Cold atmospheric plasma fumigation suppresses postharvest apple Botrytis cinerea by triggering intracellular reactive oxygen species and mitochondrial calcium

Infection by Botrytis cinerea poses a great threat to the postharvest life of apple fruit. In this study, the effects of cold atmospheric plasma (CAP) fumigation on apple B. cinerea under different exposure times and intensities were investigated. The growth of B. cinerea in vitro and in vivo was significantly suppressed by the CAP fumigation at least 700 μL/L for 5 min. To reveal the possible mechanism of antifungal activity of CAP fumigation, the pathogen was exposed to 700 μL/L and 1000 μL/L for 5 min, respectively. The results indicated that the CAP-treated spores of the pathogen underwent shrinkage, cell membrane collapse and cytoplasmic vacuolation. The results obtained from the fluorescent probe assay and flow cytometry indicated that CAP caused the accumulation of reactive oxygen species (ROS), the elevation of mitochondrial and intracellular Ca2+ levels, and the decrease in mitochondrial membrane potential of the pathogen. Investigation on statues of cell life showed that typical hallmarks of apoptosis in the CAP-treated B. cinerea spores occurred, as indicted by a large degree of increased phosphatidylserine externalization, dysfunction of membrane permeability, DNA fragmentation, distortion of morphology, chromatin condensation, and metacaspase activation observed in B. cinerea spores after CAP fumigation. Overall, CAP fumigation triggered a metacaspase-dependent apoptosis of B. cinerea spores mediated by intracellular ROS burst and Ca2+ elevation via mitochondrial dysfunction and disruption, and therefore reduced the pathogenicity of B. cinerea and suppressed postharvest Botrytis rot of apple fruit. These results would provide an insight into the underlying mechanism of CAP fumigation acting on the pathogen. The CAP fumigation makes much convenient application of CAP in storage environment to deactivate microorganism.