Resistance of Alicyclobacillus acidoterrestris spores to atmospheric cold plasma: Insights from sporulation temperature and mechanism analysis

Environmental conditions during sporulation, such as temperature, pH value, and mineral content, can significantly influence the formation production and resistance of spores. In this study, we investigated the sporulation capacity and resistance of Alicyclobacillus acidoterrestris (AAT) spores obtained at different temperatures (25 °C, 35 °C, 45 °C) to Dielectric Barrier Discharge Atmospheric cold plasma (DBD-ACP, 20 and 30 kV for 2–8 min). The results indicated that the efficiency of sporulation was highest at the optimum temperature (45 °C), deviation from optimal temperature would have a negative effect on sporulation. Otherwise, sporicidal efficiency of DBD-ACP increased with longer treatment time and lower sporulation temperatures, suggesting that AAT spores exhibit weaker resistance to DBD-ACP at lower sporulation temperature. Results from the release of DPA, and electron microscopic observations confirmed that DBD-ACP caused disruption of the cortex and membrane structure of AAT spores, leading to the release of intracellular contents and subsequent spore's death. Further analysis through proteomics revealed intriguing insights into the differences between spores formed at 25 °C and those at 45 °C. Specifically, spores formed at 25 °C exhibited lower expression levels of proteins related to sporulation, intracellular energy metabolism, membrane transport, and significant down-regulation of proteins involved in peptidoglycan and spore coat formation compared to those at 45 °C. These alterations likely contribute to the development of a thinner and looser spore structure, rendering it less effective in defending, i.e. less resistant to DBD-ACP treatment. Industrial relevance AAT spores' resistance to pasteurization and persistence in NFC juices impact juice quality and safety. This study investigates the resistance of AAT spore to DBD-ACP treatment under different sporulation temperatures. Insights into sporicidal behavior of DBD-ACP and the effect of sporulation temperature on resistance provide essential knowledge for optimizing juice processing and enhancing microbial safety. The findings offer practical guidance for the juice industry to develop improved preservation strategies, ensuring longer shelf life and consumer satisfaction.