Eradication of Pseudomonas aeruginosa Biofilms by Atmospheric Pressure Non-Thermal Plasma

Abstract

Bacteria exist, in most environments, as complex, organised communities of sessile cells embedded within a matrix of self-produced, hydrated extracellular polymeric substances known as biofilms. Bacterial biofilms represent a ubiquitous and predominant cause of both chronic infections and infections associated with the use of indwelling medical devices such as catheters and prostheses. Such infections typically exhibit significantly enhanced tolerance to antimicrobial, biocidal and immunological challenge. This renders them difficult, sometimes impossible, to treat using conventional chemotherapeutic agents. Effective alternative approaches for prevention and eradication of biofilm associated chronic and device-associated infections are therefore urgently required. Atmospheric pressure non-thermal plasmas are gaining increasing attention as a potential approach for the eradication and control of bacterial infection and contamination. To date, however, the majority of studies have been conducted with reference to planktonic bacteria and rather less attention has been directed towards bacteria in the biofilm mode of growth. In this study, the activity of a kilohertz-driven atmospheric pressure non-thermal plasma jet, operated in a helium oxygen mixture, against Pseudomonas aeruginosa in vitro biofilms was evaluated. Pseudomonas aeruginosa biofilms exhibit marked susceptibility to exposure of the plasma jet effluent, following even relatively short (∼10′s s) exposure times. Manipulation of plasma operating conditions, for example, plasma operating frequency, had a significant effect on the bacterial inactivation rate. Survival curves exhibit a rapid decline in the number of surviving cells in the first 60 seconds followed by slower rate of cell number reduction. Excellent anti-biofilm activity of the plasma jet was also demonstrated by both confocal scanning laser microscopy and metabolism of the tetrazolium salt, XTT, a measure of bactericidal activity.

Dielectric Barrier Discharge Atmospheric Cold Plasma for Inactivation of Pseudomonas aeruginosa Biofilms

“…27,41 According to the results obtained from the XTT assay, even after extended treatment time of 300 s, an average of 30% of cells in biofilms were still metabolically active. This outcome was also observed by Alkawareek et al, 42 who reported that ACP inactivated 85% of P. aeruginosa cells according to colony counts, while the XTT absorbance value corresponded to 36%. Borges et al 42 observed similar bacterial response to stress, when P. aeruginosa biofilm was subjected to naturally derived compounds with 30% of cell metabolic activity remaining after the treatment.…”

“…This outcome was also observed by Alkawareek et al, 42 who reported that ACP inactivated 85% of P. aeruginosa cells according to colony counts, while the XTT absorbance value corresponded to 36%. Borges et al 42 observed similar bacterial response to stress, when P. aeruginosa biofilm was subjected to naturally derived compounds with 30% of cell metabolic activity remaining after the treatment. In general, the plate count assay accounts only for culturable bacterial cells and does not take into account cells that might still be metabolically active.…”

Cold Plasma Inactivation of Escherichia coli O157:H7 Biofilms

“…In previous studies using a plasma system based on a 99.5% helium, 0.05% oxygen feed gas, Alkawareek and others (Alkawareek et al, 2012), a 15 s treatment reduced Pseudomonas aeruginosa biofilms by 0.8 cfu/ml at a pulse frequency of 20 kHz and by 1.6 log cfu/ml at 40 kHz. In contrast, Niemira et al (Niemira et al, 2014), using the same equipment and feed gas (air) used in the present study, showed no clear association of antimicrobial efficacy against Salmonella biofilms for pulse frequencies ranging from 24 to 48 kHz.…”

“…A flow of feed gas (dried air) at 60 psi drives the plasma arc outward from the surrounding Teflon cowling, expanding and cooling it. Varying the pulse frequency is known to be a factor in the antimicrobial efficacy of cold plasma (Alkawareek et al, 2012). Frequencies ranging from 23 to 48 kHz were evaluated, with power consumption of 522 to 549 W.…”

Application of post-discharge region of atmospheric pressure argon and air plasma jet in the contamination control of Candida albicans biofilms

“…As an antimicrobial strategy, the advantages of non-thermal plasmas operated at atmospheric pressure are the simple design, low cost of construction/operation, usage of nontoxic gases, with operating conditions of gas at or near room temperature, and no harmful residues (Gaunt et al, 2006;Kong et al, 2009;Laroussi, 2002). In last years, much effort and progress have been done in order to elucidate the exact mechanisms leading to bacterial or fungal inactivation by the action of electric plasma (Alkawareek et al, 2012;Chen et al, 2014;Laroussi, 2002;Mai-Prochnow et al, 2014;Taghizadeh et al, 2015;Traba and Liang, 2015). It is considered that several plasma products play an important role in this process, namely reactive oxygen species (ROS), such as ozone, atomic oxygen, superoxide, hydrogen peroxide and hydroxyl radicals (Kong et al, 2009), and reactive nitrogen species (RNS), UV radiation, and charged particles (Gaunt et al, 2006;Laroussi and Leipold, 2004;Ma et al, 2008).…”