g., the oral cavity) lack the muc-like BGC, but possess functional MucF homologues to “detoxify” RTCs to MUC, including several competitive bacteria of S. mutans. We also comparably studied the distribution of TAS1, a fungal enzyme responsible for the production of phytotoxic tenuazonic acids (TeAs), a class of 3-acetylated TACs with similar structure but distinct biosynthetic mechanism to MUC, and found that it mainly exists in plants or crops.ConclusionsThe in vivo and in vitro experiments revealed that the pyrrolidine-2,4-dione ring of MUC is closed via lactam bond formation, which may be adopted by many TACs without 3-acyl decorations. Besides, we found that muc-like BGCs are widespread in human-associated bacteria and their shapes and main products can be influenced by the habitat environment and vice versa. By comparing with TeAs, we provided thought-provoking insights into how ecological and evolutionary forces drive bacteria and fungi to construct a common 3-acetylated pyrrolidine-2,4-dione core through different routes, and how the biosynthetic processes are delicately controlled to generate diverse 3-acetylated TACs for environmental adaptation.2PMNhDpkP3itX1xdtgrn3CVideo

Antibiofilm Efficacies of Flavonoid-Rich Sweet Orange Waste Extract against Dual-Species Biofilms.

The current study evaluated the antibacterial properties of industrial sweet orange waste extracts (ISOWEs), which are a rich source of flavonoids. The ISOWEs exhibited antibacterial activity towards the dental cariogenic pathogens Streptococcus mutans and Lactobacillus casei with 13.0 ± 2.0 and 20.0 ± 2.0 mg/mL for MIC (minimum inhibitory concentration) and 37.7 ± 1.5 and 43.3 ± 2.1 mg/mL for MBC (minimum bactericidal concentration), respectively. When evaluated in a 7-day dual-species oral biofilm model, ISOWEs dose-dependently reduced the viable bacteria count, and demonstrated strong synergistic effects when combined with the anti-septic chlorhexidine (at 0.1 and 0.2%). Similarly, confocal microscopy confirmed the anti-cariogenic properties of ISOWEs, alone and in combination with chlorhexidine. The citrus flavonoids contributed differently to these effects, with the flavones (nobiletin, tangeretin and sinensetin) demonstrating significantly lower MICs and MBCs compared to the flavanones hesperidin and narirutin. In conclusion, our study demonstrated the potential of citrus waste as a currently underutilised source of flavonoids for antimicrobial applications, such as in dental health.

Stoichiometric models of sucrose and glucose fermentation by oral streptococci: Implications for free acid formation and enamel demineralization.

The objective of this study is to develop stoichiometric models of sugar fermentation and cell biosynthesis for model cariogenic Streptococcus mutans and non-cariogenic Streptococcus sanguinis to better understand and predict metabolic product formation. Streptococcus mutans (strain UA159) and Streptococcus sanguinis (strain DSS-10) were grown separately in bioreactors fed brain heart infusion broth supplemented with either sucrose or glucose at 37°C. Cell mass concentration and fermentation products were measured at different hydraulic residence times (HRT) to determine cell growth yield. Sucrose growth yields were 0.080±0.0078g cell/g and 0.18±0.031g cell/g for S. sanguinis and S. mutans, respectively. For glucose, this reversed, with S. sanguinis having a yield of 0.10±0.0080g cell/g and S. mutans 0.053±0.0064g cell/g. Stoichiometric equations to predict free acid concentrations were developed for each test case. Results demonstrate that S. sanguinis produces more free acid at a given pH than S. mutans due to lesser cell yield and production of more acetic acid. Greater amounts of free acid were produced at the shortest HRT of 2.5hr compared to longer HRTs for both microorganisms and substrates. The finding that the non-cariogenic S. sanguinis produces greater amounts of free acids than S. mutans strongly suggests that bacterial physiology and environmental factors affecting substrate/metabolite mass transfer play a much greater role in tooth or enamel/dentin demineralization than acidogenesis. These findings enhance the understanding of fermentation production by oral streptococci and provide useful data for comparing studies under different environmental conditions.

Dual function of anti-biofilm and modulating biofilm equilibrium of orthodontic cement containing quaternary ammonium salt.

The objectives of this study were to incorporate dimethylaminohexadecyl methacrylate (DMAHDM) into resin-modified glass ionomer cement (RMGI) to develop a novel orthodontic cement which endowed RMGI with strong antibacterial ability and investigated its modulation biofilm equilibrium from cariogenic state to non-cariogenic state for the first time. Cariogenic Streptococcus mutans (S. mutans), and non-cariogenic Streptococcus sanguinis (S. sanguinis) and Streptococcus gordonii (S. gordonii) were selected to form a tri-species biofilm model. RMGI incorporated with different mass fraction of DMAHDM was examined: biofilm colony-forming units, metabolic activity, live/dead staining, lactic acid and exopolysaccharides productions. TaqMan real-time polymerase chain reaction was used to determine changes of biofilm species compositions. The results showed RMGI containing 3% DMAHDM achieved strong antibacterial ability and suppressed the cariogenic species in biofilm, modulating biofilm equilibrium from cariogenic state to non-cariogenic state tendency. The novel bioactive cement containing DMAHDM is promising in fixed orthodontic treatments and protecting tooth enamel.

Anti-Bacterial and Anti-Biofilm Activities of Anandamide against the Cariogenic Streptococcus mutans

Streptococcus mutans is a cariogenic bacterium in the oral cavity involved in plaque formation and dental caries. The endocannabinoid anandamide (AEA), a naturally occurring bioactive lipid, has been shown to have anti-bacterial and anti-biofilm activities against Staphylococcus aureus. We aimed here to study its effects on S. mutans viability, biofilm formation and extracellular polysaccharide substance (EPS) production. S. mutans were cultivated in the absence or presence of various concentrations of AEA, and the planktonic growth was followed by changes in optical density (OD) and colony-forming units (CFU). The resulting biofilms were examined by MTT metabolic assay, Crystal Violet (CV) staining, spinning disk confocal microscopy (SDCM) and high-resolution scanning electron microscopy (HR-SEM). The EPS production was determined by Congo Red and fluorescent dextran staining. Membrane potential and membrane permeability were determined by diethyloxacarbocyanine iodide (DiOC2(3)) and SYTO 9/propidium iodide (PI) staining, respectively, using flow cytometry. We observed that AEA was bactericidal to S. mutans at 12.5 µg/mL and prevented biofilm formation at the same concentration. AEA reduced the biofilm thickness and biomass with concomitant reduction in total EPS production, although there was a net increase in EPS per bacterium. Preformed biofilms were significantly affected at 50 µg/mL AEA. We further show that AEA increased the membrane permeability and induced membrane hyperpolarization of these bacteria. AEA caused S. mutans to become elongated at the minimum inhibitory concentration (MIC). Gene expression studies showed a significant increase in the cell division gene ftsZ. The concentrations of AEA needed for the anti-bacterial effects were below the cytotoxic concentration for normal Vero epithelial cells. Altogether, our data show that AEA has anti-bacterial and anti-biofilm activities against S. mutans and may have a potential role in preventing biofilms as a therapeutic measure.

Photodynamic nano hydroxyapatite with biofilm penetration capability for dental plaque eradication and prevention of demineralization

Dental caries represents one of the most prevalent diseases worldwide, characteristic of the growth of dental plaque and demineralization of tooth enamel. Current medications for eradication of dental plaques and prevention of demineralization suffer from several limitations to overcome, calling for novel strategies with great potency in eliminating cariogenic bacteria and dental plaque that forms, as well as in inhibiting the demineralization of enamel, into an integrated system. Considering the potency of photodynamic therapy in bacteria inactivation and the composition of enamel, we herein report that the novel photodynamic nano hydroxyapatite (nHAP), named Ce6 @QCS/nHAP, was useful for this purpose. Ce6 @QCS/nHAP, comprised of quaternary chitosan (QCS)-coated nHAP loaded with chlorin e6 (Ce6), exhibited good biocompatibility and non-compromised photodynamic activity. In vitro studies revealed that Ce6 @QCS/nHAP could effectively associate with cariogenic Streptococcus mutans (S. mutans), leading to a significant antibacterial effect through photodynamic killing and physical inactivation against the planktonic microbe. Three-dimensional fluorescence imaging suggested that Ce6 @QCS/nHAP exhibited a superior S. mutans biofilm penetration capacity to free Ce6, resulting in effective dental plaque eradiation when light irradiation was applied. The number of surviving bacteria in biofilm was at least 2.8 log units lower in the Ce6 @QCS/nHAP group compared to that in the free Ce6 group. Further, in the S. mutans biofilm-infected artificial tooth model, treatment with Ce6 @QCS/nHAP also resulted in the significant prevention of hydroxyapatite disks from demineralization, with lower percentage of fragmentation and weight loss These data suggest that our photodynamic nanosystem can effectively eradicate dental plaque while also significantly protecting artificial tooth from demineralization, opening up new possibilities in treating bacterium-associated dental caries.

The Antibacterial Effect of Cannabigerol toward Streptococcus mutans Is Influenced by the Autoinducers 21-CSP and AI-2

Bacteria can communicate through an intercellular signaling system referred to as quorum sensing (QS). The QS system involves the production of autoinducers that interact with their respective receptors, leading to the induction of specific signal transduction pathways. The QS systems of the oral cariogenic Streptococcus mutans regulate the maturation of biofilms and affect its virulent properties. We have previously shown that the non-psychoactive compound cannabigerol (CBG) of the Cannabis sativa L. plant has anti-bacterial and anti-biofilm activities towards S. mutans. Here we were interested in investigating the effect of the two QS systems ComCDE and LuxS on the susceptibility of S. mutans to CBG and the anti-QS activities of CBG. This was assessed by using various comCDE and luxS mutant strains and complementation with the respective autoinducers, competence stimulating peptide (CSP) and (S)-4,5-dihydroxy-2,3-pentandione (DPD, pre-AI-2). We found that S. mutans comCDE knockout strains were more sensitive to the anti-bacterial actions of CBG compared to the WT strain. Exogenously added 21-CSP prevented the anti-bacterial actions caused by CBG on the ΔcomC, ΔcomE and ΔluxS mutants, while having no effect on the susceptibility of the WT and ΔcomCDE strains to CBG. Exogenously added DPD increased the susceptibility of WT and ΔluxS to CBG. Vice versa, CBG significantly reduced the 21-CSP-induced expression of comCDE genes and ComE-regulated genes and suppressed the expression of luxS with concomitant reduction in AI-2 production. DPD induced the expression of comCDE genes and ComE-regulated genes, and this induction was repressed by CBG. 21-CSP alone had no significant effect on luxS gene expression, while ΔcomCDE strains showed reduced AI-2 production. In conclusion, our study shows that the susceptibility of S. mutans to CBG is affected by the ComCDE and LuxS QS pathways, and CBG is a potential anti-QS compound for S. mutans. Additionally, we provide evidence for crosstalk between the ComCDE and LuxS QS systems.