Isolation and characterization of Salmonella Typhimurium SL1344 variants with increased resistance to different stressing agents and food processing technologies.

In this study, resistant variants of Salmonella enterica serovar Typhimurium SL1344 to different stressors were selected. In addition, a genetic and phenotypic study was performed to explore the mechanisms underlying the acquisition of resistance. We isolated 4 variants with increased stable resistance to acid, osmotic stress, high hydrostatic pressure (HHP) and Ultraviolet-C light (UV-C) after repeated rounds of exposure to these agents and outgrowth of survivors. A PEF-resistant variant (SL1344-RS), previously isolated by Sagarzazu et al. (2013), was also included in the analysis. The results indicated that the isolated variants showed resistance to at least one other agent. This increased resistance, in general terms, had a fitness cost in growth, and exerted a variable impact on virulence (mainly in cell adhesion capacity), increased antibiotic resistance but did not influence in biofilm formation capacity. Whole Genome Sequencing (WGS) analysis allowed us to identify the genetic changes responsible for these phenotypic differences, and revealed that in 3 out of the 5 variants (including SL1344-RS) a mutation was found in hnr gene, an anti-sigma factor that promotes RpoS proteolysis. Hence the expression of several rpoS-regulated genes was quantified and found higher in these variants. This increase in RpoS activity would explain the lower growth rates observed in these 3 variants, as it would lead to increased transcription of genes involved in growth arrest and resistance to various types of stress. However, further analysis of a set of 22 additional Salmonella strains obtained from different culture collections indicated that a direct relationship between RpoS activity and stress resistance might not exist within Salmonella.