Total IgG prevalence against <i>La</i>PSA-38S in different endemic areas of leishmaniasis.

<p>The cut-off value of reactivity for <i>La</i>PSA-38S antigen was calculated as the mean plus 3 SD of the OD values observed in naive controls from each country. These cut-off values allowed us to distinguish between positive and negative sera and consequently to estimate performance parameters of each ELISA test.</p

Total Soluble <i>Leishmania</i> antigen (TSLA) specific IFN-γ, granzyme B, TNF-α responses.

<p>IFN-γ (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092708#pone-0092708-g002" target="_blank">Fig. 2a</a>), granzyme B (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092708#pone-0092708-g002" target="_blank">Fig. 2b</a>), were detected and quantified from culture supernatants of PBMC exposed for 120h to local TSLA (10 µg/ml) and TSLA Ldd8 (10 µg/ml), by Cytokine Beads Array test (CBA) using Flow cytometry. Statistically significant differences between stimulated and non stimulated cultures and between groups (p≤0.03) are showed.</p

SDS-PAGE and silver staining of purified recombinant <i>La</i>-PSA38S protein expressed in <i>L. tarentolae</i>.

<p>Purified <i>La</i>-PSA38S was separated by electrophoresis in a 12% SDS-PAGE and silver stained under reducing (R) and non reducing (NR) conditions. The arrow indicates the position of <i>La</i>-PSA-38S (45 kDa) and Molecular Weight markers were marked in kDa.</p

<i>La</i>PSA-38S specific IFN-γ, granzyme B, TNF-α and IL-10 responses.

<p>IFN-γ (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092708#pone-0092708-g003" target="_blank">Fig. 3a</a>), granzyme B (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092708#pone-0092708-g003" target="_blank">Fig. 3b</a>), TNF-α (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092708#pone-0092708-g003" target="_blank">Fig. 3c</a>) and IL-10 (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092708#pone-0092708-g003" target="_blank">Fig. 3d</a>) were detected and quantified from culture supernatants of PBMC exposed for 120 h to SLA (10 µg/ml) or <i>La</i>PSA-38S (10 µg/ml) using Cytokine Beads Array test (CBA). Data were analyzed by Flow cytometry. PHA (10 µg/ml) was used for all cultures as positive control (data not shown). Statistically significant differences between stimulated and non stimulated cultures (p≤0.003) and between groups (p≤0.01) are showed.</p

Study population.

<p>The recruitment and sampling collection (186 donors) of different groups (patients, cured, immunes without clinical symptoms and naives) were performed in endemic and non-endemic areas in each country.</p><p>*CVLd: Cured Visceral Leishmaniasis due to <i>L. donovani</i> (India), aVLd: active Visceral Leishmaniasis due to <i>L. donovani</i> (India), HLR-I: Healthy Low Responders from India, CCLb: Cured Cutaneous Leishmaniasis due to <i>L. brasiliensis</i> (Peru), aCLb: active Cutaneous Leishmaniasis due to <i>L. brasiliensis</i> (Peru), HLR-P: Healthy Low Responders from Peru, CCLm: Cured Cutaneous Leishmaniasis due to <i>L. major</i> (Tunisia), HHR-Lm: Healthy High Responders living in an endemic area for <i>L. major</i> (Tunisia), HHR-LiT: Healthy High Responders living in an endemic area for <i>L. infantum</i> (Tunisia), aVLiT: active Visceral Leishmaniasis due to <i>L. infantum</i> (Tunisia), H-T: Healthy Low Responders from Tunisia, HHR-LiF: Healthy High Responders living in an endemic area for <i>L. infantum</i> (France), HLR-F: Healthy Low Responders from France, HHR-LiS: Healthy High Responders living in an endemic area for <i>L. infantum</i> (Spain), aVLiS: active Visceral Leishmaniasis due to <i>L. infantum</i> (Spain), HLR-S: Healthy Low Responders from Spain.</p