Abstract
Fasciola hepatica, commonly known as liver fluke, is a trematode that causes Fasciolosis in ruminants and humans. The outer tegumental coat of F. hepatica (FhTeg) is a complex metabolically active biological matrix that is continually exposed to the host immune system and therefore makes a good vaccine target. F. hepatica tegumental coat is highly glycosylated and helminth-derived immunogenic oligosaccharide motifs and glycoproteins are currently being investigated as novel vaccine candidates. This report presents the first systematic characterization of FhTeg glycosylation using lectin microarrays to characterize carbohydrates motifs present, and lectin histochemistry to localize these on the F. hepatica tegument. We discovered that FhTeg glycoproteins are predominantly oligomannose oligosaccharides that are expressed on the spines, suckers and tegumental coat of F. hepatica and lectin blot analysis confirmed the abundance of N- glycosylated proteins. Although some oligosaccharides are widely distributed on the fluke surface other subsets are restricted to distinct anatomical regions. We selectively enriched for FhTeg mannosylated glycoprotein subsets using lectin affinity chromatography and identified 369 proteins by mass spectrometric analysis. Among these proteins are a number of potential vaccine candidates with known immune modulatory properties including proteases, protease inhibitors, paramyosin, Venom Allergen-like II, Enolase and two proteins, nardilysin and TRIL, that have not been previously associated with F. hepatica. Furthermore, we provide a comprehensive insight regarding the putative glycosylation of FhTeg components that could highlight the importance of further studies examining glycoconjugates in host-parasite interactions in the context of F. hepatica infection and the development of an effective vaccine.
Highlights
From the ‡Fundamental and Translational Immunology, School of Biotechnology, Faculty of Science and Health, Dublin City University, Glasnevin, Dublin 9, Ireland; §School of Biological Sciences, Medical Biology Centre (MBC), Queen’s University Belfast, Belfast, Northern Ireland, UK; ¶Glycoselect, Dublin City University, Glasnevin, Dublin 9; ʈGlycoscience Group, National Centre for Biomedical Engineering Science, National University of Ireland Galway, Ireland; **Regenerative Medicine Institute, NUI Galway, Ireland; ‡‡Carbohydrate Signalling Group, Microbiology, NUI Galway, Ireland; §§Department of Parasitology, Leiden University Medical Center, Leiden, The Netherlands
Lectin Microarray Profiling of the F. hepatica Tegument Demonstrates Characteristic Glycoprotein Content—The glycosylation of FhTeg was characterized on a lectin microarray featuring a comprehensive panel of plant, bacterial and fungal lectins (Fig. 1A and supplemental Table S1)
FhTeg exhibited a high affinity for the Man-binding lectins (Calsepa, NPA, GNA, HHA and Con A; supplemental Table S1), which suggested a predominance of glycoproteins modified with oligomannose type N-linked oligosaccharide structures at the parasite surface
Summary
From the ‡Fundamental and Translational Immunology, School of Biotechnology, Faculty of Science and Health, Dublin City University, Glasnevin, Dublin 9, Ireland; §School of Biological Sciences, Medical Biology Centre (MBC), Queen’s University Belfast, Belfast, Northern Ireland, UK; ¶Glycoselect, Dublin City University, Glasnevin, Dublin 9; ʈGlycoscience Group, National Centre for Biomedical Engineering Science, National University of Ireland Galway, Ireland; **Regenerative Medicine Institute, NUI Galway, Ireland; ‡‡Carbohydrate Signalling Group, Microbiology, NUI Galway, Ireland; §§Department of Parasitology, Leiden University Medical Center, Leiden, The Netherlands. We provide a comprehensive insight regarding the putative glycosylation of FhTeg components that could highlight the importance of further studies examining glycoconjugates in host-parasite interactions in the context of F. hepatica infection and the development of an effective vaccine. Fasciola hepatica is a parasitic flatworm of livestock and the causative agent of Fasciolosis, a disease that results in major economic losses to the agricultural industry globally, estimated at $3 billion annually [1]. It is only recently that the fluke surface tegument has been the subject of close investigations by molecular techniques [4, 5, 6, 7] with the aim of discovering novel potential targets for both drug and vaccine development. The spines facilitate feeding of Molecular & Cellular Proteomics 15.10. It is highly likely that the shed tegumental coat is released into the blood stream and studies have shown the presence of antitegumental antibodies in serum form infected animals [9]
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