Alexander Welk

Treatment of Candida albicans biofilms with low-temperature plasma induced by dielectric barrier discharge and atmospheric pressure plasma jet

Because of some disadvantages of chemical disinfection in dental practice (especially denture cleaning), we investigated the effects of physical methods on Candida albicans biofilms. For this purpose, the antifungal efficacy of three different low-temperature plasma devices (an atmospheric pressure plasma jet and two different dielectric barrier discharges (DBDs)) on Candida albicans biofilms grown on titanium discs in vitro was investigated. As positive treatment controls, we used 0.1% chlorhexidine digluconate (CHX) and 0.6% sodium hypochlorite (NaOCl). The corresponding gas streams without plasma ignition served as negative treatment controls. The efficacy of the plasma treatment was determined evaluating the number of colony-forming units (CFU) recovered from titanium discs. The plasma treatment reduced the CFU significantly compared to chemical disinfectants. While 10 min CHX or NaOCl exposure led to a CFU log10 reduction factor of 1.5, the log10 reduction factor of DBD plasma was up to 5. In conclusion, the use of low-temperature plasma is a promising physical alternative to chemical antiseptics for dental practice.

Antimicrobial efficacy of non‐thermal plasma in comparison to chlorhexidine against dental biofilms on titanium discs in vitro – proof of principle experiment

AIM Dental biofilms play a major role in the pathogenesis of peri-implant mucositis. Biofilm reduction is a pre-requisite for a successful therapy of peri-implant mucosal lesions. In this study, we evaluated the effect of three different plasma devices on the reduction of Streptococcus mutans (S. mutans) and multispecies human saliva biofilms. MATERIAL AND METHODS We assessed the efficacy of three different non-thermal atmospheric pressure plasma devices against biofilms of S. mutans and saliva multispecies grown on titanium discs in vitro in comparison with a chlorhexidine digluconate (CHX) rinse. Efficacy of plasma treatment was determined by the number of colony forming units (CFU) and by scanning electron microscopy. The results were reported as reduction of CFU (CFU(untreated) -CFU(treated) ). RESULTS The application of plasma was much more effective than CHX against biofilms. The maximum reduction of CHX was 3.36 for S. mutans biofilm and 1.50 for saliva biofilm, whereas the colony forming units (CFU) reduction of the volume dielectric barrier discharge argon plasma was 5.38 for S. mutans biofilm and 5.67 for saliva biofilm. CONCLUSIONS Treatment of single- and multispecies dental biofilms on titanium discs with non-thermal atmospheric pressure plasma was more efficient than CHX application in vitro. Thus, the development of plasma devices for the treatment of peri-implant mucositis may be fruitful.

The Peritoneal Explant Test for Evaluating Tissue Tolerance to Mouthrinses

The tissue cultures of explants of neonatal rat peritoneum have been demonstrated to be a sensitive test for tissue compatibility with wound antiseptics. The present study investigated the suitability of this method to assess the relative toxicity of mouthrinses to tissue. Mouthrinses containing 0.1% chlorhexidine (Chlorhexamed® Fluid 0.1%) (A), 0.3% triclosan (Colgate®) (B), essential oil in ethanolic solution (Listerine®) (C), and amine/stannous fluoride (Meridol®) (D) were tested at use concentration and in dilutions of 10, 1, and 0.1% with exposure times of 1, 10, and 30 min, respectively. The mouthrinses (test) and Ringer’s solutions (control) were applied to opened rat peritoneum. After thorough irrigation with Ringer’s solution, a piece of peritoneum was removed and 1 × 1 mm explants were cut. The explants were cultivated with a bovine serum culture medium in 24-well plates at 37°C in a CO2 incubator (95% air, 5% CO2). After 10 days, the tissue proliferation for the explants was assessed by a stereo microscope at 10× magnification after ethanol fixing and hemalaun staining. With 24 grafts per test, the proliferation rate was calculated relative to a control, which was run for each mouthrinse and concentration/time combination. Data were analyzed using ANOVA (SPSS 11.0) and post-hoc paired t test. Statistical significance of all correlations was tested by setting the significance level at p < 0.05. At most concentrations, D caused significantly less tissue damage than A or B. There was no difference between C and A or C and B at 100%. However, the toxicity of C was significantly less than A or B at 10, 1, and 0.1%. C and D behaved similarly except for the 10% (30 min) and the 1% (10 min) solutions in which C was significantly less toxic. We concluded that the rat peritoneum explant test was demonstrated to be a sensitive test to assess the relative toxicity of mouthrinses to tissue.

In vitro efficacy of cold atmospheric pressure plasma on S. sanguinis biofilms in comparison of two test models

Dental plaque critically affects the etiology of caries, periodontitis and periimplantitis. The mechanical removal of plaque can only be performed partially due to limited accessibility. Therefore, plaque still represents one of the major therapeutic challenges. Even though antiseptic mouth rinses reduce the extent of biofilm temporarily, plaque removal remains incomplete and continuous usage can even result in side effects. Here we tested argon plasma produced by kinpen09 as one option to inactivate microorganisms and to eliminate plaque. S. sanguinis biofilms cultivated in either the European Biofilm Reactor (EUREBI) or in 24 well plates were treated with argon plasma. In both test systems a homogeneous, good analyzable and stable biofilm was produced on the surface of titan plates within 72 h (>6,9 log10 CFU/ml). Despite the significantly more powerful biofilm production in EUREBI, the difference of 0.4 log10 CFU/ml between EUREBI and the 24 well plates was practically not relevant. For that reason both test models were equally qualified for the analysis of efficacy of cold atmospheric pressure plasma. We demonstrate a significant reduction of the biofilm compared to the control in both test models. After plasma application of 180 s the biofilm produced in EUREBI or in 24 well plates was decreased by 0.6 log10 CFU/ml or 0.5 log10 CFU/ml, respectively. In comparison to recently published studies analyzing the efficacy of kinpen09, S. sanguinis produces a hardly removable biofilm. Future investigations using reduced distances between plasma source and biofilm, various compositions of plasma and alternative plasma sources will contribute to further optimization of the efficacy against S. sanguinis biofilms.

Antimicrobial efficacy of amine fluoride based tooth gels compared to a toothpaste in a phase 2/step 2 in-vitro test model.

Introduction: The aim of the present study was to determine the antimicrobial effect of various gel formulations on plaque formation; different tooth gels were compared to a toothpaste containing comparable antimicrobial ingredients with regard to its microbiocidal activity. The study was conducted under the assumption, that a chief requirement for the prevention of plaque formation is the combination of mechanical removal and antimicrobial activity, and not the sole capability of mechanical plaque removal. Methods: Ledermix® fluoride gel as commercially available with preservative, and without preservative and perfume oils, Elmex® gelée, and Meridol® toothpaste were tested in a standardized in-vitro test modification of the quantitative suspension test EN 1040. Instead of testing in a suspension, the respective product was directly placed on a standardized contaminated sterile stainless steel disk without adding any bio-burden. 50% egg yolk in Aqua dest. was used as a neutralizer. Results: Within 1 min, Elmex® gelée showed a RF >5 log10 against S. pyogenes and S. sanguinis. Against S. mutans, a log10 RF of ≥5 was achieved after 2 min, against C. albicans after 5 min, and against P. aeruginosa after 10 min S. aureus was the most difficult organisms to be reduced. After an application time of 10 min, only a log10 RF of 2.4 was achieved. Ledermix exceeded the antimicrobial efficacy of Elmex® gelée against S. mutans and C. albicans and was already effective against these organisms after 1 min, but did not show the same antimicrobial efficacy as Elmex® gelée against P. aeruginosa. Similar to Elmex® gelée, a required reduction of >5 log10 for antimicrobials under no organic challenge was not achieved against S. aureus. Ledermix® fluoride gel without preservatives and Ledermix® fluoride gel without preservatives and perfume oil did not show the antimicrobial efficacy of the standard Ledermix® fluoride gel formulation, indicating that the observed antimicrobial efficacy is chiefly based on the preservative, and possibly the perfume oil. Compared to the tested gels, Meridol® toothpaste was less effective and reached any antimicrobial effect >5 log10 only against S. sanguinis after 10 min. Conclusion: All unmodified tested gels showed an antimicrobial effect. Because no relevant antimicrobial efficacy against plaque forming bacteria was achieved within 2 min, in practice, an anti-plaque forming effect based on the antimicrobial action of gels cannot be assumed when used in the oral cavity. However, the results of the present study indicate that the antimicrobial efficacy of gels is determined by their formulation and that for the prevention of plaque formation the combination of mechanical removal and antimicrobial activity is not the chief requirement only, but a sustained antimicrobial effect may be of greater importance.