Treg and Tact are distinguished by a combination of several markers.

<p>(<b>A</b>) PBMC were electronically gated on lymphocyte population (left panel) followed by gate on total CD4⁺ cells (middle panel). CD4⁺ cells were analysed for the expression of CD25 and CD127 (right panel). (<b>A–C</b>) CD25⁺FOXP3⁺ cells were identified as Treg, whereas CD25⁺FOXP3⁻ counted as Tact. A dot plot for one individual is shown for group 1 (<b>B</b>) and group 2 (<b>C</b>). Gating on Treg and Tact subsets was verified by further analysis of level of expression of CD25 (<b>B–C</b>, middle panel) and CD127 (<b>B–C</b>, right panel) expressed on Treg (red) and Tact (green). Gray line - isotype control.</p

Influence of host attributes on T cell subsets.

<p>Results from the multivariate analysis of variance determining the effect of host sex and age on percentages of activated (Tact) and regulatory (Treg) cells. df  =  degrees of freedom and tests significant at p<0.05 are highlighted in bold.</p

The association of Treg and Tact proportions with infection intensity are presented partitioned by age group.

<p>The variation in infection intensity ((log₁₀ (egg count+1) transformed) was analysed for the potential confounding effects sex and village using ANOVA and calculated residuals are plotted against the proportion of Treg (<b>A, B</b>) or Tact (<b>C, D</b>; arcsin√ transformed) after dividing into two age groups as indicated. Obtained residuals were used for further used in a regression analysis whose p- and β- values are given.</p

Study population was divided in two age groups.

<p>The 8–13 year age group (N = 30) infection intensity is rising and peaking, whereas in the 14+ year age group (N = 19) infection intensity is lower. Dots represent individual values of infection intensity to illustrate distribution within the age groups and gray line indicates means. Means were compared by Student's t-Test and p-value is shown.</p

The proportion of Treg does not differ between age group regardless of markers employed.

<p>Dot plots show the fraction of Treg identified by a combination of different markers: (<b>A</b>) CD25⁺FOXP3⁺ cells, (<b>B</b>) FOXP3⁺CD127^dim cells and (<b>C</b>) CD25^high cells as percentage of CD4⁺ T cells in the different age groups. In (<b>D</b>) percentages of CD25⁺FOXP3^neg Tact are presented.</p

Proportions of ILC2s identified by CD127+CD294+CD161+ are lower in <i>S</i>. <i>haematobium</i> egg positive young children.

<p>(A) The participants were compared between <i>S</i>. <i>haematobium</i> egg negative (ve-, N = 36) and egg positive (ve+, N = 36) individuals by a non-parametric Mann-Whitney <i>U</i> test. (B) The cohort was divided into three age groups (age in years) and egg positive (ve+, open symbols) were compared to egg negative (ve-, closed symbols) people. Grey lines indicate median and the interquartile range. ILC2s proportions were analysed using a Kruskal-Wallis test followed by a multiple comparison of egg positive <i>versus</i> egg negative children.</p

Comparison of plasma TSLP and IL-33 levels in relation to <i>S</i>. <i>haematobium</i> infection.

<p>Plasma levels of (A) TSLP and (B) IL-33 were measured by ELISA and data were divided into three age group and <i>S</i>. <i>haematobium</i> egg negative (ve-, closed symbols) compared to egg positive (ve+, open symbols) individuals. Grey lines indicate median and the interquartile range. Levels were compared using a Kruskal-Wallis test followed by a multiple comparison of egg positive <i>versus</i> egg negative children. Plasma levels of (C) TSLP and (D) IL-33 from 12 individuals (as described for <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003627#pntd.0003627.g003" target="_blank">Fig. 3</a>) were compared pre- and 6 week’s post-treatment. P-values are obtained from a Wilcoxon signed-rank test and positive/negative/unchanged ranks are shown in the lower-right corner of panel (C) and (D).</p

<i>In vitro</i> generation of effector T cells (E) and co-culture with LD-BCG infected MDM (M) at different E/M ratios.

<p>(A) Workflow depiction for the generation of effector T cells, LD-BCG infected MDM, E/M co-culture, flow cytometry analyses of infected MDM and effector T cells. (B) Analyses of MDM infected with LD-BCG after co-culture with effector T cells stimulated with <i>Staphylococcus</i> Enterotoxin B (SEB), <i>M</i>. <i>tuberculosis</i> Purified Protein Derivative (PPD), and without stimulation (w/o). Different E/M ratios are shown on the x-axes. Proportions of infected MDM with live (grey) or dead LD-BCG (open) are shown as stacked boxes (left graph). Absolute numbers of infected MDM are shown as symbols (middle graph) and numbers of MDM infected with live LD-BCG are shown as boxes (right graph). The dotted line in the middle graph indicates MDM numbers infected with LD-BCG w/o effector T-cell co-culture. Median with range of triplicates from a representative experiment are depicted.</p

Characterisation of human ILC2 by flow cytometry.

<p>(A) Human PBMC were gated on live cells using forward (FSC) and side (SSC) scatter. Gated live cells were analysed using the area against height of the FSC to discriminate singlet’s from duplicates. Singlet’s were subsequently analysed by a lineage cocktail (Lin) and CD45 to gate on Lin-CD45+ cells (as indicated in the box) (B). Lin-CD45+ cells were gated on CD127+ cells, which were analysed for CD294 and CD161 (C). CD127+CD294+CD161+ were analysed for CD117 (grey histogram) versus isotype control (open histogram) (D). Flow charts as presented are from the PBMC analysis of a 9 year old female who was negative for <i>S</i>. <i>haematobium</i> eggs.</p

Non-metric multidimensional scaling (NMDS) ordination in 2-dimensional configurations by sex, age-group and <i>S. haematobium</i> infection status determined using parasitological (A) and serological diagnostic techniques (B).

<p>Subgroup centres are represented by the bigger closed (●), or open (○) points, and the distance between these centres is proportional to the level of dissimilarities between subgroups.</p