Effects of combining lysozyme, ultrasound, and heat treatments on the inactivation of Bacillus subtilis spores.

The inactivation of bacterial spores remains a critical challenge in food sterilization. This study aimed to investigate the effects of combining ultrasound with heat and lysozyme treatment on the key structures of Bacillus subtilis spores. Spores were subjected to 600 W ultrasound combined with heat treatments at 75 ℃ and 90 ℃ and lysozyme at concentrations of 0.2% and 0.4% for 30 min. The combined treatments exhibited a synergistic effect on spores inactivation, with the most pronounced effect observed with the 600W-90 ℃-0.4% lysozyme treatment, achieving a reduction of 4.7 log CFU/mL. This treatment significantly increased the permeability of the inner membrane, leading to severe hydration of the spore core and substantial release of dipicolinic acid. Flow cytometry results revealed significant damage to the spore's inner membrane. Fourier transform infrared spectroscopy analysis indicated a decrease in the stability of the cortex and cell wall peptidoglycan, a phase transition of inner membrane phospholipids from gel to liquid crystal state, and increased fluidity of the inner membrane. The obtained results demonstrated that combining ultrasound, heat, and lysozyme treatments caused substantial damage to the inner membrane, cortex, cell wall and coat of spores, leading to effective spore inactivation.