R script written for some of the analyses

This is an R script that was written for some of the analysis, namely: 1) hierarchical f-statistics 2) tests for inbreeding with multilocus heterozygosity and 3) tests for family structure with relatedness estimate

Microsatellite data from parasites

This data consists genotypes obtained from Schistosoma mansoni parasites that were type at nine microsatellite markers. Each locus (e.g. L46951) is coded by 6 digits (e.g. 169175). Each allele is coded by 3 digits (e.g. 169)

Human host data

Village: location where human host lived. Host: population number that was assigned to the human host. HostIdentifier: code that is linked with that host. SamplingTime: time when parasites were sampled (e.g. Aug-09 or 09-Aug means that parasites were sampled from this host in August 2009. Age: age of the human host. EPG: eggs per gram of feces, i.e. a measure of infection intensity. CoInfectionWithShaematobium: 1 = when human host was coinfected with Schistosoma haematobium, 0 = when human host was NOT coinfected

Cytokine Responses to <i>Schistosoma mansoni</i> and <i>Schistosoma haematobium</i> in Relation to Infection in a Co-endemic Focus in Northern Senegal

<div><p>Background</p><p>In Africa, many areas are co-endemic for the two major <i>Schistosoma</i> species, <i>S. mansoni</i> and <i>S. haematobium</i>. Epidemiological studies have suggested that host immunological factors may play an important role in co-endemic areas. As yet, little is known about differences in host immune responses and possible immunological interactions between <i>S. mansoni</i> and <i>S. haematobium</i> in humans. The aim of this study was to analyze host cytokine responses to antigens from either species in a population from a co-endemic focus, and relate these to <i>S. mansoni</i> and <i>S. haematobium</i> infection.</p><p>Methodology</p><p>Whole blood cytokine responses were investigated in a population in the north of Senegal (n = 200). Blood was stimulated for 72 h with schistosomal egg and adult worm antigens of either <i>Schistosoma</i> species. IL-10, IL-5, IFN-γ, TNF-α, and IL-2 production was determined in culture supernatants. A multivariate (i.e. multi-response) approach was used to allow a joint analysis of all cytokines in relation to <i>Schistosoma</i> infection.</p><p>Principal Findings</p><p><i>Schistosoma haematobium</i> egg and worm antigens induced higher cytokine production, suggesting that <i>S. haematobium</i> may be more immunogenic than <i>S. mansoni</i>. However, both infections were strongly associated with similar, modified Th2 cytokine profiles.</p><p>Conclusions/Significance</p><p>This study is the first to compare <i>S. mansoni</i> and <i>S. haematobium</i> cytokine responses in one population residing in a co-endemic area. These findings are in line with previous epidemiological studies that also suggested <i>S. haematobium</i> egg and worm stages to be more immunogenic than those of <i>S. mansoni</i>.</p></div

Spatial distribution of <i>S. mansoni</i>- and <i>S. haematobium</i>-specific morbidity.

<p>Black circles and crosses indicate households that were included and excluded from the analysis, respectively. Continuous colored circles are statistically significant clusters (<i>p</i><0.05) and dotted circles are borderline significant (<i>p</i><0.06). Roman numerals indicate water contact sites. <b>Panel A</b> depicts the unadjusted clusters for the prevalence of morbidity. Both <i>S. mansoni</i>-specific hepatic fibrosis (pink dotted circle; RR = 1.9) and <i>S. haematobium</i>-specific urinary tract morbidity clusters (green dotted circle; RR = 1.2) were borderline significant (<i>p</i> = 0.054 and <i>p</i> = 0.053, respectively). The prevalence of hepatic fibrosis was 41% (59/145) in- and 21% (31/146) outside, and that of urinary tract morbidity was 89% (117/131) in- and 73% (116/160) outside the cluster. Gender- and age-adjusted analysis revealed no (borderline) significant clusters for the prevalence of morbidity. <b>Panel B</b> depicts the clusters of morbidity by severity. The risk of severe hepatic fibrosis was elevated in the circle with a RR of 0.3 for liver image pattern A, 1.3 for B, 1.4 for C, 2.7 for D and 4.3 for E, and the only person with pattern F lived here (<i>p</i> = 0.001; unadjusted ordinal model). The gender- and age-adjusted cluster for patterns D–F (as opposed to A–C) in adults constituted of the households indicated in pink (<i>p</i> = 0.031; Bernoulli model).</p

Variation in <i>Schistosoma</i> antigen-induced cytokine responses in relation to <i>Schistosoma</i> infection intensity.

<p>Each three-dimensional (3D) nMDS ordination is represented in two 2D planes (<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003080#pntd.0003080.s001" target="_blank">Supporting Information S1</a>). Left and right panels represent the 1^st and 2^nd, and 2^nd and 3^rd dimensions, respectively. <b>Panels A</b> and <b>B</b> show the <i>S. mansoni</i> egg antigen (SEAm)-induced cytokine profile, <b>Panels C</b> and <b>D</b> that of <i>S. haematobium</i> SEA(h), <b>Panels E</b> and <b>F</b> that of <i>S. mansoni</i> adult worm antigens (AWAm), and <b>Panels G</b> and <b>H</b> show <i>S. haematobium</i> AWA(h)-induced cytokine profiles. Green dots represent individuals. Distances between dots approximate the rank order of dissimilarities in cytokine profiles between the respective individuals with stress values (i.e. discrepancies) of 0.051 for SEAm, 0.041 for SEAh, 0.058 for AWAm, and 0.061 for AWAh. Red arrows indicate linear gradients of normalized net cytokine responses on which the nMDS is based. Green dot sizes are proportional to individual values of normalized infection intensity of <i>S. mansoni</i> (for simplicity dots were only labelled with <i>S. mansoni</i> (not <i>S. haematobium</i>) infection intensity). Black arrows indicate linear gradients of post hoc fitted normalized infection intensity of <i>S. mansoni</i> (‘Sm’) and <i>S. haematobium</i> (‘Sh’). The length of the arrows is proportional to the goodness of fit onto the cytokine profile within one 2D plane, but lengths cannot be compared between cytokine and infection intensity arrows. Arrows are only depicted if their fit was significant at the level of <i>p</i> = 0.05 in 3D ordinations (see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003080#pntd-0003080-t004" target="_blank">Table 4</a>), as well as in the respective 2D planes. In Panel H, the arrows of IL-5 response and <i>S. mansoni</i> infection intensity are overlapping and their labels are therefore illegible. ^aThe biological a posteriori interpretation of nMDS1 (left x-axis) and nMDS2 (y-axis) were added between brackets on the axis labels, but nMDS3 (right x-axis) could not be interpreted.</p

Spatial distribution of <i>S. mansoni</i> and <i>S. haematobium</i> infection densities.

<p>Black circles and crosses indicate households that were included in and excluded from the analysis, respectively. Roman numerals indicate water contact sites. <b>Panel A</b> depicts the unadjusted clusters (<i>p</i> = 0.001 for both <i>S. mansoni</i> and <i>S. haematobium</i>). The geometric mean (GM) <i>S. mansoni</i> infection density was 33 epg for those living in inside the northern <i>S. mansoni</i> cluster (n = 285) compared to12 epg in the rest of the community (n = 314). The GM <i>S. haematobium</i> infection density was 4.7 ep10ml inside the southern <i>S. haematobium</i> cluster (n = 34) and 0.7 ep10ml outside (n = 565). <b>Panel B</b> depicts the gender- and age-adjusted clusters (<i>p</i> = 0.002 for <i>S. mansoni</i> (north), and <i>p</i> = 0.023 for <i>S. haematobium</i> (south).</p

Distribution of <i>Schistosoma</i> infection in the study population.

<p>Distribution of <i>Schistosoma</i> infection in the study population.</p

Levels of <i>Schistosoma</i>-induced cytokine responses in 72 h whole blood cultures (n = 200).

<p>Blood samples from one individual were divided into five and stimulated with <i>Schistosoma</i> antigens (SEAm, SEAh, AWAm, or AWAh), and with medium only (negative control; see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003080#s2" target="_blank">Materials and Methods</a>).</p>a<p>Crude cytokine levels are reported. IQR: Interquartile range (Tukey's hinges).</p>b<p>Wilcoxon Signed Rank test comparing <i>S. mansoni</i>- and <i>S. haematobium</i>-induced cytokine levels within individuals (either for SEA or AWA).</p>c<p><i>Schistosoma</i> egg antigen.</p>d<p>Adult worm antigen.</p

Characteristics of the 6 datasets used for the gender- and age-adjusted spatial analyses.

<p>^a Total study population in the first columns and numbers of cases in subsequent columns.</p