UV-C and hydration state drive pulsed light-induced proteome damage in

Introduction: Pulsed light (PL) is a non-thermal processing technology that inactivates microorganisms through high-intensity pulses of polychromatic light, including ultraviolet-C (UV-C). While the germicidal effect of PL has been widely studied, its impact on microbial proteomes remains poorly understood. Here, we investigate the proteomic response of Methods: Results: PL treatment induced a significantly greater proteomic alteration compared to UV-C, particularly in spores suspended in water, where the number of proteins with decreased abundance was ~6-fold higher than in spores sprayed on a polystyrene surface. Proteomic analysis revealed that the effect of PL in water was primarily due to UV-C 254 nm, whereas on polystyrene, UV-C 254 nm had no significant impact. Furthermore, proteins most affected by PL were enriched in photosensitive amino acids such as tryptophan, histidine, tyrosine, cysteine, and methionine, suggesting oxidation and photoreactivity as key degradation mechanisms. Discussion: Although the overall inactivation rate could not be directly correlated with proteome damage, we identified that core proteins involved in DNA and RNA protection and repair were specifically targeted by PL. These findings provide new insights into the molecular mechanisms underlying PL-mediated microbial inactivation and highlight the role of protein photodamage in spore susceptibility.