Glycans present on glycoproteins and glycolipids of the major human parasite Schistosoma mansoni induce innate as well as adaptive immune responses in the host. To be able to study the molecular characteristics of schistosome infections it is therefore required to determine the expression profiles of glycans and antigenic glycan-motifs during a range of critical stages of the complex schistosome lifecycle. We performed a longitudinal profiling study covering schistosome glycosylation throughout worm- and egg-development using a mass spectrometry-based glycomics approach. Our study revealed that during worm development N-glycans with Galβ1–4(Fucα1–3)GlcNAc (LeX) and core-xylose motifs were rapidly lost after cercariae to schistosomula transformation, whereas GalNAcβ1–4GlcNAc (LDN)-motifs gradually became abundant and predominated in adult worms. LeX-motifs were present on glycolipids up to 2 weeks of schistosomula development, whereas glycolipids with mono- and multifucosylated LDN-motifs remained present up to the adult worm stage. In contrast, expression of complex O-glycans diminished to undetectable levels within days after transformation. During egg development, a rich diversity of N-glycans with fucosylated motifs was expressed, but with α3-core fucose and a high degree of multifucosylated antennae only in mature eggs and miracidia. N-glycan antennae were exclusively LDN-based in miracidia. O-glycans in the mature eggs were also diverse and contained LeX- and multifucosylated LDN, but none of these were associated with miracidia in which we detected only the Galβ1–3(Galβ1–6)GalNAc core glycan. Immature eggs also exhibited short O-glycan core structures only, suggesting that complex fucosylated O-glycans of schistosome eggs are derived primarily from glycoproteins produced by the subshell envelope in the developed egg. Lipid glycans with multifucosylated GlcNAc repeats were present throughout egg development, but with the longer highly fucosylated stretches enriched in mature eggs and miracidia. This global analysis of the developing schistosome's glycome provides new insights into how stage-specifically expressed glycans may contribute to different aspects of schistosome-host interactions.

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The synthesis of a collection of 33 xylosylated and core-fucosylated N-glycans found only in nonmammalian organisms such as plants and parasitic helminths has been achieved by employing a highly convergent chemo-enzymatic approach. The influence of these core modifications on the interaction with plant lectins, with the human lectin DC-SIGN (Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Nonintegrin), and with serum antibodies from schistosome-infected individuals was studied. Core xylosylation markedly reduced or completely abolished binding to several mannose-binding plant lectins and to DC-SIGN, a C-type lectin receptor present on antigen presenting cells. Employing the synthetic collection of core-fucosylated and core-xylosylated N-glycans in the context of a larger glycan array including structures lacking these core modifications, we were able to dissect core xylose and core fucose specific antiglycan antibody responses in S. mansoni infection sera, and we observed clear and immunologically relevant differences between children and adult groups infected with this parasite. The work presented here suggests that, quite similar to bisecting N-acetylglucosamine, core xylose distorts the conformation of the unsubstituted glycan, with important implications for the immunogenicity and protein binding properties of complex N-glycans.

BackgroundSchistosomiasis (bilharzia) is a chronic and potentially deadly parasitic disease that affects millions of people in (sub)tropical areas. An important partial immunity to Schistosoma infections does develop in disease endemic areas, but this takes many years of exposure and maturation of the immune system. Therefore, children are far more susceptible to re-infection after treatment than older children and adults. This age-dependent immunity or susceptibility to re-infection has been shown to be associated with specific antibody and T cell responses. Many antibodies generated during Schistosoma infection are directed against the numerous glycans expressed by Schistosoma. The nature of glycan epitopes recognized by antibodies in natural schistosomiasis infection serum is largely unknown.Methodology/Principal FindingsThe binding of serum antibodies to glycans can be analyzed efficiently and quantitatively using glycan microarray approaches. Very small amounts of a large number of glycans are presented on a solid surface allowing binding properties of various glycan binding proteins to be tested. We have generated a so-called shotgun glycan microarray containing natural N-glycan and lipid-glycan fractions derived from 4 different life stages of S. mansoni and applied this array to the analysis of IgG and IgM antibodies in sera from children and adults living in an endemic area. This resulted in the identification of differential glycan recognition profiles characteristic for the two different age groups, possibly reflecting differences in age or differences in length of exposure or infection.Conclusions/SignificanceUsing the shotgun glycan microarray approach to study antibody response profiles against schistosome-derived glycan elements, we have defined groups of infected individuals as well as glycan element clusters to which antibody responses are directed in S. mansoni infections. These findings are significant for further exploration of Schistosoma glycan antigens in relation to immunity.

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