Due to the great increase in the non-thermal plasma (NTP) bio-applications, especially thanks to its antimicrobial properties, many types of NTP generating devices have been developed recently. However, a comparison of these devices is difficult due to the differences in the setup of studies testing them, e.g., in species of microorganisms used and sample preparations. In this study, we optimized a robust and reproducible standard protocol using Bacillus subtilis spores and applied it to compare seven different NTP generating devices in terms of technical parameters and sporicidal properties. Inhibition zones determined using the Aurora software and the complete inhibition of bacteria growth induced by the NTP treatment were analyzed to determine both local and overall effects, respectively. The highest sporicidal efficacy of the tested devices was achieved by the volume dielectric barrier discharge from Wroclaw, which inhibited 99.9% of colony forming units after 30 min of exposure. To our knowledge, a comparative study of this extent has not been published to date. The presented protocol is based on an established bacterial method and can therefore serve as a general standard for an effective comparison of NTP sources across laboratories worldwide.

Consorzio RFX Padua Italy

Department of Biotechnology University of Chemistry and Technology Prague Czech Republic

Department of Electrical Engineering Fundamentals Wroclaw University of Science and Technology Wrocław Poland

Department of Physics and Measurements University of Chemistry and Technology Prague Czech Republic

Department of Physics Giuseppe Occhialini University of Milano Bicocca Milan Italy

Faculty of Nuclear Sciences and Physical Engineering Czech Technical University Prague Prague Czech Republic

Istituto per la Scienza e la Tecnologia dei Plasmi CNR Padua Italy

Zobrazit více v PubMed

Scholtz, V. et al. Non-thermal plasma treatment of ESKAPE pathogens: a review. Front. Microbiol.12, 89 (2021). PubMed PMC

Varilla, C., Marcone, M. & Annor, G. A. Potential of cold plasma technology in ensuring the safety of foods and agricultural produce: a review. Foods9, 1435 (2020). PubMed PMC

von Woedtke, T., Emmert, S., Metelmann, H. R., Rupf, S. & Weltmann, K. D. Perspectives on cold atmospheric plasma (CAP) applications in medicine. Phys. Plasmas27, 070601 (2020).

Boekema, B. K. H. L. et al. A new flexible DBD device for treating infected wounds: in vitro and ex vivo evaluation and comparison with a RF argon plasma jet. J. Phys. D: Appl. Phys.49, 044001 (2016).

Hahn, V., Brandenburg, R. & von Woedtke, T. Springer, DIN SPEC 91315: a first attempt to implement mandatory test protocols for the characterization of plasma medical devices. In Comprehensive Clinical Plasma Medicine (eds. Metelmann, H. R., von Woedtke, T., Weltmann, K. D.) (2018). 10.1007/978-3-319-67627-2_35.

Mann, M. S., Schnabel, U., Weihe, T. & Weltmann, K. D. & von Woedtke, T. A reference technique to compare the antimicrobial properties of atmospheric pressure plasma sources. Plasma Med.5, 1 (2015).

Alves, L. L. et al. Foundations of plasma standards. Plasma Sources Sci. Technol.32, 023001 (2023).

Mann, M. et al. Introduction to DIN-Specification 91315 based on the characterization of the plasma jet kINPen® MED. Clin. Plasma Med.4, 789 (2016).

Jablonowski, H. et al. Characterization and comparability study of a series of miniaturized neon plasma jets. J. Phys. D: Appl. Phys.57, 195202 (2024).

Shaw, A., Seri, P., Borghi, C. A., Shama, G. & Iza, F. A reference protocol for comparing the biocidal properties of gas plasma generating devices. J. Phys. D: Appl. Phys.48, 484001 (2015).

Raguse, M. et al. Improvement of biological indicators by uniformly distributing Bacillus subtilis spores in monolayers to evaluate enhanced spore decontamination technologies. Appl. Environ. Microbiol.82(7), 2031–2038 (2016). PubMed PMC

Ulrich, N. et al. Experimental studies addressing the longevity of Bacillus subtilis spores—the first data from a 500-year experiment. PLOS ONE13, 12 (2018). PubMed PMC

Khun, J., Jirešová, J., Kujalová, L., Hozák, P. & Scholtz, V. Comparing the biocidal properties of non-thermal plasma sources by reference protocol. Eur. Phys. J. D71, 263 (2017).

Fridman, G. et al. Comparison of direct and indirect effects of non-thermal atmospheric-pressure plasma on bacteria. Plasma Processes Polym.4, 370–375 (2007).

Georgescu, N., Apostol, L. & Gherendi, F. Inactivation of Salmonella enterica serovar typhimurium on egg surface, by direct and indirect treatments with cold atmospheric plasma. Food Control76, 52–61 (2017).

Martusevich, A. K. et al. Cold argon athmospheric plasma for biomedicine: biological effects, applications and possibilities. Antioxidants11, 1262 (2022). PubMed PMC

Lou, B-S. et al. Helium/argon-generated cold atmospheric plasma facilitates cutaneous wound healing. Front. Bioeng. Biotechnol.8, 683 (2020). PubMed PMC

Shimizu, T., Sakiyama, Y., Graves, D. B., Zimmermann, J. L. & Morfill, G. E. The dynamics of Ozone generation and mode transition in air surface micro-discharge plasma at atmospheric pressure. New. J. Phys.14, 103028 (2012).

Lou, B-S. et al. Parameters affecting the antimicrobial properties of cold atmospheric plasma jet. J. Clin. Med.8(11), 1930 (2019). PubMed PMC

De Masi, G., Gareri, C. & Cordaro, L. Plasma coagulation controller: a low- power atmospheric plasma source for accelerated blood coagulation. Plasma Med.8, 3 (2018).

Brun, P. et al. Antibacterial efficacy and mechanisms of action of low power atmospheric pressure cold plasma: membrane permeability, biofilm penetration and antimicrobial sensitization. J. Appl. Microbiol.125, 2 (2018). PubMed

do Nascimento, F. et al. Plasma electrode dielectric barrier discharge: development, characterization and preliminary assessment for large surface decontamination. Plasma Chem. Plasma Process.43, 1791–1817 (2023).

Trebulova, K. et al. Antimycotic effects of the plasma gun on the yeast Candida glabrata tested on various surfaces. Plasma Process. Polym.21(9), 2400057 (2024).

Hrudka, J. et al. Automatic image analysis of the effects of non-thermal plasma on mold growth. In Proceedings of 25th International Symposium on Plasma Chemistry, Kyoto, Japan (2023).

KC, S. K., Sharma, S., Shrestha, R. & Subedi, D. P. Electrical characterization of an atmospheric pressure plasma jet. J. Nep Phys. Soc.5, 85–90 (2019).

Martines, E. et al. A novel plasma source for sterilization of living tissues. New. J. Phys.11, 115014 (2009).

Neretti, G. et al. Characterization of a plasma source for biomedical applications by electrical, optical, and chemical measurements. Plasma Process. Polym.15, 1800105 (2018).

Brun, P. et al. Disinfection of ocular cells and tissues by atmospheric-pressure cold plasma. PLoS One7(3), e33245 (2012). PubMed PMC

Brun, P. et al. Antibacterial efficacy and mechanisms of action of low power atmospheric pressure cold plasma: membrane permeability, biofilm penetration and antimicrobial sensitization. J. Appl. Microbiol.125(2), 398–408 (2018). PubMed

Cordaro, L. et al. On the electrical and optical features of the plasma coagulation controller low temperature atmospheric plasma jet. Plasma2, 156–167 (2019).

Zampieri, L. & Martines, E. Design and optimization of a radio frequency plasma jet for biomedical applications. In 48th EPS Conference on Plasma Physics (2022). http://ocs.ciemat.es/EPS2022PAP/pdf/P1b.303.pdf.

Hofmann, S., van Gessel, A. F. H., Verreycken, T. & Bruggeman, P. Power dissipation, gas temperatures and electron densities of cold atmospheric pressure helium and argon RF plasma jets. Plasma Sources Sci. Technol.20, 065010 (2011).

Czapka, T., Maliszewska, I. & Winkler, A. Decontamination of polymeric surgical sutures covered with bacterial biofilms using nonthermal plasma. Plasma Chem. Plasma Process.41, 227–243 (2021).

Nowinski, D., Czapka, T. & Maliszewska, I. Effect of multiple nonthermal plasma treatments of filamentous fungi on cellular phenotypic changes and phytopathogenicity. Int. J. Food Microbiol.2(408), 110428 (2024). PubMed

Kogelschatz, U. Dielectric-barrier discharges: their history, discharge physics, and industrial applications. Plasma Chem. Plasma Process.23, 1–46 (2003).

Czapka, T., Maliszewska, I. & Olesiak-Bańska, J. Influence of atmospheric pressure non-thermal plasma on inactivation of biofilm cells. Plasma Chem. Plasma Process.38, 1181–1197 (2018).

Klenivskyi, M. et al. Portable and affordable cold air plasma source with optimized bactericidal effect. Sci. Rep.14, 15930 (2024). PubMed PMC

MiniJet, Heuermann HF-Technik GmbH. https://hhft.de/10w-minijet (2024).

Brotankova, J., Mlynar, J., Pfeifer, M. & Svoboda, V. Plasmalab@ctu - new facilities in support of fusion education. In 20th Conference of Czech and Slovak Physicists Proceedings 186–187 (2020). https://www.plasmalab.cz.