The effect of the antimicrobial mechanism of ultrasound (US) in combination with heat (T) and potassium sorbate (PS) treatment on Pichia membranifaciens (P. membranifaciens) is investigated in this study. It is found that ultrasound combined with heat (thermoultrasound [TS]) exhibits a synergistic inactivation efficiency, and the combinations of US, T, and TS with PS also improve inactivation efficacy through reductions of 0.32 ± 0.02 log, 0.73 ± 0.04 log, and 4.50 ± 0.04 log in P. membranifaciens achieved using US + PS, T + PS, and TS + PS, respectively. Further, use of the TS + PS treatment leads to cytoplasmic leakage and a decrease in dehydrogenase activity, as well as a significant increase in cell membrane permeability, causing membrane depolarization and the inward flow of extracellular calcium ions. In addition, the cell membrane composition is detected by infrared spectroscopy, demonstrating that TS + PS treatment results in a decrease in lipid and protein content and an increase in phospholipid content in the cell membrane. Scanning and transmission electron microscopy reveals a deformed cell structure and damage to cell membrane integrity after TS + PS treatment. In addition, the expression of heat shock protein (Hsp82 and Hsp70) genes is shown to decrease after TS and TS + PS treatment compared to the T and T + PS treatment groups, indicating cellular homeostasis has been broken and cells are responding to the damage by activating the protective gene expression program.
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