On the inactivation of Bacillus subtilis spores by surface streamer discharge in humid air caused by reactive species

Abstract

The inactivation of Bacillus subtilis (ATCC 6633) spores deposited on a filter membrane was studied by using low-temperature plasma produced via surface dielectric barrier discharge. Spore samples were carefully prepared to avoid the formation of cell aggregates, and their inactivation was induced by multiple surface streamer discharge driven in a coplanar dielectric barrier discharge electrode geometry by an amplitude-modulated AC high voltage waveforms in humid air at atmospheric pressure. At a discharge duty cycle of 0.4, the surface dielectric barrier discharge is characterised by an average total power of 1.7 W (power density 1.5 W cm−2 and energy density ∼0.3 Wh l−1) and a low gas temperature of the plasma filaments of about 320 K. The spores were exposed by placing a sample holder at a fixed distance of 3 mm from the electrode surface covered by plasma filaments. Particular attention was paid to identifying sporicidal agents employed in the process of inactivation. Since treated samples did not come into direct contact with the streamer filaments and excessive heating was excluded thanks to the low energy density, our results indicate that the spores were inactivated mainly by reactive oxygen and nitrogen species such as O3, H2O2 and NO2–. Discharge-induced damage of the spore structure was evidenced via the detection of dipicolinic acid and leaking of intracellular components. We therefore conclude that B. subtilis spores were inactivated chemically, probably due to failure of the coat structure or membrane of the spore.