Transcriptomic and metabolomic analyses to study the key role by which Ralstonia insidiosa induces Listeria monocytogenes to form suspended aggregates

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Abstract
Ralstonia insidiosa can survive in a wide range of aqueous environments, including food processing areas, and is harmful to humans. It can induce Listeria monocytogenes to form suspended aggregates, resulting from the co-aggregation of two bacteria, which allows for more persistent survival and increases the risk of L. monocytogenes contamination. In our study, different groups of aggregates were analyzed and compared using Illumina RNA sequencing technology. These included R. insidiosa under normal and barren nutrient conditions and in the presence or absence of L. monocytogenes as a way to screen for differentially expressed genes (DEGs) in the process of aggregate formation. In addition, sterile supernatants of R. insidiosa were analyzed under different nutrient conditions using metabolomics to investigate the effect of nutrient-poor conditions on metabolite production by R. insidiosa. We also undertook a combined analysis of transcriptome and metabolome data to further investigate the induction effect of R. insidiosa on L. monocytogenes in a barren environment. The results of the functional annotation analysis on the surface of DEGs and qPCR showed that under nutrient-poor conditions, the acdx, puuE, and acs genes of R. insidiosa were significantly upregulated in biosynthetic processes such as carbon metabolism, metabolic pathways, and biosynthesis of secondary metabolites, with Log2FC reaching 4.39, 3.96, and 3.95 respectively. In contrast, the Log2FC of cydA, cyoB, and rpsJ in oxidative phosphorylation and ribosomal pathways reached 3.74, 3.87, and 4.25, respectively. Thirty-one key components were identified while screening for differential metabolites, which mainly included amino acids and their metabolites, enriched to the pathways of biosynthesis of amino acids, phenylalanine metabolism, and methionine metabolism. Of these, aminomalonic acid and Proximicin B were the special components of R. insidiosa that were metabolized under nutrient-poor conditions.

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