Effect of ozone on periodontopathogenic species—an in vitro study

The in vitro study was aimed to determine the effect of ozone on periodontopathogenic microorganisms. Ozone was generated for 6s-2 × 24s (corresponding to 0.56mg-2 × 2.24mg of ozone) against 23 mainly anaerobic periodontopathogenic species. Agar diffusion test was used as a screening method. Then, the killing activity was tested in a serum-free environment and with 25% v/v inactivated serum. Further, the effect of ozone on bactericidal activity of native serum was analyzed against Fusobacterium nucleatum, Porphyromonas gingivalis, and Aggregatibacter actinomycetemcomitans. Agar diffusion test showed a high efficacy of ozone against microorganisms, especially against Porphyromonas gingivalis. This result was confirmed by the killing tests; most of the strains in a concentration of 105 were completely eliminated after twofold 18-s application of ozone. Only four of the six potentially "superinfecting” species (Staphylococcus aureus, Enterococcus faecalis, Enterobacter cloacae, Candida albicans) survived in part. Addition of heat-inactivated serum reduced the killing rate of ozone by 78% after 6-s and by 47% after twofold 18-s exposures; no strain was completely eradicated after any application of ozone. The bactericidal effect of native serum was enhanced after application of ozone; no effect was visible on the included A. actinomycetemcomitans strain which was found to be completely resistant to the bactericidal action of serum. In conclusion, (a) ozone has a strong antibacterial activity against putative periodontopathogenic microorganisms, and (b) the bactericidal effect is reduced in the presence of serum. Ozone may have potential as an adjunctive application to mechanical treatment in periodontitis patient

Oral microbiota in Swiss adolescents

Objectives: The purpose of the study was to determine the prevalence of different oral microbes in gingival plaque samples and in samples from the dorsum of the tongue in a Swiss adolescent population. Materials and methods: Ninety-nine adolescents between 15 and 18years were enrolled. Plaque index, bleeding on probing (BOP), the periodontal screening index, and decayed missed filled tooth (DMFT) index were recorded. Samples from subgingival plaque and swabs from the tongue were analyzed by the Checkerboard DNA-DNA hybridization method. Additionally, counts of Streptococus mutans and Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola were determined by real-time PCR. Results: Periodontitis was not diagnosed in any of the subjects but all of them presented signs of gingival inflammation displaying a mean BOP of 28%. Ten (10.1%) subjects were tested positive for P. gingivalis, each 22 (22.2%) for A. actinomycetemcomitans and T. forsythia, (47.5%) for T. denticola. T. denticola and S. mutans showed a high affinity to the gingival plaque, whereas T. forsythia was often detected from the dorsum of the tongue. DMFT was associated with S. mutans counts, and BOP correlated with counts of P. gingivalis and T. denticola. Conclusions: The present data indicate that: (a) gingivitis but not periodontitis is a common finding among Swiss adolescents, and (b) bacteria associated with periodontitis were frequently detected in the subgingival dental plaque and on the dorsum of the tongue in Swiss adolescents with gingivitis. Clinical relevance: Although gingivitis was a frequent finding in Swiss adolescents, periodontitis was not detected in this population. The dorsum of the tongue appears to represent an important reservoir for periodontopathic bacteri

In-vitro effects of different hyaluronic acids on periodontal biofilm-immune cell interaction.

INTRODUCTION Recent studies have demonstrated a positive role of hyaluronic acid (HA) on periodontal clinical outcomes. This in-vitro study aimed to investigate the impact of four different HAs on interactions between periodontal biofilm and immune cells. METHODS The four HAs included: high-molecular-weight HA (HHA, non-cross-linked), low-molecular-weight HA (LHA), oligomers HA (OHA), and cross-linked high-molecular-weight HA (CHA). Serial experiments were conducted to verify the influence of HAs on: (i) 12-species periodontal biofilm (formation and pre-existing); (ii) expression of inflammatory cytokines and HA receptors in monocytic (MONO-MAC-6) cells and periodontal ligament fibroblasts (PDLF) with or without exposure to periodontal biofilms; (iii) generation of reactive oxygen species (ROS) in MONO-MAC-6 cells and PDLF with presence of biofilm and HA. RESULTS The results indicated that HHA and CHA reduced the bacterial counts in a newly formed (4-h) biofilm and in a pre-existing five-day-old biofilm. Without biofilm challenge, OHA triggered inflammatory reaction by increasing IL-1β and IL-10 levels in MONO-MAC cells and IL-8 in PDLF in a time-dependent manner, whereas CHA suppressed this response by inhibiting the expression of IL-10 in MONO-MAC cells and IL-8 in PDLF. Under biofilm challenge, HA decreased the expression of IL-1β (most decreasing HHA) and increased IL-10 levels in MONO-MAC-6 cells in a molecular weight dependent manner (most increasing CHA). The interaction between HA and both cells may occur via ICAM-1 receptor. Biofilm stimulus increased ROS levels in MONO-MAC-6 cells and PDLF, but only HHA slightly suppressed the high generation of ROS induced by biofilm stimulation in both cells. CONCLUSION Overall, these results indicate that OHA induces inflammation, while HHA and CHA exhibit anti-biofilm, primarily anti-inflammatory, and antioxidant properties in the periodontal environment

Antimicrobial Efficiency of Pistacia lentiscus L. Derivates against Oral Biofilm-Associated Diseases-A Narrative Review.

Pistacia lentiscus L. (PlL) has been used for centuries in traditional medicine. The richness in antimicrobial biomolecules of Pll derivates can represent an alternative to chemically formulated agents used against oral infections. This review summarizes the knowledge on the antimicrobial activity of PlL essential oil (EO), extracts, and mastic resin against microorganisms being of relevance in oral biofilm-associated diseases. Results demonstrated that the potential of PlL polyphenol extracts has led to increasing scientific interest. In fact, the extracts are a significantly more effective agent than the other PlL derivates. The positive findings regarding the inhibition of periodontal pathogens and C. albicans, together with the antioxidant activity and the reduction of the inflammatory responses, suggest the use of the extracts in the prevention and/or reversal of intraoral dysbiosis. Toothpaste, mouthwashes, and local delivery devices could be effective in the clinical management of these oral diseases

In- vitro-activity of additive application of hydrogen peroxide in antimicrobial photodynamic therapy using LED in the blue spectrum against bacteria and biofilm associated with periodontal disease.

BACKGROUND Although antimicrobial photodynamic therapy (aPDT) has been shown to be efficient in killing planktonic periodontopathogenic bacteria, its activity on established biofilms is very limited. The aim of the present in-vitro study was to evaluate the potential effect of hydrogen peroxide as a pretreatment for aPDT. METHODS aPDT consisting of riboflavin as photosensitizer and illumination by a LED lamp emitting in the blue spectrum for 30 s and 60 s (aPDT60) was combined with a pretreatment with 0.25% and 3% hydrogen peroxide. The antimicrobial activity of these treatments was determined against eight oral species (incl. Porphyromonas gingivalis and Tannerella forsythia) and against eight-species biofilms. Treatment of biofilms in an artificial pocket model included a mechanical removal of the biofilm. RESULTS Against planktonic bacteria, pretreatment with hydrogen peroxide increased killing of planktonic bacteria, after aPDT60 no viable bacteria were detected in 7 of 8 strains. In biofilms formed on well-plates, aPDT60 reduced bacterial counts only by 0.53 log10 cfu, whereas reduction was closed to 4 log10 or higher when 3% hydrogen peroxide was used. When biofilms were treated in the periodontal-pocket model, reduction of cfu was less than 0.5 log10 after mechanical therapy or aPDT60 only, however no bacteria were detected after mechanical biofilm removal followed by the use 3% of hydrogen peroxide and aPDT60. CONCLUSIONS aPDT using riboflavin and blue LED light applied after mechanical removal of biofilm and adjunctive 3% hydrogen peroxide solution appears to represent an alternative for antimicrobial periodontal therapy

Non-Surgical Periodontal Therapy with Adjunctive Amoxicillin/Metronidazole or Metronidazole When No Aggregatibacter actinomycetemcomitans Is Detected-A Randomized Clinical Trial.

BACKGROUND The aim was to compare two different systemic antibiotics regimens adjunctive to non-surgical periodontal therapy when Aggregatibacter actinomycetemcomitans was not detected in the subgingival biofilm. METHODS A total of 58 patients with periodontitis and with no A. actinomycetemcomitans in the subgingival biofilm were treated with full-mouth subgingival instrumentation and either metronidazole (MET; n = 29) or amoxicillin/metronidazole (AMX/MET; n = 29). Probing depth (PD), clinical attachment level (CAL) and bleeding on probing (BOP) were recorded at baseline, as well as after three and six months. Subgingival biofilm and gingival crevicular fluid were collected and analyzed for major periodontopathogens and biomarkers. RESULTS PD, CAL and BOP improved at 3 and 6 months (each p < 0.001 vs. baseline) with no difference between the groups. Sites with initial PD ≥ 6 mm also improved in both groups after 3 and 6 months (p < 0.001) with a higher reduction of PD in the AMX/MET group (p < 0.05). T. forsythia was lower in the AMX/MET group after 3 months (p < 0.05). MMP-8 and IL-1β were without significant changes and differences between the groups. CONCLUSION When A. actinomycetemcomitans was not detected in the subgingival biofilm, the adjunctive systemic use of amoxicillin/metronidazole results in better clinical and microbiological outcomes of non-surgical periodontal therapy when the application of systemic antibiotics is scheduled

A novel in vitro periodontal pocket model to evaluate the effect of root surface instrumentation on biofilm-epithelial cell interactions.

OBJECTIVE To develop a novel in vitro periodontal pocket model for evaluating the effect of two different root surface instrumentation modalities on biofilm-epithelial cell interactions. MATERIALS AND METHODS An artificial periodontal pocket model was created using an impression material. Dentin discs were prepared and incubated for 3.5 days with a biofilm consisting of 12 bacterial strains. Then, the discs were inserted into the pocket model and instrumented for 10 s or 10 strokes either with ultrasonics (US) or hand instruments (HI). Subsequently, a glass slide coated with epithelial cells was placed in close vicinity to the discs. After incubation of the pocket model in a 5% CO2 atmosphere for 6 h, residual bacteria of the biofilm as well as bacteria adhering to or invaded into epithelial cells were determined using colony-forming unit (cfu) counts and real-time PCR. Further, as a parameter of the pro-inflammatory cell response, interleukin (IL)-8 expression was determined by ELISA. RESULTS Compared to untreated control, HI reduced the cfu counts by 0.63 log10 (not significant) and US by 1.78 log10 (p = 0.005) with a significant difference between the treatment modalities favoring US (p = 0.048). By trend, lower detection levels of Tannerella forsythia were detected in the US group compared to HI. Concerning the interaction with epithelial cells, half of the control and the HI samples showed epithelial cells with attaching or invading bacteria, while US displayed bacteria only in two out of eight samples. In addition, US resulted in significantly lower IL-8 secretion by epithelial cells compared to the untreated control. Between HI and controls, no statistically significant difference in IL-8 secretion was found. CONCLUSION This newly developed in vitro model revealed in terms of biofilm-epithelial cell interaction after root surface instrumentation that compared to hand curettes, ultrasonic instrumentation appeared to be more effective in removing bacterial biofilm and in decreasing the inflammatory response of epithelium to biofilm. CLINICAL RELEVANCE Ultrasonic instrumentation might be more advantageous to reduce cellular inflammatory response than hand instruments

In vitro evaluation of surface roughness, adhesion of periodontal ligament fibroblasts, and Streptococcus gordonii following root instrumentation with Gracey curettes and subsequent polishing with diamond-coated curettes

Objectives: The objective of the study was to evaluate the efficacy of an additional usage of a diamond-coated curette on surface roughness, adhesion of periodontal ligament (PDL) fibroblasts, and of Streptococcus gordonii in vitro. Materials and methods: Test specimens were prepared from extracted teeth and exposed to instrumentation with conventional Gracey curettes with or without additional use of diamond-coated curettes. Surface roughness (Ra and Rz) was measured before and following treatment. In addition, the adhesion of PDL fibroblasts for 72h and adhesion of S. gordonii ATCC 10558 for 2h have been determined. Results: Instrumentation with conventional Gracey curettes reduced surface roughness (median Ra before: 0.36μm/after: 0.25μm; p < 0.001; median Rz before: 2.34μm/after: 1.61μm; p < 0.001). The subsequent instrumentation with the diamond-coated curettes resulted in a median Ra of 0.31μm/Rz of 2.06μm (no significance in comparison to controls). The number of attached PDL fibroblasts did not change following scaling with Gracey curettes. The additional instrumentation with the diamond-coated curettes resulted in a two-fold increase in the number of attached PDL fibroblasts but not in the numbers of adhered bacteria. Conclusions: Treatment of root surfaces with conventional Gracey curettes followed by subsequent polishing with diamond-coated curettes may result in a root surface which provides favorable conditions for the attachment of PDL fibroblasts without enhancing microbial adhesion. Clinical relevance: The improved attachment of PDL fibroblasts and the limited microbial adhesion on root surfaces treated with scaling with conventional Gracey curettes followed by subsequent polishing with diamond-coated curettes may favor periodontal wound healin

Efficacy of taurolidine against periodontopathic species—an in vitro study

The antimicrobial effect of taurolidine was tested against periodontopathic species in comparison to chlorhexidine digluconate in the presence or absence of serum. Minimal inhibitory concentrations (MIC), microbiocidal concentrations (MBC), as well as killing were determined against 32 different microbial strains including 3 Porphyromonas gingivalis, 3 Aggregatibacter actinomycetemcomitans, and 15 potentially superinfecting species with and without 25% v/v human serum. The MIC50 of taurolidine against the tested microbial strains was 0.025% and the MIC90 0.05%. The respective values for the MBCs were 0.05% and 0.1%. Addition of 25% serum (heat-inactivated) did not change the MIC and MBC values of taurolidine. In contrast, MICs and MBCs of chlorhexidine (CHX) increased by two steps after addition of serum. Taurolidine killed microorganisms in a concentration and time-dependent manner, the killing rate of 1.6% taurolidine was 99.08% ± 2.27% in mean after 2h. Again, killing activity of taurolidine was not affected if serum was added, whereas addition of inactivated serum clearly reduced the killing rate of all selected bacterial strains by CHX. Therefore, taurolidine possesses antimicrobial properties which are not reduced in the presence of serum as a main component in gingival crevicular fluid and wound fluid. Taurolidine may have potential as an antimicrobial agent in non-surgical and surgical periodontal treatmen

In vitro activity of taurolidine gel on bacteria associated with periodontitis.

OBJECTIVES The purpose of this in vitro study was to determine the antimicrobial activity of two different taurolidine gel formulations in comparison with minocycline microspheres. METHODS Three percent taurolidine gel (TLG3) and 2 % taurolidine gel (TLG2) were compared to minocycline microspheres (MINO) against single bacterial species and a 12-species-mixture. The antimicrobial activity was proven by determination of minimal inhibitory concentrations (MICs), killing assays, after exposure of the antimicrobials as well as within a biofilm. RESULTS The MICs against the single species were between 0.5 and 2 mg/ml of taurolidine. MICs of the used mixed microbiota were 1.5 mg/ml (TLG3) and 4 mg/ml (TLG2). Fusobacterium nucleatum and Porphyromonas gingivalis were completely killed by 10 % TLG3 and TLG2 (equivalent to 3 and 2 mg/ml taurolidine) after 6 h. The mixture of 12 species was not completely killed by any of the test substances. Taurolidine gels showed a post-antimicrobial activity, however being less than that of MINO. On biofilms, taurolidine gels reduced concentration dependently the colony forming unit (CFU) counts (multi-species biofilms by 3.63 log10 after 100 % (30 mg/ml) of TLG3), reductions were 2.12 log10 after MINO (1000 μg/ml minocycline). CONCLUSIONS Taurolidine gel formulations exert antimicrobial activity against bacteria associated with periodontal disease. Nevertheless, a complete elimination of biofilms seems to be impossible and underlines the importance of mechanical removal of biofilms prior to application of the antimicrobial. CLINICAL RELEVANCE Taurolidine gels may represent a potential alternative for adjunctive topical antimicrobial treatment in periodontitis and infectious peri-implant diseases