In vitro effect of low-level laser therapy on typical oral microbial biofilms

The aim of this study was to evaluate the effect of specific parameters of low-level laser therapy (LLLT) on biofilms formed by Streptococcus mutans, Candida albicans or an association of both species. Single and dual-species biofilms - SSB and DSB - were exposed to laser doses of 5, 10 or 20 J/cm 2 from a near infrared InGaAsP diode laser prototype (LASERTable; 780 ± 3 nm, 0.04 W). After irradiation, the analysis of biobilm viability (MTT assay), biofilm growth (cfu/mL) and cell morphology (SEM) showed that LLLT reduced cell viability as well as the growth of biofilms. The response of S. mutans (SSB) to irradiation was similar for all laser doses and the biofilm growth was dose dependent. However, when associated with C. albicans (DSB), S. mutans was resistant to LLLT. For C. albicans, the association with S. mutans (DSB) caused a significant decrease in biofilm growth in a dose-dependent fashion. The morphology of the microorganisms in the SSB was not altered by LLLT, while the association of microbial species (DSB) promoted a reduction in the formation of C. albicans hyphae. LLLT had an inhibitory effect on the microorganisms, and this capacity can be altered according to the interactions between different microbial species.

Effect of brushing with conventional versus whitening dentifrices on surface roughness and biofilm formation of dental ceramics

The aim of this study was to evaluate the effect of conventional and whitening dentifrices on the weight loss, surface roughness, and early in situ biofilm formation on the surface of dental ceramics. Standardized feldspar ceramic specimens (Vita VM7 and Vita VM13) were submitted to the following experimental conditions: no brushing; brushing without a dentifrice; brushing with a conventional dentifrice; and brushing with a whitening dentifrice. A brushing machine was used to simulate brushing. The mass and surface roughness of all specimens from the test groups were evaluated prior to and after brushing. Ten participants used an oral device for eight hours to evaluate the biofilm formed in situ on the specimens. Scanning electron microscopy was used for qualitative and quantitative analysis of the biofilm. ANOVA and Tukey tests were used to analyze the results of weight loss, surface roughness, and presence of bacteria. A one-way Kruskal-Wallis test was used for bacterial colonization results. For both ceramics, brushing with a whitening dentifrice resulted in weight loss that was significantly greater when compared to brushing without a dentifrice or with a conventional dentifrice. Increased surface roughness was noticed on VM13 ceramic samples with both dentifrices, whereas only conventional dentifrice had a significant effect on the surface roughness of VM7 samples. For both VM7 and VM13, no difference was found between the experimental conditions with regard to the presence or number of bacteria. Cocci and short rods were the predominant microbial morphotypes. Granular or fibrillar acellular material partially covered the specimens. Brushing with a whitening dentifrice resulted in significant weight loss of ceramic restorations, while brushing with both conventional and whitening dentifrices can roughen ceramic surfaces. The increase in roughness was not clinically significant to contribute to increased biofilm formation.

Daily biofilm control and oral health: Consensus on the epidemiological challenge-Latin American Advisory Panel

Our understanding of dental plaque biofilm has evolved since the nonspecific plaque hypothesis that considered plaque as a nonspecific mass of native microorganisms that, because of lack of oral hygiene, builds up in proportions great enough to overcome the host resistance threshold and affect the tooth structure and tooth supporting tissues. A great diversity of microorganisms-over 700 species-was detected in the oral cavity, and evidence shows that the investigation of specific microorganisms or associations of microorganisms as etiological agents for periodontal diseases and caries is not a simplistic approach. Although clinical evidence shows that oral mechanical hygiene is fundamental to prevent and control caries and periodontal disease, it is important to highlight that optimal control is not achieved by most individuals. Thus the complementary use of chemotherapeutic agents has been investigated as a way to overcome the deficiencies of mechanical oral hygiene habits, insofar as they reduce both plaque formation and gingival inflammation, and represent a valid strategy to change the biofilm and maintain dental and periodontal health. The role of the dental professional is to monitor patients and offer them the best recommendations to preserve oral health throughout their life. With this in mind, chemical control should be indicated as part of daily oral hygiene, together with mechanical procedures, for all individuals who present supragingival and/or subgingival biofilm, taking into account age, physical and/or psychological limitations, allergies, and other factors.

Candida species: Current epidemiology, pathogenicity, biofilm formation, natural antifungal products and new therapeutic options

The incidence of fungal infections has increased significantly, so contributing to morbidity and mortality. This is caused by an increase in antimicrobial resistance and the restricted number of antifungal drugs, which retain many side effects. Candida species are major human fungal pathogens that cause both mucosal and deep tissue infections. Recent evidence suggests that the majority of infections produced by this pathogen are associated with biofilm growth. Biofilms are biological communities with a high degree of organization, in which micro-organisms form structured, coordinated and functional communities. These biological communities are embedded in a self-created extracellular matrix. Biofilm production is also associated with a high level of antimicrobial resistance of the associated organisms. The ability of Candida species to form drugresistant biofilms is an important factor in their contribution to human disease. The study of plants as an alternative to other forms of drug discovery has attracted great attention because, according to the World Health Organization, these would be the best sources for obtaining a wide variety of drugs and could benefit a large population. Furthermore, silver nanoparticles, antibodies and photodynamic inactivation have also been used with good results. This article presents a brief review of the literature regarding the epidemiology of Candida species, as well as their pathogenicity and ability to form biofilms, the antifungal activity of natural products and other therapeutic options. © 2013 SGM.

Response of molars and non-molars to a strict supragingival control in periodontal patients

The posterior position in the arches is one of the factors that underlies the poor prognosis of molar teeth (M). It is speculated that M do not benefit from the oral hygiene routine as well as non-molars (NM) do. This study evaluated the response of M and NM to supragingival control during a 6-month period in 25 smokers (S) and 25 never-smokers (NS) with moderate-to-severe periodontitis. One calibrated examiner assessed visible plaque (VPI) and gingival bleeding (GBI) indexes, periodontal probing depth (PPD), bleeding on probing (BOP), and clinical attachment loss (CAL) at days 0 (baseline), 30 and 180. At baseline, M showed significantly higher mean values of VPI (p = 0.017) and PPD (p < 0.001) compared with NM; CAL was also greater in M (p < 0.001) and was affected by smoking (p = 0.007). The reductions obtained for periodontal indicators at day 180 showed similar responses between M and NM. For CAL, M (NS 0.57 ± 0.50; S 0.67 ± 0.64) and NM (NS 0.38 ± 0.23; S 0.50 ± 0.33) reached an almost significant difference (p = 0.05). Smoking did not influence the response to treatment. Multilevel analysis revealed that, only for PDD reductions, the interaction between sites, teeth and patient was significant (p < 0.001). It was concluded that M benefit from an adequate regimen of supragingival biofilm control; therefore, supragingival condition should be considered in the prognosis of molar teeth.

Recovery of mutans streptococci on MSB, SB-20 and SB-20M agar media

Objective: The recovery of mutans streptococci in saliva and dental biofilm samples depends, in part, on the culture medium used. In this study, we compared (i) the culture media Sucrose-Bacitracin agar (SB-20), Modified SB-20 (SB-20M) and Mitis Salivarius Bacitracin agar (MSB) in the count of colony forming units (cfu) of mutans streptococci and (ii) in the morphological and biochemical differentiation between Streptococcus mutans and Streptococcus sobrinus. Design: Samples of non-stimulated saliva from 20 children were plated on SB-20, SB-20M and MSB, and incubated in microaerophilia at 37 °C for 72 h. Identification of microorganisms was based on analysis of colony morphology under stereomicroscopy. The biochemical identification of colonies was done by biochemical tests using sugar fermentation, resistance to bacitracin and hydrogen peroxide production. Results: There was no significant difference (p > 0.05) in the number of cfu of mutans streptococci recovered on SB-20 and SB-20M agar. Comparing the media, SB-20 and SB-20M yielded a larger number of mutans streptococci colonies (p < 0.05) and were more effective than MSB in the identification of S. sobrinus (p < 0.05), but not of S. mutans (p > 0.05). Conclusion: There was no significant difference between SB-20 and SB-20M culture media in the count of mutans streptococci, demonstrating that the replacement of sucrose by coarse granular cane sugar did not alter the efficacy of the medium. Compared with MSB, SB-20 and SB-20M allowed counting a larger number of mutans streptococci colonies and a more effective morphological identification of S. sobrinus. © 2012 Elsevier Ltd.

N-acetylcysteine and vancomycin alone and in combination against staphylococci biofilm

Introduction: The ability of staphylococci to produce biofilm is an important virulence mechanism that allows bacteria both to adhere and to live on artificial surfaces and to resist to the host immune factors and antibiotics. Staphylococcal infections have become increasingly difficult to treat due their antibiotic resistance. Therefore, there is a continuous need for new and effective treatment alternatives against staphylococcal infections. The main goal of this study was to test N-acetylcysteine (NAC) and vancomycin alone and in combination against S. epidermidis and S. aureus biofilms. Methods: Biofilms were treated with NAC at minimum inhibitory concentration (MIC) and 10 × MIC concentrations and vancomycin at MIC and peak serum concentrations. Results: The use of NAC 10 × MIC alone showed a significant antibactericidal effect, promoting a 4-5 log10 CFU/ mL reduction in biofilm cells. The combination of NAC 10 × MIC with vancomycin (independently of the concentration used) reduced significantly the number of biofilm cells for all strains evaluated (5-6 log10). Conclusion: N-acetylcysteine associated to vancomycin can be a potential therapeutic strategy in the treatment of infections associated to biofilms of S. epidermidis or S. aureus.

Bifunctional silica nanoparticles for the exploration of biofilms of Pseudomonas aeruginosa

Luminescent silica nanoparticles are frequently employed for biotechnology applications mainly because of their easy functionalization, photo-stability, and biocompatibility. Bifunctional silica nanoparticles (BSNPs) are described here as new efficient tools for investigating complex biological systems such as biofilms. Photoluminescence is brought about by the incorporation of a silylated ruthenium(II) complex. The surface properties of the silica particles were designed by reaction with amino-organosilanes, quaternary ammonium-organosilanes, carboxylate-organosilanes and hexamethyldisilazane. BSNPs were characterized extensively by DRIFT, 13C and 29Si solid state NMR, XPS, and photoluminescence. Zeta potential and contact angle measurements exhibited various surface properties (hydrophilic/hydrophobic balance and electric charge) according to the functional groups. Confocal laser scanning microscopy (CLSM) measurements showed that the spatial distribution of these nanoparticles inside a biofilm of Pseudomonas aeruginosa PAO1 depends more on their hydrophilic/hydrophobic characteristics than on their size. CLSM observations using two nanosized particles (25 and 68 nm) suggest that narrow diffusion paths exist through the extracellular polymeric substances matrix. © 2013 Copyright Taylor and Francis Group, LLC.

Antibiofilm activity, pH and solubility of endodontic sealers

Aim: To evaluate antibiofilm activity against Enterococcus faecalis, pH and solubility of AH Plus, Sealer 26, Epiphany SE, Sealapex, Activ GP, MTA Fillapex (MTA-F) and an experimental MTA-based Sealer (MTA-S). Methodology: Sealer samples were manipulated and stored for 2 or 7 days. Prepared sealers were evaluated by a modified direct contact test (DCT) for 5 h, 10 h or 15 h with biofilm previously induced on bovine dentine for 14 days. In the control group, the biofilm was not exposed to the sealers. The number of colony-forming units (CFU mL-1) in the remaining biofilm was determined. Sealer solubility was assessed by the percentage of mass loss after 15 h of immersion in distilled water. Sealer pH was measured at 5 h, 10 h and 15 h. Statistical analysis was performed using Kruskal-Wallis and Dunn or anova and Tamhane's T2 tests, at 5% significance. Results: At 2 days post-manipulation, the DCT showed that Sealapex and MTA-F were associated with a reduction in the number of bacteria in all 3 contact periods evaluated, compared with the control group (P < 0.05). At 7 days, Sealapex had the greatest antibiofilm action at 10 h and 15 h. Sealapex had the highest pH values 2 and 7 days post-manipulation. Regarding the solubility, at 2 days the highest values were observed for MTA-F, MTA-S, Sealapex and Activ GP (P < 0.05). At 7 days, MTA-S and MTA-F had greater solubility than the other materials (P < 0.05). AH Plus had the lowest solubility for both post-manipulation periods (P < 0.05). Conclusion: Sealapex and MTA-F were associated with a reduction in the number of bacteria in biofilms and had greater solubility. The high solubility and pH may be related to the antibacterial activity of these materials. © 2012 International Endodontic Journal.

Production of biofilm by Listeria monocytogenes in different materials and temperatures

Listeria monocytogenes, considered as one of the most important foodborne pathogens, is easily found on surfaces, particularly in the form of a biofilm. Biofilms are aggregates of cells that facilitate the persistence of these pathogens in food processing environments conferring resistance to the processes of cleaning and may cause contamination of food during processing, thus, representing a danger to public health. Little is known about the dynamics of the formation and regulation of biofilm production in L.monocytogenes, but several authors reported that the luxS gene may be a precursor in this process. In addition, the product of the inlA gene is responsible for facilitating the entry of the microorganism into epithelial cells that express the receptor E-cadherin, also participates in surface attachment. Thus, 32 strains of L.monocytogenes isolated from different foods (milk and vegetables) and from food processing environments were analyzed for the presence of these genes and their ability to form biofilms on three different surfaces often used in the food industry and retail (polystyrene, glass and stainless steel) at different temperatures (4, 20 and 30°C). All strains had the ilnA gene and 25 out of 32 strains (78.1%) were positive for the presence of the luxS gene, but all strains produced biofilm in at least one of the temperatures and materials tested. This suggests that genes in addition to luxS may participate in this process, but were not the decisive factors for biofilm formation. The bacteria adhered better to hydrophilic surfaces (stainless steel and glass) than to hydrophobic ones (polystyrene), since at 20°C for 24h, 30 (93.8%) and 26 (81.3%) produced biofilm in stainless steel and glass, respectively, and just 2 (6.2%) in polystyrene. The incubation time seemed to be an important factor in the process of biofilm formation, mainly at 35°C for 48h, because the results showed a decrease from 30 (93.8%) to 20 (62.5%) and from 27 (84.4%) to 12 (37.5%), on stainless steel and glass, respectively, although this was not significant (. p=0.3847). We conclude that L.monocytogenes is capable of forming biofilm on different surfaces independent of temperature, but the surface composition may be important factor for a faster development of biofilm. © 2013 Elsevier Ltd.

NaOCl effect on biofilm produced by Staphylococcus aureus isolated from the milking environment and mastitis infected cows

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Highlights in pathogenic fungal biofilms

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Adhesion of Candida biofilm cells to human epithelial cells and polystyrene after treatment with silver nanoparticles

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Fibrinogen-Induced Streptococcus mutans Biofilm Formation and Adherence to Endothelial Cells

Streptococcus mutans, the predominant bacterial species associated with dental caries, can enter the bloodstream and cause infective endocarditis. The aim of this study was to investigate S. mutans biofilm formation and adherence to endothelial cells induced by human fibrinogen. The putative mechanism by which biofilm formation is induced as well as the impact of fibrinogen on S. mutans resistance to penicillin was also evaluated. Bovine plasma dose dependently induced biofilm formation by S. mutans. Of the various plasma proteins tested, only fibrinogen promoted the formation of biofilm in a dose-dependent manner. Scanning electron microscopy observations revealed the presence of complex aggregates of bacterial cells firmly attached to the polystyrene support. S. mutans in biofilms induced by the presence of fibrinogen was markedly resistant to the bactericidal effect of penicillin. Fibrinogen also significantly increased the adherence of S. mutans to endothelial cells. Neither S. mutans cells nor culture supernatants converted fibrinogen into fibrin. However, fibrinogen is specifically bound to the cell surface of S. mutans and may act as a bridging molecule to mediate biofilm formation. In conclusion, our study identified a new mechanism promoting S. mutans biofilm formation and adherence to endothelial cells which may contribute to infective endocarditis.

Ability of Salmonella spp. to Produce Biofilm Is Dependent on Temperature and Surface Material

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)