Cross-stress adaptation enhances the survival and stability of freeze-dried lactic acid bacteria

Lactic acid bacteria (LAB), widely used as probiotics in the food industry, are typically freeze-dried to extend shelf life, although this process often compromises cell viability. While individual stress treatments have been shown to enhance bacterial survival, the potential of combined stresses remains underexplored. In this study, we investigated the effect of acid-cold cross-stress adaptation on the freeze-drying tolerance of LAB. The results demonstrated that cross-stress pretreatment significantly improved survival rates, fermentation performance, and storage stability. Specifically, Lacticaseibacillus rhamnosus exhibited a 1.65-fold increase in survival and 20% higher viability after 21 days of storage. Mechanistic analysis revealed that this strategy preserved cell membrane integrity, elevated lactate dehydrogenase and ATPase activity, and increased the unsaturated-to-saturated fatty acid ratio. These findings highlight the synergistic benefits of multi-stress conditioning and provide a promising strategy for improving the industrial preservation of probiotic cultures.

The online version contains supplementary material available at 10.1007/s44463-025-00007-y.