Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: Adhesion and invasion activity of Salmonella Typhimurium in response to prebiotic pre-treatment. (A) Prebiotic pre-treatment of Caco2 cells reduces combined invasion and adhesion of Salmonella LT2 in an in-vitro setting. 0% prebiotic addition (white bars) are used as the control for comparison of increasing (0.1, 0.5, and 1%) prebiotic treatment concentrations. BioMos and HMO were added at the noted concentrations to differentiated Caco2 cells for 15 min, followed by co-incubation of S. Typhimurium LT2 for 60 min. Invasion and adhesion was measured with a gentamicin protection assay and % associated bacteria values represent the combined percentage of Salmonella cells which invaded or adhered to host cells. * p < 0.05, ** p < 0.02. (B) Expression of virulence factors in S. Typhimurium 14028 ss was evaluated for each prebiotic treatment, using S. Typhimurium 14028 ss without prebiotics as the control. Green up arrows represent an upregulation of genes related to each factor while red down arrows represent repression. (C) The TLR expression pathway for Caco2 cells was overlayed with both HMO and BioMos treated Caco2 expression profiles, with no prebiotic Caco2s as the control for both cases. Teal represents decreased measurement for the gene while red represents increased measurement. Blue and orange genes represent predicted inhibition or activation, respectively. Inset graphs display the log2 fold-change comparison between HMO (1) and BioMos (2).

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Activity Assay, In Vitro, Control, Comparison, Incubation, Bacteria, Expressing, Inhibition, Activation Assay

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: Expression of virulence factors in S. Typhimurium 14028 ss and TLRs in Caco2 Cells. (A) Virulence factor expression in S. Typhimurium added to prebiotic treated Caco2 cells was measured as log 2 fold change from S. Typhimurium exposed to untreated Caco2 cells after 60 min of co-incubation. Orange represents S. Typhimurium add to the BioMos condition and purple represent the HMO condition S. Typhimurium. (B) Toll-like receptor expression in prebiotic treated then infected Caco2 cells was evaluated by log 2 fold change expression data with non-treated but infected Caco2 cells as control. Green represents BioMos treatment and purple represents HMO treatment.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Expressing, Incubation, Infection, Control

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: BioMos addition rescues invasion phenotype of S. Typhimurium LT2 glycosyl hydrolase knockouts. Single gene knockouts of an amylase encoding gene (malS) and sialidase encoding gene (nanH) decreased the ability of S. Typhimurium LT2 to both adhere to and invade Caco2 cells. Pretreatment of the Caco2 cells with 1% BioMos for 60 min restored the adhesion and invasion abilities of both the malS and nanH knockouts as compared to a wild-type control. S. Typhimurium was co-incubated with the Caco2 cells for 60 min and invasion and adhesion was measured with a gentamicin protection assay. * p < 0.05, ** p < 0.02.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Control, Incubation

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: Regulation of gene expression and metabolism of Caco2 cells in response to prebiotic pretreatment. (A) HMO and BioMos treatment of Caco2 cells resulted in 3,745 shared significantly expressed genes, 191 additional unique genes in HMO treated cells and 3,533 in BioMos treated cells. (B,C) IPA was used to evaluated the expression patterns of the most significant (−log10 P > 1.3) metabolic pathways in both prebiotic treatments. Bars represent the percentage of each pathway differentially expressed in each condition, red sections being upregulation, blue downregulation, and white as the percentage of genes found to be unsignificant in the treatment for that pathway. Right hand side numbers represent the total genes in each pathway. (D–F) The metabolic profiles of prebiotic treated Caco2 cells were analyzed using Metabolite Set Enrichment Analysis (MSEA) in MetaboAnalyst. Time 0 with no prebiotic (D) , HMO (E) , or BioMos (F) was used as the control comparison for the same condition at Time 60. MSEA graph display the enriched pathways at Time 60 for each prebiotic condition, with the dot size representing the portion of the pathway enriched in the dataset and color as p -value.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Gene Expression, Expressing, Control, Comparison

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: Regulation of gene expression in S. Typhimurium 14028 ss in response to addition to Caco2 cells with prebiotic pretreatment. All expression data was collected after 60 min of co-incubation with Caco2 cells, with or without prebiotic treatment and in four biological replicates. (A) Enriched metabolic pathway in S. Typhimurium added to Caco2 cells pretreated with either BioMos (orange) or HMO (purple). Enrichment determined using BioCyc with a cut of –log10 P > 1.3 for significance and S. Typhimurium added to Caco2 cells and no prebiotic treatment as control condition. (B) Broad evaluation of S. Typhimurium gene expression in response to either prebiotic condition using log2 fold change data, significance cut off of –log10 P > 1.3 and Euclidean distance clustering for genes. (C) 2,107 genes were significantly expressed in both the HMO and BioMos treated S. Typhimurium, while an additional 2,713 unique genes were significantly expressed in the HMO condition and 0 were uniquely expressed in BioMos S. Typhimurium.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Gene Expression, Expressing, Incubation, Control

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: Effect of prebiotic treatment on three major energy-producing metabolic pathways across both infected Caco2 cells and S. Typhimurium 14028 ss. Expression of genes related to the TCA cycle, glycolysis and oxidative phosphorylation in response to prebiotic treatment was evaluated for both Caco2 cells (top) and S. Typhimurium (bottom). Caco2 expression of these metabolic respiration-related genes was evaluated using S. Typhimurium infected Caco2 cells without any prebiotic treatment added as the control for both HMO and BioMos. S. Typhimurium expression was determined using S. Typhimurium added to Caco2 cells without any prebiotics as control. All heatmaps used Euclidean distance clustering, log2 fold change expression data, and only significant genes with a cutoff of −log10 P > 1.3. Underlying data for the genetic regulation of the TCA cycle (right) was mapped using IPA for the Caco2 cells and BioCyc for S. Typhimurium. All genes represented in orange and blue in the diagrams were significantly expressed compared to the control with a cutoff of –log10 P > 1.3.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Infection, Expressing, Phospho-proteomics, Control

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: The pentose phosphate pathway is differentially regulated by prebiotic treatment in both Caco2 cells and S. Typhimurium 14028 ss. Genetic regulation of the pentose phosphate pathway was evaluated in both S. Tyhpimurium (left within each panel) and Caco2 cells (right within each panel) across HMO addition (left) and BioMos addition (right). Expression is represented as log2 fold change with no prebiotic add matched host/pathogen cells as control and all regulated genes were significant at –log10 P > 1.3. Increased expression is represented by yellow to red and decreased expression by blue to purple. Black text and teal arrows indicate those genes were non-significant or not annotated in the dataset.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Expressing, Control

Journal: Frontiers in Microbiology

Article Title: Cellular respiration and amino acid metabolism is altered by dietary oligosaccharides in Salmonella with epithelial cell association

doi: 10.3389/fmicb.2025.1672770

Figure Lengend Snippet: Prebiotic treatment of Caco2 cells challenged with S. Typhimurium 14028 ss alters oxidative phosphorylation and expression patterns in the electron transport chain. The canonical oxidative phosphorylation pathway in IPA was overlayed with expression data from Caco2 cells treated with either HMO (top) or BioMos (bottom) and exposed to S. Typhimurium for 60 min. Red shading in the complexes indicates induction of genes responsible for that complex’s activity and teal shading indicates repression of that complex. Orange shading on the products indicates a predicted increase in concentration due to the expression pattern.

Article Snippet: Human colonic carcinoma (Caco2) cell lines were obtained from ATCC (HTB-37) and grown as described previously ( ).

Techniques: Phospho-proteomics, Expressing, Activity Assay, Concentration Assay