Evaluation of the C-Terminal Fragment of Entamoeba histolytica Gal/GalNAc Lectin Intermediate Subunit as a Vaccine Candidate against Amebic Liver Abscess

Abstract

BackgroundEntamoeba histolytica is an intestinal protozoan parasite that causes amoebiasis, including amebic dysentery and liver abscesses. E. histolytica invades host tissues by adhering onto cells and phagocytosing them depending on the adaptation and expression of pathogenic factors, including Gal/GalNAc lectin. We have previously reported that E. histolytica possesses multiple CXXC sequence motifs, with the intermediate subunit of Gal/GalNAc lectin (i.e., Igl) as a key factor affecting the amoeba's pathogenicity. The present work showed the effect of immunization with recombinant Igl on amebic liver abscess formation and the corresponding immunological properties.Methodology/Principal FindingsA prokaryotic expression system was used to prepare the full-length Igl and the N-terminal, middle, and C-terminal fragments (C-Igl) of Igl. Vaccine efficacy was assessed by challenging hamsters with an intrahepatic injection of E. histolytica trophozoites. Hamsters intramuscularly immunized with full-length Igl and C-Igl were found to be 92% and 96% immune to liver abscess formation, respectively. Immune-response evaluation revealed that C-Igl can generate significant humoral immune responses, with high levels of antibodies in sera from immunized hamsters inhibiting 80% of trophozoites adherence to mammalian cells and inducing 80% more complement-mediated lysis of trophozoites compared with the control. C-Igl was further assessed for its cellular response by cytokine-gene qPCR analysis. The productions of IL-4 (8.4-fold) and IL-10 (2-fold) in the spleen cells of immunized hamsters were enhanced after in vitro stimulation. IL-4 expression was also supported by increased programmed cell death 1 ligand 1 gene.Conclusions/SignificanceImmunobiochemical characterization strongly suggests the potential of recombinant Igl, especially the C-terminal fragment, as a vaccine candidate against amoebiasis. Moreover, protection through Th2-cell participation enabled effective humoral immunity against amebic liver abscesses.

Evaluation on Elongation Factor 1 Alpha of Entamoeba histolytica Interaction with the Intermediate Subunit of the Gal/GalNAc Lectin and Actin in Phagocytosis

“…Mouse original polyclonal antibodies against rEF1a were prepared after immunization with target proteins described in a previous study [37]. The specificity of polyclonal antibodies examined by Western blotting following the process described above, using mouse anti-rEF1a polyclonal antibodies (1:4000) as the primary and goat anti-mouse IgG as the secondary antibody.…”

Interaction between human mucins and parasite glycoproteins: the role of lectins and glycosidases in colonization by intestinal protozoa

“…On the other hand, igl lacks a carbohydrate recognition motif; however, it has galactose-binding activity and contains several CXXC and CXC domains, which indicates protein-protein interactions; monoclonal antibodies against igl significantly inhibit ameba adherence, phagocytosis and toxicity 25 . In addition, 96% of hamsters immunized with the C-terminal fragment of igl were protected against liver abscess formation, and the antibodies generated inhibited up to 80% the adhesion of trophozoites to mammalian cells in vitro, supporting the use of this fragment as a vaccine to prevent amebiasis 26 .…”

Characteristics of inflammatory reactions during development of liver abscess in hamsters inoculated with Entamoeba nuttalli

“…histolytica SAW755CR and E . nuttalli GY4 strains were grown under axenic conditions at 36.5°C in YIMDHA-S medium [ 22 ] containing 15% (v⁄v) heat-inactivated adult bovine serum. Trophozoites were harvested during the logarithmic growth phase (48 to 72 h) by chilling on ice for 5 min.…”

“…Gene expression of IFN-γ, TNF-α, IL-1β, IL-4, IL-6, IL-10, nitric oxide synthase (iNOS), arginine enzyme I (Arg-1) and mannose receptor I (MRC-I) in liver tissues was examined by quantitative real-time PCR (qRT-PCR) using the primers listed in Table 1 [ 22 , 25 – 26 ]. Briefly, total RNA (1 μg) of tissue from the edge of a liver abscess was purified with an RNeasy Plus Mini kit (Qiagen).…”