In vitro evaluation of membranes for regenerative procedures against oral bacteria.

The current literature on guided bone regeneration (GBR) and guided tissue regeneration (GTR) membrane contamination reports that the physicochemical characteristics of these biomaterials might influence affinity to bacteria, which appears to be a major drawback for the clinical outcome of the regenerative procedures. Thus, this study aimed to evaluate, in vitro, a multispecies biofilm adherence and passage of bacteria through different types of commercially available membranes for GTR/GBR. Four types of membranes were tested (n=12): LC) Lumina Coat®; JS) Jason®; BG) Biogide®; and LP) Lumina PTFE®. Aluminum foil (AL) simulated an impermeable barrier and was used as the control. The membranes were adapted to specific apparatus and challenged with a mixed bacterial culture composed of A. actinomycetemcomitans b, S. mutans, S. mitis, and A. israelii. After 2 h or 7 days, bacterial adhesion and passage of bacteria were evaluated through CFU counting, which was analyzed by two-way ANOVA e post hoc Tukey, at a 5% significance level. Representative areas of two membranes of each group were analyzed through scanning electron microscopy (SEM) to assess the morphology and organization of the biofilm over the membrane fibers. LC and LP presented similar values of adhered bacterial cells (p > 0.05), significantly inferior when compared to the other groups, in both time points (p < 0.05). All the tested groups were permeable to bacterial cells, with no significant difference between the trial period of 2 h and 7 days (p > 0.05). SEM analyses demonstrated that adhered bacteria number increased throughout the time points (2 h < 7 days). Commercially available biological membranes demonstrated intense bacterial adherence and passage of bacteria, which increased throughout the trial period.

In vitro evaluation of membranes for regenerative procedures against oral bacteria

Abstract The current literature on guided bone regeneration (GBR) and guided tissue regeneration (GTR) membrane contamination reports that the physicochemical characteristics of these biomaterials might influence affinity to bacteria, which appears to be a major drawback for the clinical outcome of the regenerative procedures. Thus, this study aimed to evaluate, in vitro, a multispecies biofilm adherence and passage of bacteria through different types of commercially available membranes for GTR/GBR. Four types of membranes were tested (n=12): LC) Lumina Coat®; JS) Jason®; BG) Biogide®; and LP) Lumina PTFE®. Aluminum foil (AL) simulated an impermeable barrier and was used as the control. The membranes were adapted to specific apparatus and challenged with a mixed bacterial culture composed of A. actinomycetemcomitans b, S. mutans, S. mitis, and A. israelii. After 2 h or 7 days, bacterial adhesion and passage of bacteria were evaluated through CFU counting, which was analyzed by two-way ANOVA e post hoc Tukey, at a 5% significance level. Representative areas of two membranes of each group were analyzed through scanning electron microscopy (SEM) to assess the morphology and organization of the biofilm over the membrane fibers. LC and LP presented similar values of adhered bacterial cells (p > 0.05), significantly inferior when compared to the other groups, in both time points (p < 0.05). All the tested groups were permeable to bacterial cells, with no significant difference between the trial period of 2 h and 7 days (p > 0.05). SEM analyses demonstrated that adhered bacteria number increased throughout the time points (2 h < 7 days). Commercially available biological membranes demonstrated intense bacterial adherence and passage of bacteria, which increased throughout the trial period.

Resumo O objetivo deste estudo foi avaliar, in vitro, a aderência do biofilme multiespécie e a passagem de bactérias através dos diferentes tipos de membranas disponíveis comercialmente para RTG/ROG. Quatro tipos de membranas foram testados (n=12): LC) Lumina Coat®; JS) Jason®; BG) Biogide®; e LP) Lumina PTFE®. Papel alumínio (AL) simulou uma barreira impermeável e foi usado como controle negativo. As membranas foram adaptadas à um aparato específico e desafiadas com uma cultura bacteriana mista composta de A. actinomycetemcomitans b, S. mutans, S. mitis, e A. israelii. Após 2 h ou 7 dias, a aderência e passagem bacteriana foi avaliada através da contagem de UFCs. Duas membranas de cada grupo foram analisadas através da microscopia eletrônica de varredura (MEV). LC e LP apresentaram valores semelhantes de células bacterianas aderidas (p < 0.05), significativamente inferiores quando comparados aos outros grupos, em ambos os períodos experimentais (p < 0.05). Desde a análise inicial, todos os grupos testados foram permeáveis às células bacterianas, sem diferença significativa entre o período experimental de 2 h e 7 dias (p > 0.05). As análises em MEV demonstraram que o número de bactérias aderidas aumentou com o tempo (2 h < 7 days). Membranas biológicas comercialmente disponíveis demonstraram intensa aderência bacteriana e passagem de bactérias, que aumentou durante os períodos experimentais.

Recent Updates on Microbial Biofilms in Periodontitis: An Analysis of In Vitro Biofilm Models.

The development of oral biofilm models has been extremely important to study the specific role of most microbial species at the early stages of periodontitis. The current knowledge on monospecies or multispecies biofilms originates mainly from the observation of in vitro dynamic or static biofilm model systems, which were engineered to mimic clinical oral conditions. In the last few decades, mounting evidence has confirmed that biofilms are the major form of bacterial lifestyle, and more importantly, that microorganisms dwelling in sessile mixed-species aggregates display completely different phenotypes and physiological characteristics than when living in planktonic pure cultures. Interspecies interactions within these communities, mediated by chemical communication systems, have been shown to affect biofilm physiology and increase antimicrobial resistance by up to 1000 fold. These aspects reinforce the importance of developing multispecies biofilm models to better understand and control biofilms. Literature reports demonstrate that while monospecies models are still most commonly used in caries research, authors have used different multispecies models to study periodontal diseases. Periodontitis is a polymicrobial biofilm-dependent disease mainly associated with Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola. Interestingly, these species hardly adhere to substrates commonly used for biofilm formation, which makes multispecies models essential for an accurate analysis of periodontitis-related biofilms. The multispecies models currently available are generally composed of 6-10 species, but a more recent 34-species model was developed to better examine the dynamics within oral biofilms. The complexity of such polymicrobial biofilm models mimics more consistently the oral microbiome and different aspects of the oral environment. Collectively, the evidence on multispecies biofilm models described herein may support future studies on the use of antimicrobials for biofilm control as well as provide research opportunities to expand the current knowledge on interspecies interactions. The present manuscript reviews the most recent updates on in vitro biofilm model systems for periodontitis.

A novel antimicrobial-containing nanocellulose scaffold for regenerative endodontics.

OBJECTIVES: The aim of this study was to evaluate bacterial nanocellulose (BNC) membranes incorporated with antimicrobial agents regarding cytotoxicity in fibroblasts of the periodontal ligament (PDLF), antimicrobial activity, and inhibition of multispecies biofilm formation. MATERIALS AND METHODS: The tested BNC membranes were BNC + 1% clindamycin (BNC/CLI); BNC + 0.12% chlorhexidine (BNC/CHX); BNC + nitric oxide (BNC/NO); and conventional BNC (BNC; control). After PDLF culture, the BNC membranes were positioned in the wells and maintained for 24 hours. Cell viability was then evaluated using the MTS calorimetric test. Antimicrobial activity against Enterococcus faecalis, Actinomyces naeslundii, and Streptococcus sanguinis (S. sanguinis) was evaluated using the agar diffusion test. To assess the antibiofilm activity, BNC membranes were exposed for 24 hours to the mixed culture. After sonicating the BNC membranes to remove the remaining biofilm and plating the suspension on agar, the number of colony-forming units (CFU)/mL was determined. Data were analyzed by 1-way analysis of variance and the Tukey, Kruskal-Wallis, and Dunn tests (α = 5%). RESULTS: PDLF metabolic activity after contact with BNC/CHX, BNC/CLI, and BNC/NO was 35%, 61% and 97%, respectively, compared to BNC. BNC/NO showed biocompatibility similar to that of BNC (p = 0.78). BNC/CLI showed the largest inhibition halos, and was superior to the other BNC membranes against S. sanguinis (p < 0.05). The experimental BNC membranes inhibited biofilm formation, with about a 3-fold log CFU reduction compared to BNC (p < 0.05). CONCLUSIONS: BNC/NO showed excellent biocompatibility and inhibited multispecies biofilm formation, similarly to BNC/CLI and BNC/CHX.

Development of a multispecies periodontal biofilm model within a stirred bioreactor.

The objective of this work was to develop a subgingival biofilm model using a stirred bioreactor. Discs of bovine teeth were adapted to a stirred bioreactor filled with a culture medium containing bacterial species associated with periodontal health or disease. After anaerobic incubation, the biofilms growing on the substratum surfaces were collected and analyzed. The mean number of Colony-forming Units (CFUs) varied, but with no difference between 3 and 7 days of biofilm formation (p > 0.05). Scanning Electron Microscopy (SEM) analysis showed a uniform biofilm layer covering the cement layer of the root surface containing bacteria with diverse morphology. In checkerboard DNA-DNA hybridization, bacterial species were identified in both biofilms. In conclusion, a subgingival biofilm model was developed using a stirred bioreactor, allowing the in vitro reproduction of complex microbial communities. This is an advanced model that may be useful to mimic complex clinical periodontal biofilms.