Transcriptome analysis to study expression under desiccation stress of Salmonella enterica subsp. enterica serovar Agona str. 460004 2-1 on low moisture food (cereal)

Salmonella enterica serovar Agona (S. Agona) is a foodborne pathogen that caused recurrent multistate outbreaks associated with cereal between 1998 and 2008, underscoring the endurance of Salmonella over time in low-moisture food (LMF) processing facilities. In this study, we aimed to determine the molecular mechanism of survival of S. Agona in LMF and confirm their impact on phenotype by the knockout study. S. Agona strain (CFSAN 000477), isolated from cereal, was selected for this study. A 100µl suspension with a concentration of ~10^11 cfu/ml was inoculated into 3g of rice cereals. Three replications of inoculated cereals were subjected to desiccation stress (aw ≤ 0.25) for 24h at room temperature (25⁰C). Inoculated cereal samples were collected at 6 timepoints post-inoculation. Cells were separated from the food matrix for RNA extraction. RNA sequencing was performed using the NextSeq 2000 platform. Read counts were generated with Salmon v1.9.0. Downstream analysis was conducted with R and KEGG mapper. There were 1120 differentially expressed genes (DEGs) in S. Agona in response to desiccation stress (Padj < 0.01, |log2FoldChange| >1), with 647 downregulated and 473 upregulated. Functional analysis of downregulated DEGs revealed that most of the genes were associated with metabolic pathways, followed by translation, suggesting slower growth in the surviving population. The top 3 upregulated genes/operons: kdp and ccm operon, and tisB were knocked out and checked for survival study. Approximately 1-2 log reduction (p>0.05) was noticed in the survival of the mutants compared with the wild type. This transcriptome data suggests that Salmonella Agona survives in low-moisture food by conserving energy, lowering metabolism, and reducing replication. RNA sequencing of Salmonella Agona inoculated on cereal at 0-, 0.5-, 1-, 2-, 4- and 24- hour post-drying time points.