Fig 1.
Workflow of gene co-expression network analyses for STEC strains and Caco-2 cells.
(A) Network analysis for STEC strains. Two comparative analyses were done to investigate if enterocyte soluble mediators modify global gene expression after bacterial growth in Caco-2-conditioned (C) medium for 3h: i) EH41 in C medium X EH41 in F medium; ii) Ec472/01 in C medium X Ec472/01 in F medium. Another comparative analysis was done to assess global gene expression differences between EH41 and Ec472/01 in C medium. (B) Network analyses for Caco-2 cells after 3h of interaction with EH41 or Ec472/01. Two comparative analyses were done: Caco-2 exposed to EH41 X Caco-2 control and Caco-2 exposed to Ec472/01 X Caco-2 control.
Table 1.
Differentially expressed (DE) and exclusively expressed (EE) valid transcripts obtained for each of the comparison groups.
Fig 2.
Differential gene expression profiles for STEC strains.
Venn diagram analysis for DE and EE genes obtained from two comparisons: EH41 in C medium X EH41 in F medium or Ec472/01 in C medium X Ec472/01 in F medium.
Fig 3.
Functional profile analyses of DE and EE transcripts obtained from the comparison of EH41 X Ec472/01 in C medium.
Pie charts show: (A) the DE transcripts set; (B) the EE transcripts in EH41; (C) the EE transcripts in Ec472/01. Functional categories are identified by roman numerals as follows: I, metabolic process; II, chaperone; III, fimbrin/ outer membrane protein; IV, ion transport/ protein transport; V, transcription; VI, qin prophage/ prophage; VII, acid resistance; VIII, transposase; IX, putative protein. The number of genes belonging to a particular functional category is indicated between parentheses in each slice. Transcripts described as hypothetical or unknown proteins are not represented in this figure.
Fig 4.
DE and EE gene co-expression networks (GCNs) for STEC strains.
GCNs for EH41 and Ec472/01 are depicted in (A) and (B) respectively. The most relevant hubs (high number of gene-gene links) are graphically represented by the proportionally larger nodes. Positive or negative Pearson’s coefficients—indicating positive or inverse covariation between gene-pairs—are represented by gray or red lines respectively. Nodes in red or green indicate hyper or hypo expressed genes, respectively. Circle or diamond nodes indicate DE or EE genes respectively. Node borders in blue indicate hub genes; node borders in yellow indicate common hubs between the two GCNs.
Table 2.
Gene co-expression network characteristics for each group.
Table 3.
Hubs in EH41 and Ec472/01 networks.
Fig 5.
Hierarchical structure of the EH41 network.
Node hierarchical arrangement represents the main direction within a network (A). In (B) and (C) the EE transcriptional module is displayed in detail. Links in red represent the first node connections, centered in dicA (B) or in dicC (C). Nodes in red or green indicate hyper or hypo expressed genes, respectively. Circle or diamond nodes indicate DE or EE genes respectively. Node borders in blue indicate hub genes; node borders in yellow indicate common hubs between EH41 and Ec472/02 GCNs.
Table 4.
PCR gene detection in a panel of STEC O113:H21 strains.
Fig 6.
DE gene co-expression network (GCN) for Caco-2 cells after interaction with STEC strains.
GCNs for Caco-2 interacting with EH41 or with Ec472/01 are shown in (A) and (B) respectively. Hubs are graphically represented by the proportionally larger nodes. Positive or negative Pearson’s coefficients are indicated by gray or red lines respectively. Nodes in red or green indicate hyper or hypo expressed genes, respectively. Blue node border indicates a hub gene; yellow node border indicates a common hub between the two GCNs.
Fig 7.
DE gene co-expression network (GCN) for Caco-2 cells control group.
Caco2-control network was constructed considering all DE genes obtained from two comparisons: Caco-2 with EH41 X Caco-2 control and Caco-2 with Ec472/01 X Caco-2 control. Hubs (square nodes) are graphically represented by the proportionally larger nodes. Positive or negative Pearson’s coefficient is indicated by gray or red lines respectively. Nodes in blue, pink or purple indicate, respectively: DE genes present only in Caco-2 interacting with EH41, or with Ec472/01 or common for the two groups. Node borders in green or red indicate hypo or hyper expressed genes in the control group.
Table 5.
Hubs in Caco-2 cells with EH41 or with Ec472/01 and control networks.
Fig 8.
SEM visualization of Caco-2 cells after 3h of interaction with STEC strains.
The images depict the Caco-2 cells control (A) and cells interacting with EH41 (B) or Ec472/01 (C).