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Lectin-FITC labelling of the F. hepatica miracidia. ConA (A) and LCA (B). Staining located in the anterior end of the miracidia. The N-acetylglucosamine binding lectins WGA (C) and LEA (D) reacted intensely with the whole miracidial surface. Scale bars = 80 µm
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The lectin binding properties of Fasciola hepatica miracidia were studied by a panel of fluorescein- and gold-conjugated lectins (ConA, LCA, WGA, LEA, SBA, HPA and UEA-I). The presence of mannose and/or glucose residues was demonstrated with ConA and LCA as weak diffuse fluorescence of the miracidial surface, which was more intense at the anterior...
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Context 1
... results of the lectin labelling experiments are shown in Fig. 1. ConA and LCA binding resulted in very weak dif- fuse fluorescence of the miracidial surface, more intense at the anterior part of the larva (Fig. 1A, B). The N-acetylglu- cosamine-binding lectins (WGA and LEA) reacted intensely with the miracidial surface (Fig. 1C, D). No labelling of N- acetylgalactosamine and/or galactose-specific ...
Context 2
... results of the lectin labelling experiments are shown in Fig. 1. ConA and LCA binding resulted in very weak dif- fuse fluorescence of the miracidial surface, more intense at the anterior part of the larva (Fig. 1A, B). The N-acetylglu- cosamine-binding lectins (WGA and LEA) reacted intensely with the miracidial surface (Fig. 1C, D). No labelling of N- acetylgalactosamine and/or galactose-specific (SBA and HPA) and fucose-specific UEA-I lectins was observed. Specificity of the labelling reactions was confirmed by the ...
Context 3
... results of the lectin labelling experiments are shown in Fig. 1. ConA and LCA binding resulted in very weak dif- fuse fluorescence of the miracidial surface, more intense at the anterior part of the larva (Fig. 1A, B). The N-acetylglu- cosamine-binding lectins (WGA and LEA) reacted intensely with the miracidial surface (Fig. 1C, D). No labelling of N- acetylgalactosamine and/or galactose-specific (SBA and HPA) and fucose-specific UEA-I lectins was observed. Specificity of the labelling reactions was confirmed by the ...
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Fasciola hepatica infection is associated with T helper (Th) 2/T regulatory (Treg) immune responses and increased mast cell numbers. The aim of this study was to examine the interaction between F. hepatica tegumental coat antigen (FhTeg) and mast cells in vivo and in vitro. Firstly, BALB/C, C57BL/6 or STAT6(-/-) mice were infected with F. hepatica...
Citations
... The metacercaria were examined by light microscopy for viability whereas the excretory granules have existed in viable metacercaria [1] [14]. ...
... Parasites and snails. Fasciola hepatica miracidia and Galba truncatula snails were obtained from laboratory maintained life cycles, which were described in detail earlier [ 4 ]. ...
... Parasites and snails. Fasciola hepatica miracidia and Galba truncatula snails were obtained from laboratory maintained life cycles, which were described in detail earlier [ 4 ]. ...
As an alternative to the chemical treatment of farm animals, the control of Fasciola hepatica can be shifted to the botanical treatment of waterborne infectious larval stages of the parasite, realizing the infection at the intermediate host level. In this study, the authors present an experiment to evaluate an efficacy of methanol extract (ME) and essential oil (EO) of Origanum vulgare subsp. hirtum against F. hepatica infection of Galba truncatula snails. In the aquatic environment, miracidia and snails are subjected to ME or EO at different concentrations. At preserved viability of snails, ME and EO have a dose-and time-dependent effect provoking complete prevention of snail invasion. As a larvicide, ME showed an effect with LC 50 values of 23 µg/mL, 8.3 µg/mL, 0.8 µg/mL and 0.3 µg/mL at 15 min, 30 min, 60 min and 120 min, respectively. EO showed a rapid effect with LC 50 values of 0.17 µL/mL, 0.07 µL/mL, 0.03 µL/mL at 15 min, 30 min and 60 min, respectively. In conclusion, targeted miracidial treatment can be a new opportunity to control fasciolosis. ME and EO from O. vulgare subsp. hirtum can be applied as effective suppressors of the transmission of liver fluke F. hepatica at the intermediate host level and this can be a valuable tool for the control of fasciolosis.
... The Wistar rats (Rattus norvegicus) used were aged 30 days, male and weighed 100 ± 10 g each. Fasciola hepatica used where obtained from laboratory maintained life cycle of the parasite, using Galba truncatula snails as intermediate host, and male Wistar rats as definitive host [2]. ...
... September 2021 | Volume 9 | Issue 10 | Page 1696 still need more studies. Researchers have identified reactivity of miracidia, redia and sporocysts surface (Georgieva et al., 2012) and gut of adult flukes (Farahnak et al., 2010;McAllister et al., 2011) with lectin this explains the presence of Glc and Man, GlcNAc or GalNAc residues. F. hepatica adult stage contains many types of glycan structure like, LDN [LacdiNAc (GalNAcβ1,4GlcNAc], fucosylated LDN, LDNF, [GalNAcβ1,4(Fucβ1-3)GlcNAc], truncated O-glycans, known as T antigen (Galβ1-3GalNAcβ1-O-Thr/Ser) LDN and LDNF (Nyame et al., 1998;Van Die and Cummings, 2010a). ...
... Parasites and snails. Fasciola hepatica miracidia and Galba truncatula snails were obtained from laboratory maintained life cycles, which were described in detail earlier [ 4 ]. ...
As an alternative to the chemical treatment of farm animals, the control of Fasciola hepatica can be shifted to the botanical treatment of waterborne infectious larval stages of the parasite, realizing the infection at the intermediate host level. In this study, the authors present an experiment to evaluate an efficacy of methanol extract (ME) and essential oil (EO) of Origanum vulgare subsp. hirtum against F. hepatica infection of Galba truncatula snails. In the aquatic environment, miracidia and snails are subjected to ME or EO at different concentrations. At preserved viability of snails, ME and EO have a dose-and time-dependent effect provoking complete prevention of snail invasion. As a larvicide, ME showed an effect with LC 50 values of 23 µg/mL, 8.3 µg/mL, 0.8 µg/mL and 0.3 µg/mL at 15 min, 30 min, 60 min and 120 min, respectively. EO showed a rapid effect with LC 50 values of 0.17 µL/mL, 0.07 µL/mL, 0.03 µL/mL at 15 min, 30 min and 60 min, respectively. In conclusion, targeted miracidial treatment can be a new opportunity to control fasciolosis. ME and EO from O. vulgare subsp. hirtum can be applied as effective suppressors of the transmission of liver fluke F. hepatica at the intermediate host level and this can be a valuable tool for the control of fasciolosis.
... In the present work, we conducted an experiment by subjecting F. hepatica miracidia and G. truncatula snails to the presence of different monosaccharides to test their ability to impact on lectin/carbohydrate recognition. Monosaccharides were selected on the basis of our previous studies identifying carbohydrates exposed on the surface of miracidia and snails with lectin-binding techniques (Georgieva et al. 2012(Georgieva et al. , 2014(Georgieva et al. , 2016. Finding appropriate inhibitors of parasite-snail recognition is a key to developing a strategy for control of fasciolosis at the intermediate host levels. ...
... truncatula) susceptible to Fasciola infection as intermediate host and male Wistar rats as definitive host. The procedures were described previously in details by Georgieva et al. (2012). Briefly, rats were fed with 15 metacercariae each. ...
... Carbohydrates in the form of monosaccharides (thick arrow) which are added to the aquatic environment, interfere with the ability of the parasite to recognize its snail host, probably by blocking of specific receptors. Abbreviations: Manmannose, Glcglucose, GlcNAc -N-acetylglucosamine. * Georgieva et al. (2012), ** Georgieva et al. (2016) interactions. These results are taken as evidence that surface oligosaccharides play an essential role in the recognition events that accompany the miracidial hostfinding, penetration and invasion of specific snail hosts. ...
Fasciola hepatica is one of the etiological agents of fasciolosis, a widespread disease in domestic animals and occasionally in humans. Fasciolosis may be reduced by blocking the parasite transmission through its intermediate hosts. In the present work, different monosaccharides have been tested for their ability to impact on ligand/receptor interactions at the interface between the parasite and the intermediate host. Laboratory snails were subjected to miracidia in the presence of methyl-α-D-mannopyranoside (MetMan), α-D-glucose (Glc), N-acetyl-D-glucosamine (GlcNAc), D-(+)-galactose (Gal), N-acetyl-D-galactosamine (GalNAc) or L-(−)-fucose (Fuc), in 10 mM concentration. The snail survival rates and the prevalence of infection were determined after 50 days. Survived snails in the study groups varied from 78% to 97%. A remarkable reduction in the number of parasite-infected snails was observed in groups subjected to MetMan, Glc, or GlcNAc – 36.9%, 10.9%, and 11.9%, respectively, compared to 92.7% in the control group. Other tested monosaccharides had a low impact on snail infection. The results point to the implication that surface carbohydrate/receptor interactions are among the determining factors concerning the transmission of F. hepatica by the specific vector Galba truncatula. Biological recognition between the two organisms can be interfered with appropriate monosaccharides and this can be used to develop an alternative method for control of fasciolosis at the intermediate host level.
... Because these lectins play important roles in innate immunity, their recognition of cercarial structures may be an inseparable part of host-parasite interactions which lead to, e.g., complement activation, opsonization, and cell adherence. They need not always represent a signal for cercarial transformation, although this was clearly demonstrated in our study with exogenous lectins, and a similar effect was observed in some other larval trematode transformations (e.g., Fasciola hepatica miracidia to mother sporocysts) [59]. ...
The invasive larvae (cercariae) of schistosomes penetrate the skin of their definitive hosts. During the invasion, they undergo dramatic ultrastructural and physiological transitions. These changes result in the development of the subsequent stage, schistosomulum, which migrates through host tissues in close contact with host’s immune system. One of the striking changes in the transforming cercariae is the shedding of their thick tegumental glycocalyx, which represents an immunoattractive structure; therefore its removal helps cercariae to avoid immune attack. A set of commercial fluorescently labeled lectin probes, their saccharide inhibitors and monoclonal antibodies against the trisaccharide Lewis-X antigen (LeX, CD15) were used to characterize changes in the surface saccharide composition of the neuropathogenic avian schistosome Trichobilharzia regenti during the transformation of cercariae to schistosomula, both in vitro and in vivo. The effect of various lectins on glycocalyx shedding was evaluated microscopically. The involvement of peptidases and their inhibitors on the shedding of glycocalyx was investigated using T. regenti recombinant cathepsin B2 and a set of peptidase inhibitors. The surface glycocalyx of T. regenti cercariae was rich in fucose and mannose/glucose residues. After the transformation of cercariae in vitro or in vivo within their specific duck host, reduction and vanishing of these epitopes was observed, and galactose/N-acetylgalactosamine emerged. The presence of LeX was not observed on the cercariae, but the antigen was gradually expressed from the anterior part of the body in the developing schistosomula. Some lectins which bind to the cercarial surface also induced secretion from the acetabular penetration glands. Seven lectins induced the shedding of glycocalyx by cercariae, among which five bound strongly to cercarial surface; the effect could be blocked by saccharide inhibitors. Mannose-binding protein, part of the lectin pathway of the complement system, also bound to cercariae and schistosomula, but had little effect on glycocalyx shedding. Our study did not confirm the involvement of proteolysis in glycocalyx shedding.
... F. hepatica were obtained from experimental life cycle of the parasite maintained using Galba snails as intermediate and male Wistar rats as definitive hosts. The procedures have been previously described in detail by Georgieva et al. (2012). Tissue samples of adult G. truncatula snails from either uninfected snails, or snails infected with F. hepatica were taken eight, 14 and 50 days post infection. ...
... The sections were dewaxed and hydrated. After blocking of non-specific binding with 5% bovine serum albumin (Sigma-Aldrich) the samples were labelled with lectinfluorescein isothiocyanate (FITC) conjugates following the procedures described by Georgieva et al. (2012). The applied lectins (Sigma-Aldrich or Vector Labs), their major carbohydrate specificities and the final concentrations in PBS [or PBS supplemented with 0.1 mM CaCl 2 and MnCl 2 for Concanavalin A (ConA)] are listed in Table I. ...
... Intensity of staining: ++ = intense; + = moderate; ± = weak; − = none. * Georgieva et al. (2012Georgieva et al. ( , 2014); 1 it refers to the labelling of entire larvae; 2 the labelling of the tissue sections. amine, N-acetylgalactosamine and galactose residues. ...
The presence and distribution of surface carbohydrates in the tissues of Galba truncatula snails uninfected or after infection with Fasciola hepatica as well as on the surface of the snail-pathogenic larval stages of the parasite were studied by lectin labelling assay. This is an attempt to find similarities that indicate possible mimicry, utilised by the parasite as an evasion strategy in this snail-trematode system. Different binding patterns were identified on head-foot-mantle, hepatopancreas, genital glands, renopericardial complex of the host as well as of the snail-pathogenic larval stages of F. hepatica. The infection with F. hepatica leads to changes of labelling with Glycine max in the head-mantle cells and Arachis hypogaea in the tubular epithelium of the hepatopancreas. The lectin binding on the other snail tissues is not changed by the development of the larvae. Our data clearly demonstrated the similarity in labelling of G. truncatula tissues and the surface of the snail-pathogenic larval stages of F. hepatica. The role of glycosylation of the contact surfaces of both organisms in relation to the host-parasite interactions is also discussed.
... It has been shown that the adult stage of F. hepatica contains at least several types of glycans and glycan motifs such as fucosylated LacdiNAc (LDN-F) motifs [26], mucin-type O-glycosylated proteins [27], the glycolipid CD77 [28] and other glycoceramides [29,30]. Even though the full range of F. hepatica glycoconjugates has not been characterised, the presence of various glycans has been confirmed by lectin binding studies not only in adult stages [25,29,31] but also in the other lifecycle stages (miracidia, rediae, sporocysts) [32][33][34]. However, systematic studies of glycosylation in the NEJ have not yet been carried out. ...
Recently, the prevalence of Fasciola hepatica in some areas has increased considerably and the availability of a vaccine to protect livestock from infection would represent a major advance in tools available for controlling this disease. To date, most vaccine-target discovery research on this parasite has concentrated on proteomic and transcriptomic approaches whereas little work has been carried out on glycosylation. As the F. hepatica tegument (Teg) may contain glycans potentially relevant to vaccine development and the Newly Excysted Juvenile (NEJ) is the first lifecycle stage in contact with the definitive host, our work has focused on assessing the glycosylation of the NEJTeg and identifying the NEJTeg glycoprotein repertoire. After in vitro excystation, NEJ were fixed and NEJTeg was extracted. Matrix-assisted laser desorption ionisation-time of flight-mass spectrometry (MALDI-TOF-MS) analysis of released N-glycans revealed that oligomannose and core-fucosylated truncated N-glycans were the most dominant glycan types. By lectin binding studies these glycans were identified mainly on the NEJ surface, together with the oral and ventral suckers. NEJTeg glycoproteins were affinity purified after targeted biotinylation of the glycans and identified using liquid chromatography and tandem mass spectrometry (LC-MS/MS). From the total set of proteins previously identified in NEJTeg, eighteen were also detected in the glycosylated fraction, including the F. hepatica Cathepsin B3 (FhCB3) and two of the Cathepsin L3 (FhCL3) proteins, among others. To confirm glycosylation of cathepsins, analysis at the glycopeptide level by LC-ESI-ion-trap-MS/MS with collision-induced dissociation (CID) and electron-transfer dissociation (ETD) was carried out. We established that cathepsin B1 (FhCB1) on position N80, and FhCL3 (BN1106_s10139B000014, scaffold10139) on position N153, carry unusual paucimannosidic Man2GlcNAc2 glycans. To our knowledge, this is the first description of F. hepatica NEJ glycosylation and the first report of N-glycosylation of F. hepatica cathepsins. The significance of these findings for immunological studies and vaccine development is discussed.
