Genes identified by more than 1 independent study.

<p>Genes identified by more than 1 independent global genome-wide gene expression analysis. Manuscript numbers refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone-0073230-t002" target="_blank">Table 2</a>. Classification into modules, functional groups according to Ingenuity and GSEA was performed according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone-0073230-t003" target="_blank">Tables 3</a> & <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone-0073230-t004" target="_blank">4</a> and identical to Genes identified by more than 1 independent global genome-wide gene expression analysis. Manuscript numbers refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone-0073230-t002" target="_blank">Table 2</a>. Classification into modules, functional groups according to Ingenuity and GSEA was performed according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone-0073230-t003" target="_blank">Tables 3</a> & <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone-0073230-t004" target="_blank">4</a> and identical to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0073230#pone.0073230.s001" target="_blank">Table S1</a>.</p

Functional classification of individual genes identified by gene expression analysis on TB patients.

<p>Categories have been based on combined output from Ingenuity and GSEA software modules and may include multiple canonical pathways or cell processes. Myeloid cells includes the following canonical pathways: role of macrophages, fibroblasts and endothelial cells in rheumatoid arthritis; Fcg Receptor mediated phagocytosis in macrophages and monocytes; role of pattern recognition receptors in recognition of bacteria and viruses; IL12 signaling and production in macrophages; Dendritic cell maturation; production of Nitrox Oxide and Reactive Oxygen Species in Macrophages; Toll like receptor signaling. T cells includes: T cell receptor signaling; CD28 signaling in T helper cells; iCOS-iCOSL signaling in T helper cells. B cells includes: B cell receptor signaling; PI3K signaling in B lymphocytes. Interferon related pathways include: Interferon signaling, role of jak1, jak2 and tyk2 in interferon signaling, role of PKR in interferon induction and antiviral response. Inflammation includes: IL-8 signaling; NF-kB signaling; altered T cell and B cell signaling in Rheumatoid Arthritis; systemic lupus erythematosus signaling; chemokine signaling; IL-6 signaling. TREM1 includes specifically TREM1 signaling and mitochondrial dysfunction also only contains mitochondrial dysfunction. Finally, hematopoiesis includes: erythropoietin signaling; IL-3 signaling; FLT3 signaling in hematopoietic progenitor cells; prolactin signaling; HGF signaling.</p

Schematic representation of events during active TB Disease.

<p>Pathway and process based analysis suggests that these processes are key players in TB disease pathogenesis.</p

Information on studies included.

<p>Information on studies included.</p

Ingenuity pathway analysis of all genes identified by unbiased methods related to TB disease.

<p>All 409 biomarkers were analysed by integrated pathway analysis using Ingenuity and the most dominant network is depicted here. Signalling pathways were coloured according to functional classification into myeloid cells, T cells and B cells and type I interferon related genes.</p

TREM1 canonical pathway.

<p>Genes involved in TREM1 signaling canonical pathway according to Ingenuity Integrated Pathway analysis. Genes identified in our 409 gene total geneset are indicated in the right column. * TLR genes include TLR2,4,5,6,7,8 and were identified by multiple studies, all genes identified count in overlap with pathway count according to Ingenuity</p

Results from GSEA.

<p>Gene sets with an FDR <5% were included in this analysis. SIZE indicates the number of genes in both the gene set and the expression dataset. NES the primary result of the gene set enrichment analysis is the enrichment score (ES), which reflects the degree to which a gene set is overrepresented in a list of genes. Normalizing the enrichment score (NES) accounts for differences in gene set size and in correlations between gene sets and the expression dataset.</p

Factors that may affect TB Biomarkers.

<p>Factors that may affect TB Biomarkers.</p

Results of Ingenuity pathway analysis.

<p>All canonical pathways significantly associated with the dataset are depicted (p<0.001), after application of the following filter criteria: gene set comprises at least 50 genes, at least 10 genes from dataset are retrieved in gene set and at least 10% of genes from gene set are present in the data set of 409 genes.</p

Immunogenicity and protective efficacy of BCG + CT – MVA85A.

<p>Balb/c mice received BCG±CT <i>i.n.</i> followed by 1×10⁶ CFU MVA85A 10 weeks later. Lungs (<b>A</b>) and spleen (<b>B</b>) were taken at 10 (black circles), 11 (dark grey circles) and 14 (light grey circles) weeks post-BCG and cytokine-producing cells responding to an Ag85A peptide pool quantified using ICS. Responses from animals receiving BCG – MVA85A (closed circles) were compared with those receiving BCG + CT followed by MVA85A (open circles). Statistical analysis was performed using a Mann Whitney test. n = 10, five each from two experiments. (<b>C</b>) Balb/c mice were vaccinated as above. Control groups included unvaccinated and BCG <i>i.d.</i> A group receiving BCG <i>i.n.</i> was included to compare BCG – MVA85A <i>i.n.</i> to BCG <i>i.n</i>. Animals were exposed to ∼100 CFU <i>M.tb</i> via aerosol four weeks post-MVA85A. Four weeks post-challenge, lungs and spleen were homogenised and plated for CFU quantitation. (<b>D</b>) Balb/c mice were vaccinated and challenged as described above. Groups receiving BCG – MVA85A and BCG + CT – MVA85A received an anti-IL-17 blocking antibody (MAB421; R&D Systems) administered <i>i.p.</i> every three days post-challenge. One group receiving BCG – MVA85A received an IgG2a isotype control antibody (MAB006; R&D Systems) on the same regimen. Mice were culled four weeks post-challenge and lung CFU quantitated as described above. Statistical analysis was performed using a one way ANOVA and post-hoc tests on the vaccinated groups (n = 8–16).</p