Low-temperature plasma and plasma-activated water as alternative novel technologies for postharvest disease control

The need for new sustainable technologies to reduce losses and maintain the quality of fruits and vegetables during postharvest storage has grown during recent years. Low-Temperature Plasma (LTP) and Plasma-Activated Water (PAW) are currently proposed as promising tools for inactivation of microbial contaminants, including foodborne pathogens and microorganisms responsible for postharvest decay. The effects of both direct and indirect applications of atmospheric air plasma against major fungal pathogens (Botrytis cinerea, Monilinia fructicola, Fusarium graminearum, Aspergillus carbonarius, Penicillium italicum, Penicillium digitatum, Alternaria alternata) and contaminant bacteria (e.g., Bacillus subtilis) were investigated in in-vitro assays using different treatment conditions and exposure times. Three different plasma sources, i.e., Surface Dielectric Barrier Discharge (SDBD), Atmospheric Pressure Plasma Jet (APPJ), and Volume Dielectric Barrier Discharge (VDBD), were applied. Albino mutants of Botrytis cinerea and Aspergillus carbonarius were used to explore the protective role of fungal melanin on sensitivity to plasma exposure, with mutants showing higher sensitivity to treatments compared to the melanized wild-type strains. As expected, inhibitory effects increased with higher exposure times. The complete spore inactivation was obtained after a few seconds (VDBD) or minutes (SDBD and APPJ) of exposure. Major structural damages to the conidia surface after plasma treatment were assessed by Scanning Electron Microscopy (SEM) analysis. An early etching and later perforation of cell walls up to complete cell disruption was observed. A decrease in viability of conidia and an increase in their membrane permeability was assessed by fluorescence-based assays. Differences in the response to plasma among species that could be related to the fungal cell structure and composition were observed, with the strongly melanized conidia of A. carbonarius and A. alternata showing the lowest sensitivity to the treatments. Antibacterial and antifungal activities of PAW having different contents in reactive species (hydrogen peroxide, nitrite and nitrate), pH, and ORP under different treatment conditions (i.e., plasma source, exposure time, gas flow) were evaluated. Results revealed a close correlation between chemical properties of PAW and inhibition rates for bacteria and fungi. Furthermore, inhibitory effect of PAW against all the tested microbial species was proved stable, with a persistence of at least 30 min after water activation. The efficacy of PAW was also tested on fruits of Citrus sinensis cv. Fukumoto artificially inoculated with P. italicum with a total inhibition of the infection development and symptoms appearance recorded on treated fruit. Experiments on cherry fruits artificially inoculated with B. cinerea and M. fructicola and exposed to SDBD air plasma demonstrated that the application of cold plasma on fruits may significantly extend their shelf life by direct inactivation of fungi and possible activation of plant defense responses.