Morphological and transcriptomic responses of aflatoxin-producing Aspergillus flavus to atmospheric cold plasma treatment

Mycotoxins contaminate about 60% of the world’s grain and legume are a health and economic burden. Atmospheric cold plasma (ACP) is a rapidly growing technology that could be used as a dry, non-thermal, and rapid process for fungal decontamination of food commodities. However, the specific mechanisms and modes of action that result in fungal inactivation by ACP are not well understood. The purposes of this study were to evaluate: 1). the effectiveness of the ACP on aflatoxin-producing Aspergillus flavus at different distances, different exposure times, and with a particular type of actuator; 2). the transcriptomic response of Aspergillus flavus to a surface dielectric barrier discharge (SDBD) atmospheric cold plasma treatment, thereby gaining insights on the possible mechanisms of inactivation through RNA sequencing. In this study, ACP’s inactivation of spores and mycelia were evaluated, and the morphological effects of ACP treatment were observed by scanning electron microscopy. ACP treatment resulted in up to 4 log reduction (Killing 99.99%) of the A. flavus spores with decreasing inactivation efficiency at increasing treatment distance. A significant reduction in A. flavus mycelia growth after ACP treatment was observed. A. flavus spores treated by ACP appeared warped with some of the treated spores sticking together, and ACP treated mycelia appeared wrinkled. Some treated spores had smooth surfaces. In ACP-treated spores, the KEGG pathway indicated up-regulated genes associated with ribosome and ribosomal biogenesis, whereas the down-regulated genes were in carbon metabolism and pentose phosphate pathways. ACP treatment of wet mycelia of A. flavus triggered the upregulation of nitrogen metabolism and aflatoxin biosynthesis and the down-regulation of carbon metabolism and peroxisome. For dry mycelia of A. flavus, ACP treatment caused the up-regulation of pentose and glucuronate interconversion and the down-regulation of steroid biosynthesis. DEGs in each group revealed regulatory changes in genes involved in aflatoxin biosynthesis, a variety of types of antioxidants, stress defense mechanisms, and antioxidant related mechanisms such as superoxide dismutase, catalase, glutathione, vitamin, etc. Future improvements on the ACP, actuators and optimization of other parameters will enhance inactivation efficiency. ACP would then become an alternative to the more commonly used agricultural/foodborne decontamination treatments.