Flowchart depicting conduct of baseline and two-step tuberculin testing.

<p>Flowchart depicting conduct of baseline and two-step tuberculin testing.</p

The relationship between the cytokine-producing CD4+CD45RO+ T cells and the QuantiFERON Gold In-Tube (QFT) response at baseline.

<p>The relationship between the absolute frequencies of (<b>A</b>) all IFNγ-producing and (<b>B</b>) polyfunctional (IFNγ, IL2 and TNFα) PPD-specific CD4+CD45RO+ T cells and the magnitude of the QFT response are shown. The relationship between the absolute frequencies of cytokine-producing T-cells in response to SEB is shown for comparison (<b>C and D</b>). Statistical analyses were performed by Spearman’s correlation coefficient (rho).</p

QuantiFERON Gold In-Tube (QFT) and Tuberculin Skin Test (TST) responses.

<p>(<b>A</b>) The magnitude of the QFT response and (<b>B</b>) the TST response in QFT consistent positives (n = 21), QFT reverters (n = 21) and QFT negative controls (n = 10) at baseline (BL) and after 1 year. Unpaired and paired analyses was performed with Mann-Whitney U test and Wilcoxon Signed rank test where appropriate. Horizontal lines represent median values.</p

Qualitative and quantitative analysis of PPD-specific CD4+CD45RO+ T-cell responses in QFT consistent positives and QFT reverters.

<p>(<b>A</b>) The relative frequencies of the PPD-specific CD4+CD45RO+ T-cell subsets in QFT consistent positives (red symbols) and QFT reverters (blue symbols) at baseline (filled) (n = 16, n = 7) and after 1 year (open) (n = 6, n = 6). The subsets are defined on the basis of IFNγ, TNFα and/or IL-2 production. Since relative frequencies are normalized data, un-paired T-test with Welch’s correction was applied for statistical analyses and horizontal lines represent mean values. (<b>B</b>) The absolute frequencies of PPD-specific CD4+CD45RO+ T- in QFT consistent positives (red symbols) and QFT reverters (blue symbols) at baseline (filled) (n = 16, n = 7) and after 1 year (open) (n = 6, n = 6). As the data are not normally distributed, statistical analyses were performed by the Mann-Whitney U test and horizontal lines represent median values. (<b>C</b>) Flowcytometric plots of the functional profile of PPD-specific CD4+CD45RO+ T-cells compared with negative (PBS) and positive (SEB) control in one representative QFT consistent positive subject are shown.</p

Dot-plot graph depicting genes and proteins that are differentially expressed between the three clinical groups: TB disease, NTM present and controls (defined as MTB, NTM uninfected children and negative for TST and QFTGIT).

<p>(A) The median with inter quartile range relative gene expression (log 2 transformed) of genes from peripheral blood. (B) The median concentration (pg/mL) of cytokines in the QFTGIT supernatants after stimulation of whole blood with <i>M. tuberculosis</i> antigens. p-value<0.05 (*), <0.01 (**), <0.001 (***) were considered to be significant.</p

Flow chart of patients recruited to the study.

<p>¹Referral based on cough >2weeks, FTT, history of contact with a TB case and a TST ≥10 mm at study closure ²Abnormal CXR; MTB positive by the Hain MTB test or Roche test. ³Abnormal CXR; culture negative for MTB. ⁴Normal CXR; culture positive for NTM and ⁵Normal CXR; culture negative for MTB and NTM. Abbreviations: FTT – failure to thrive; CVW- case verification ward; CXR – chest X-ray; TST - tuberculin skin test; QFTGIT – QuantiFERON Gold In-tube test; dcRT-MLPA – dual colour reverse transcriptase – multiplex ligation dependent probe amplification; MTB – <i>Mycobacterium tuberculosis</i>; NTM – non-tuberculous mycobacteria.</p

Dot-plot graph depicting genes and proteins that are differentially expressed between children with NTM present and children that were culture negative for MTB and NTM.

<p>(A) The median with inter quartile range relative gene expression (log 2 transformed) of genes from peripheral blood. (B) The median concentration (pg/mL) of cytokines in the QFTGIT supernatants of whole blood without stimulation. p-value<0.05 (*) was considered to be significant.</p

Day 1 and Day 2 induced sputum (IS) and gastric aspirate (GA) culture positive results among 210 study participants.

<p>*Mixed NTM species were isolated from day 1 and day 2 IS and GA samples.</p><p>Day 1 and Day 2 induced sputum (IS) and gastric aspirate (GA) culture positive results among 210 study participants.</p

Baseline characteristics of 210 children.

<p>Abbreviations: CXR-Chest X-ray; TST- tuberculin skin test; QFTGIT- QuantiFERON In-tube TB Gold test; IS-Induced sputum; GA-gastric aspirates; MTB- <i>Mycobacterium tuberculosis</i>; NTM-non-tuberculous mycobacteria.</p><p>Baseline characteristics of 210 children.</p

Dot-plot graph depicting genes and proteins that are differentially expressed between the three clinical groups: TB disease, NTM present and controls.

<p>(A) The median with inter quartile range relative gene expression (log 2 transformed) of genes from peripheral blood. (B) The median concentration (pg/mL) of cytokines in the QFTGIT supernatants of whole blood without stimulation. (C) The median concentration (pg/mL) of cytokines in the QFTGIT supernatants after stimulation of whole blood with <i>M. tuberculosis</i> antigens. p-value<0.05 (*), <0.01 (**), <0.001 (***) were considered to be significant; NS - not significant.</p