BIOPLASMAS & PLASMAS WITH LIQUIDS - Joint Conference of COST ACTIONS TD1208 “Electrical discharges with liquids for

future
applications” & MP1101 Biomedical Applications of Atmospheric Pressure Plasma Technology, Bertinoro, Italy, 13th-17th September 2015

Non-thermal plasma activated water (PAW) – a new mycobactericidal

agent

Andra-Cristina BOSTANARU1, Eugen HNATIUC2*, Mihai MARES1

1
Laboratory of Antimicrobial Chemotherapy. University of Agricultural Sciences
and Veterinary Medicine “Ion Ionescu de la Brad”, Iasi, 700489, Romania
2
Technical University Gh. Asachi, Iasi-Romania

Mycobacteria are still important agents of humans and animals infections. The major pathogen
is Mycobacterium tuberculosis (MTB) that infects almost 1.5 billion people and kills 1.3 million
people each year [1].

Disinfection of MTB and non-tuberculous mycobacteria (NTM) can be a problem, as

mycobacteria are among the most resistant bacteria [2]. Recently, plasma-activated water
(PAW) has emerged as an efficient biocide that could be used in pathogenic germs inactivation
[3]. Among the advantages of PAW, its sterilization effect and lack of toxicity are the most
valuable characteristics [4].

The aim of this study was to evaluate the mycobacteria disinfection efficacy of plasma activated
water obtained by the activation of water with GlidArc reactor and explore its mechanism of
action. The experiment was performed on M. tuberculosis K37Ra, Lehmann and Neumann
(ATCC 25177), M. avium subsp. avium (CECT 7407), M. smegmatis (CECT 3017), M. marinum
(CECT 7091), M. abscessus (CECT 8517), M. chelonae (CECT 8519), M. kansasii (CECT
3030), M. peregrinum (CECT 3023), strains subcultured in Middlebrook 7H9 broth and plated on
8
Löwenstein Jensen (LJ) medium. Suspensions of mycobacteria (10 CFU/mL) were treated by
plasma-activated water (PAW) for different period of time (5, 10, 15, 20, and 30 minutes).

The reduction of mycobacterial burden was evaluated using colony-forming unit (CFU) count
and the formula: log10 (No. CFU before PAW treatment / No. CFU after PAW treatment). In
order to assess the impact of PAW treatment on mycobacterial structure, scanning electron
microscopy (SEM) was used. The integrity of the cell membranes was investigated by UV
spectrophotometry. In addition, X-ray photoelectron spectroscopy (XPS) and Fourier Transform
Infrared Spectrometry (FT-IR) were used to study the interaction of PAW with cell wall of
mycobacteria.

A reduction higher than 5 logarithmic units was observed after an exposure time of 15-min, that
means a powerful mycobactericidal effect. After an exposure time of 20-min, the complete
inactivation of mycobacteria was achieved (sterilization).

Significant reductions in mycobacterial populations were achieved in all strains of tested

mycobacteria, demonstrating the effectiveness of this new approach to treat the mycobacteria.
These findings previously not described in the literature suggest that plasma-activated water
have a mycobactericidal effect exhibited against both Mycobacterium tuberculosis and non-TB
mycobacteria. The oxygen and nitrogen free radicals from PAW are responsible for modification
of the chemical state of the cells and impairing morphology of mycobacteria.

[1] World Health Organization. Global Tuberculosis Control: WHO Report 2013 (WHO, Geneva, 2013).
[2] G. McDonnell, AD. Russell , Clin. Microbiol. Rev. 12 (1999) 147–179.
[3] M. Ursache, R. Moraru, V. Nastasa, E. Hnatiuc and M. Mares, International Conference on Optimization of
Electrical and Electronic Equipment (OPTIM) (2014), 1036-104.
[4] U. K. Ercan, H. Wang, J. Haifeng, G. Freidman. A.D. Brooks and G.J. Suresh, Plasma Processes and Polymers,
10, (2013) 544-555.