Pilot-study on the influence of carrier gas and plasma application (open resp. delimited) modifications on physical plasma and its antimicrobial effect against Pseudomonas aeruginosa and Staphylococcus aureus

Introduction: Physical plasma is a promising new technology regarding its antimicrobial effects. This is especially accounting for treatment of bacterial infection of chronic wounds. Plasma can be generated with different carrier gases causing various biological effects. Screening of different carrier gases and plasma generation setups is therefore needed to find suitable compositions for highly effective antimicrobial plasma treatments and other applications

XTT assay of ex vivo saliva biofilms to test antimicrobial influences

Objective: Many dental diseases are attributable to biofilms. The screening of antimicrobial substances, in particular, requires a high sample throughput and a realistic model, the evaluation must be as quick and as simple as possible. For this purpose, a colorimetric assay of the tetrazolium salt XTT (sodium 3'-[1-[(phenylamino)-carbony]-3,4-tetrazolium]-bis(4-methoxy-6-nitro)benzene-sulfonic acid hydrate) converted by saliva biofilms is recommended. Cleavage of XTT by dehydrogenase enzymes of metabolically active cells in biofilms yields a highly colored formazan product which is measured photometrically

XTT assay of ex vivo saliva biofilms to test antimicrobial influences

Objective: Many dental diseases are attributable to biofilms. The screening of antimicrobial substances, in particular, requires a high sample throughput and a realistic model, the evaluation must be as quick and as simple as possible. For this purpose, a colorimetric assay of the tetrazolium salt XTT (sodium 3'-[1-[(phenylamino)-carbony]-3,4-tetrazolium]-bis(4-methoxy-6-nitro)benzene-sulfonic acid hydrate) converted by saliva biofilms is recommended. Cleavage of XTT by dehydrogenase enzymes of metabolically active cells in biofilms yields a highly colored formazan product which is measured photometrically

Efficiency of cold atmospheric plasma, cleaning powders and their combination for biofilm removal on two different titanium implant surfaces

Objectives: Biofilm removal is the decisive factor for the control of peri-implantitis. Cold atmospheric pressure plasma (CAP) can become an effective aid due to its ability to destroy and to inactivate bacterial biofilm residues. This study evaluated the cleaning efficiency of CAP, and air-polishing with glycine (APG) or erythritol (APE) containing powders alone or in combination with CAP (APG + CAP, APE + CAP) on sandblasted/acid etched, and anodised titanium implant surface. Materials and methods: On respective titanium discs, a 7-day ex vivo human biofilm was grown. Afterwards, the samples were treated with CAP, APG, APE, APG + CAP, and APE + CAP. Sterile and untreated biofilm discs were used for verification. Directly after treatment and after 5 days of incubation in medium at 37 °C, samples were prepared for examination by fluorescence microscopy. The relative biofilm fluorescence was measured for quantitative analyses. Results: Air-polishing with or without CAP removed biofilms effectively. The combination of air-polishing with CAP showed the best cleaning results compared to single treatments, even on day 5. Immediately after treatment, APE + CAP showed insignificant higher cleansing efficiency than APG + CAP. Conclusions: CAP supports mechanical cleansing and disinfection to remove and inactivate microbial biofilm on implant surfaces significantly. Here, the type of the powder was not important. The highest cleansing results were obtained on sandblasted/etched surfaces. Clinical relevance. Microbial residuals impede wound healing and re-osseointegration after peri-implantitis treatment. Air-polishing treatment removes biofilms very effectively, but not completely. In combination with CAP, microbial free surfaces can be achieved. The tested treatment regime offers an advantage during treatment of peri-implantitis

A three-phase in-vitro system for studying Pseudomonas aeruginosa adhesion and biofilm formation upon hydrogel contact lenses

<p>Abstract</p> <p>Background</p> <p><it>Pseudomonas aeruginosa </it>is commonly associated with contact lens (CL) -related eye infections, for which bacterial adhesion and biofilm formation upon hydrogel CLs is a specific risk factor. Whilst <it>P. aeruginosa </it>has been widely used as a model organism for initial biofilm formation on CLs, <it>in-vitro </it>models that closely reproduce <it>in-vivo </it>conditions have rarely been presented.</p> <p>Results</p> <p>In the current investigation, a novel <it>in-vitro </it>biofilm model for studying the adherence of <it>P. aeruginosa </it>to hydrogel CLs was established. Nutritional and interfacial conditions similar to those in the eye of a CL wearer were created through the involvement of a solid:liquid and a solid:air interface, shear forces and a complex artificial tear fluid. Bioburdens varied depending on the CL material and biofilm maturation occurred after 72 h incubation. Whilst a range of biofilm morphologies were visualised including dispersed and adherent bacterial cells, aggregates and colonies embedded in extracellular polymer substances (EPS), EPS fibres, mushroom-like formations, and crystalline structures, a compact and heterogeneous biofilm morphology predominated on all CL materials.</p> <p>Conclusions</p> <p>In order to better understand the process of biofilm formation on CLs and to test the efficacy of CL care solutions, representative <it>in-vitro </it>biofilm models are required. Here, we present a three-phase biofilm model that simulates the environment in the eye of a CL wearer and thus generates biofilms which resemble those commonly observed <it>in-situ</it>.</p

Efficiency of biofilm removal by combination of water jet and cold plasma: an in-vitro study

Background: Peri-implantitis therapy is a major problem in implantology. Because of challenging rough implant surface and implant geometry, microorganisms can hide and survive in implant microstructures and impede debridement. We developed a new water jet (WJ) device and a new cold atmospheric pressure plasma (CAP) device to overcome these problems and investigated aspects of efficacy in vitro and safety with the aim to create the prerequisites for a clinical pilot study with these medical devices. Methods: We compared the efficiency of a single treatment with a WJ or curette and cotton swab (CC) without or with adjunctive use of CAP (WJ + CAP, CC + CAP) to remove biofilm in vitro from rough titanium discs. Treatment efficacy was evaluated by measuring turbidity up to 72 h for bacterial re-growth or spreading of osteoblast-like cells (MG-63) after 5 days with scanning electron microscopy. With respect to application safety, the WJ and CAP instruments were examined according to basic regulations for medical devices. Results: After 96 h of incubation all WJ and CC treated disks were turbid but 67% of WJ + CAP and 46% CC + CAP treated specimens were still clear. The increase in turbidity after WJ treatment was delayed by about 20 h compared to CC treatment. In combination with CAP the cell coverage significantly increased to 82% (WJ + CAP) or 72% (CC + CAP), compared to single treatment 11% (WJ) or 10% (CC). Conclusion: The newly developed water jet device effectively removes biofilm from rough titanium surfaces in vitro and, in combination with the new CAP device, biologically acceptable surfaces allow osteoblasts to grow. WJ in combination with CAP leads to cleaner surfaces than the usage of curette and cotton swabs with or without subsequent plasma treatment. Our next step will be a clinical pilot study with these new devices to assess the clinical healing process

Effects of cold atmospheric pressure plasma and disinfecting agents on Candida albicans in root canals of extracted human teeth

Reinfection in endodontically treated teeth is linked to the complexity of the root canal system, which is problematic to reach with conventional disinfection methods. As plasma is expected to have the ability to sanitize narrow areas, the aim of this study was to analyze the effect of cold atmospheric pressure plasma (CAP) on Candida albicans in root canals of extracted human teeth. CAP was applied as mono treatment and in combination with standard endodontic disinfectants (sodium hypochlorite, chlorhexidine and octenidine). Disinfection efficiency was evaluated as reduction of the logarithm of colony forming units per milliliter (log10 CFU/mL) supported by scanning electron microscopy as imaging technique. Plasma alone showed the highest reduction of log10 CFU, suggesting the best disinfection properties of all tested agents. © 2020 The Authors. Journal of Biophotonics published by Wiley-VCH GmbH

In-Vitro Biofilm Removal Efficacy Using Water Jet in Combination with Cold Plasma Technology on Dental Titanium Implants

Peri-implantitis-associated inflammation can lead to bone loss and implant failure. Current decontamination measures are ineffective due to the implants&rsquo; complex geometry and rough surfaces providing niches for microbial biofilms. A modified water jet system (WaterJet) was combined with cold plasma technology (CAP) to achieve superior antimicrobial efficacy compared to cotton gauze treatment. Seven-day-old multi-species-contaminated titanium discs and implants were investigated as model systems. The efficacy of decontamination on implants was determined by rolling the implants over agar and determining colony-forming units supported by scanning electron microscopy image quantification of implant surface features. The inflammatory consequences of mono and combination treatments were investigated with peripheral blood mononuclear cell surface marker expression and chemokine and cytokine release profiles on titanium discs. In addition, titanium discs were assayed using fluorescence microscopy. Cotton gauze was inferior to WaterJet treatment according to all types of analysis. In combination with the antimicrobial effect of CAP, decontamination was improved accordingly. Mono and CAP-combined treatment on titanium surfaces alone did not unleash inflammation. Simultaneously, chemokine and cytokine release was dramatically reduced in samples that had benefited from additional antimicrobial effects through CAP. The combined treatment with WaterJet and CAP potently removed biofilm and disinfected rough titanium implant surfaces. At the same time, non-favorable rendering of the surface structure or its pro-inflammatory potential through CAP was not observed

Antimikrobielle Wirksamkeit von kaltem Atmosphärendruckplasma drei verschiedener Quellen an Biofilmen auf Kunststoffoberflächen im Hinblick auf eine Anwendbarkeit zur Behandlung chronischer Wunden

Die Inaktivierung von Bakterien durch Antiseptika, z. B. auf chronischen Wunden ist unter anderem aufgrund der Bildung von Biofilmen erschwert. Ebenso stellt die Entwicklung von Resistenzen gegenüber Antibiotika ein immer größer werdendes Problem bei der Behandlung von Infektionen dar. Zudem ist die antimikrobielle Behandlung nur ein Teilaspekt, um chronisch infizierte Wunden in einen regenerativen Heilungsprozess zu überführen. Daher sind neue, alternative Behandlungsstrategien von hoher Bedeutung. Hierfür scheint physikalisches Plasma aufgrund seiner antimikrobiellen wie auch wundheilungsfördernder Wirkungsweise eine aussichtsreiche Perspektive darzustellen. Zur Erzeugung von sog. Tissue Tollerablen Plasma (TTP) stehen verschiedene Plasmaquellen zur Verfügung, die zur Anwendung gegen Mikroorganismen in Biofilmen in Frage kommen. In der vorliegenden Arbeit wurden der kinpen09 und zwei Dielektrisch-Behinderte-Oberflächen-Entladungs-Quellen, die Conplas- und die Epoxidharz-Plasmaquelle, auf ihre antimikrobielle Wirkungsweise mit Argonplasma mit und ohne Sauerstoffbeimischung und mit Luftplasma v. a. an Biofilmen mit P. aeruginosa SG81 und S. epidermidis RP62A untersucht. Mit dem kinpen09 wurde zusätzlich die antimikrobielle Effektivität von Plasma mit Helium oder Stickstoffbeimischungen getestet. Bei Einsatz des kinpen09 zeigte sich Argon als das antimikrobiell effektivste Trägergas. Bei der Epoxidharz-Plasmaquelle war Luftplasma am wirksamsten. Bei der Conplas sind Luft- und Argonplasma etwa gleich effizient gegen mikrobielle Biofilme. Die Reduktionsraten bei Argonplasma mit dem kinpen09 und Conplas lagen nach 300 s Expositionszeit bei P. aeruginosa bei ca. 5 log10 und mit dem kinpen09 bei S. epidermidis bei 3 log10. Diese Reduktionsraten übersteigen mit Ausnahme von 300 s Ar+O2-Plasma (kinpen09) die Wirksamkeit von Chlorhexidin (0,1 %), einem Standard-Antiseptikum zur Behandlung von Biofilmen, nach 10 min Behandlungsdauer von ca. 1,5 log10 signifikant (p < 0,005). In Übereinstimmung zur Literatur lässt sich aus den Ergebnissen ableiten, dass v. a. Sauerstoffradikale für die antimikrobielle Wirksamkeit verantwortlich sind. Neben der alleinigen Anwendung von TTP könnten Kombinationsbehandlungen mit Antiseptika aussichtsreiche Verfahren zur gezielten Inaktivierung von Mikroorganismen in Biofilmen und zur Modulation von Wundheilungsprozessen darstellen.The inactivation of bacteria by antiseptics, like in chronic wounds, is complicated by microbial biofilms. Just as the development of bacterial resistance against antibiotics is an increasing problem in the treatment of infections. Additionally, the antimicrobial strategy is only one aspect to lead chronically infected wounds into a stable regeneration process. Therefore, new and alternative strategies are necessary and are particularly interesting. Here, physical plasma with its antimicrobial efficacy and stimulation of wound healing seems to be a promising option. Different plasma sources are available to generate tissue tolerable plasma (TTP). For this work, the tissue tolerable plasma-jet kinpen09 and two different surface dielectric barrier discharges, the ”Conplas”- and “Epoxidharz”- plasma sources, which seem to be tissue tolerable by their physical properties, were tested for their antimicrobial efficacy against the biofilm forming bacteria P. aeruginosa SG81 and S. epidermidis RP62A on biofilms and uncoated on agar. The working gas helium and admixtures of nitrogen to argon or helium were additionally tested by the kinpen09 plasma source. With the kinpen09 argon was the most antimicrobial effective working gas. The “Epoxidharz” plasma source with the ambient air as working gas showed the highest antimicrobial effect. The antimicrobial effectiveness of the Conplas was similar for argon and air plasma. The reduction rates for argon after 300 s treatment time on P. aeruginosa with the kinpen09 and Conplas was approximately 5 log10 and the treatment on S. epidermidis at 3 log10. These reductions are superior to the efficacy of Chlorhexidin (0.1 %), a standard antiseptic solution used especially for biofilm treatment. The Chlorhexidine treatment resulted in a reduction of 1.5 log10 after 10 min treatment time, which is significantly (p < 0.005) lower than the plasma treatment, except the 300 s Ar+O2-plasma treatment with the kinpen09. In comparison with the scientific literature different indications after the experiments suggest that the main antimicrobial components of plasma are chiefly caused by oxygen radicals. Besides, the single application of TTP for antimicrobial treatments, the use of a combination of plasma and different antiseptics could be promising to inactivate microorganism in biofilms and to modulate wound healing processes. Currently, in comparison to the three tested plasma sources the kinpen09 is the favorite plasma source for a future application in wound treatment, because it is flexible in usage and showed a high antimicrobial efficacy

Impact of cold atmospheric pressure argon plasma on antibiotic sensitivity of methicillin-resistant Staphylococcus aureus strains in vitro

Aim: The antimicrobial activity of cold atmospheric pressure plasma (CAP), also called tissue tolerable plasma (TTP), could be a promising option to eradicate methicillin-sensitive as well as methicillin-resistant strains, which often colonize chronic wounds. Currently, the influence of CAP on the susceptibility of to antibiotics is scarcely known, but could be important for treatment of wounds. Therefore, the aim of this study was to investigate whether CAP has an impact on the susceptibility of different strains to different antibiotics.Method: For assessment, the agar diffusion test with different antibiotic test disks (cefuroxime, gentamicin, oxacillin, vancomycin, ciprofloxacin, co-trimoxazole, clindamycin, erythromycin) was used. Test strains were spread on agar plates and CAP treated before the antibiotic disks were placed. After 24 hours cultivation, the inhibited growth zones were measured and differences statistically evaluated.Results: In most cases, CAP had a negligible influence on the susceptibility to antibiotics. For two strains, the susceptibility significantly decreased to β-lactam antibiotics. Conclusion: Because CAP can influence the antibiotic susceptibility of , before conducting combined treatment with local plasma application on wounds and systemic antibiotics, their interaction must be analysed to exclude unwanted combination effects