Characterization of the Laminated Layer of In vitro Cultivated Echinococcus vogeli Metacestodes

Ingold, Katrin; Dai, Wenjuan; Rausch, Robert L.; Gottstein, Bruno; Hemphill, Andrew

doi:10.2307/3285175

Cited by 8 publications

(14 citation statements)

References 3 publications

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“…In contrast to the Em2(G11) antigen (27,26), Em492 does not appear to be an integral part of the laminated layer but rather is just associated with it, as it was solubilized by 6 M urea extraction, a procedure normally used for the purification of the laminated layer (27,26). In addition, Em492 antigen was progressively released during metacestode in vitro culture and accumulated in the medium supernatant with increasing time of culture.…”

Section: Vol 72 2004 Secretory Component Of E Multilocularis Metacmentioning

confidence: 97%

Isolation and Characterization of a Secretory Component ofEchinococcus multilocularisMetacestodes Potentially Involved in Modulating the Host-Parasite Interface

Walker¹,

Baz

Dematteis

et al. 2004

Infect Immun

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Echinococcus multilocularis metacestodes are fluid-filled, vesicle-like organisms, which are characterized by continuous asexual proliferation via external budding of daughter vesicles, predominantly in the livers of infected individuals. Tumor-like growth eventually leads to the disease alveolar echinococcosis (AE). We employed the monoclonal antibody (MAb) E492/G1, previously shown to be directed against a carbohydraterich, immunomodulatory fraction of Echinococcus granulosus, to characterize potentially related components in E. multilocularis. Immunofluorescence studies demonstrated that MAb E492/G1-reactive epitopes were found predominantly on the laminated layer and in the periphery of developing brood capsules. The respective molecules were continuously released into the exterior medium and were also found in the parasite vesicle fluid. The MAb E492/G1-reactive fraction in E. multilocularis, named Em492 antigen, was isolated by immunoaffinity chromatography. Em492 antigen had a protein/carbohydrate ratio of 0.25, reacted with a series of lectins, and is related to the laminated layer-associated Em2(G11) antigen. The epitope recognized by MAb E492/G1 was sensitive to sodium periodate but was not affected by protease treatment. Anti-Em492 immunoglobulin G1 (IgG1) and IgG2 and, at lower levels, IgG3 were found in sera of mice suffering from experimentally induced secondary, but not primary, AE. However, with regard to cellular immunity, a suppressive effect on concanavalin A-or crude parasite extract-induced splenocyte proliferation in these mice was observed upon addition of Em492 antigen, but trypan blue exclusion tests and transmission electron microscopy failed to reveal any cytotoxic effect in Em492 antigen-treated spleen cells. This indicated that Em492 antigen could be modulating the periparasitic cellular environment during E. multilocularis infection through as yet unidentified mechanisms and could be one of the factors contributing to immunosuppressive events that occur at the host-parasite interface.Alveolar echinococcosis (AE) is caused by the metacestode (larval) stage of Echinococcus multilocularis and is a rare but life-threatening disease, which is confined to the northern hemisphere (14, 15). The adult tapeworm exists as an enteric parasite in the fox and in a few other carnivores, such as wolves, cats, and dogs. Through fecal shedding, parasite eggs are released into the environment. The eggs contain oncospheres, which upon ingestion by a suitable intermediate host and subsequent passage through stomach and intestine get activated, penetrate the mucosa, enter blood and lymphatic vessels, and end up in the liver. In the liver parenchyma, oncospheres develop over time to form mature, asexually proliferating metacestodes, which are characterized by tumor-like growth. Metastasis formation in other organs has been reported (38). Mice and other small mammals act as natural intermediate hosts for E. multilocularis, while humans acquire AE by accidentally ingesting viable parasite eggs.As for many o...

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Section: Vol 72 2004 Secretory Component Of E Multilocularis Metacmentioning

confidence: 97%

Isolation and Characterization of a Secretory Component ofEchinococcus multilocularisMetacestodes Potentially Involved in Modulating the Host-Parasite Interface

Walker¹,

Baz

Dematteis

et al. 2004

Infect Immun

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“…Freshly isolated vesicle walls were processed for TEM as described (15). Briefly, they were fixed for 4 h at room temperature in 2.5% glutaraldehyde in 100 mM phosphate buffer, pH 7.2, containing 0.5% of tannic acid, followed by postfixation in 2% OsO 4 in phosphate buffer.…”

Section: Methodsmentioning

confidence: 99%

“…In vitro-cultured metacestodes were processed for SEM analysis as described (15). Briefly, freshly isolated vesicles were fixed in 2.5% glutaraldehyde in 100 mM phosphate buffer for 4 h at room temperature, followed by postfixation in 2% OsO 4 in phosphate buffer.…”

Section: Methodsmentioning

confidence: 99%

Echinococcus multilocularis Alkaline Phosphatase as a Marker for Metacestode Damage Induced by In Vitro Drug Treatment with Albendazole Sulfoxide and Albendazole Sulfone

Stettler

Siles-Lucas

Sarciron

et al. 2001

Antimicrob Agents Chemother

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Alveolar echinococcosis (AE) is caused by the metacestode stage of the fox tapeworm Echinococcus multilocularis. The disease affects the human liver and occasionally other organs and is fatal if treatment is unsuccessful. The present chemotherapy of AE is based on the administration of benzimidazole carbamate derivatives, such as mebendazole and albendazole. Albendazole treatment has been found to be ineffective in some cases, parasitostatic rather than parasiticidal, and the recurrence rate is rather high. Therefore, chemotherapy usually involves the lifelong uptake of massive doses of albendazole and new treatment options are urgently needed. In order to avoid costly and time-consuming animal experimentation, a first step in searching for novel parasiticidal compounds could be the in vitro drug screening of novel compounds by employing metacestode cultivation. However, presently used techniques (e.g., transmission electron microscopy) for determination of parasite viability involve costly equipment and time-consuming preparation of rather large amounts of parasite material. We therefore searched for a parasite marker which can be easily traced and the presence or absence of which is indicative of parasite viability. In this study we show that the increase of E. multilocularis alkaline phosphatase activity in culture supernatants during in vitro drug treatment with albendazole derivatives correlates with the progressive degeneration and destruction of the metacestode tissue. The inexpensive and rapid assay presented here will serve as an ideal tool for performing first-round in vitro tests on the efficacy of a large number of antiparasitic compounds.

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“…Individual granules are also observed within vesicles in germinal layer cells, and seem to be exocytosed towards the LL. Whereas the microfibrillar matrix is a common feature of the LL of all Echinococcus species [Ingold et al, 2000[Ingold et al, , 2001, the granules have been described only in E. granulosus.…”

mentioning

confidence: 98%

Unique precipitation and exocytosis of a calcium salt of myo‐inositol hexakisphosphate in larval Echinococcus granulosus

Irigoı́n¹,

Casaravilla²,

Iborra³

et al. 2004

J of Cellular Biochemistry

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The ubiquitous intracellular molecule myo-inositol hexakisphosphate (IP6) is present extracellularly in the hydatid cyst wall (HCW) of the parasitic cestode Echinococcus granulosus. This study shows that extracellular IP6 is present as its solid calcium salt, in the form of deposits that are observed, at the ultrastructural level, as naturally electron dense granules some tens of nanometers in diameter. The presence of a calcium salt of IP6 in these structures was determined by two different electron microscopy techniques: (i) the analysis of the spatial distribution of phosphorus and calcium in the outer, acellular layer of the HCW (the laminated layer, LL) through electron energy loss spectroscopy, and (ii) the observation, by transmission electron microscopy, of HCW that were selectively depleted of IP6 by treatment with EGTA or phytase, an enzyme that catalyses the dephosphorylation of IP6. The deposits of the IP6-Ca(II) salt are also observed inside membrane vesicles in cells of the germinal layer (the inner, cellular layer of the HCW), indicating that IP6 precipitates with calcium within a cellular vesicular compartment and is then secreted to the LL. Thus, much as in plants (that produce vesicular IP6 deposits), the existence of transporters for IP6 or its precursors in internal membranes is needed to explain the compound's cellular localisation in E. granulosus.

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Characterization of the Laminated Layer of In vitro Cultivated Echinococcus vogeli Metacestodes

Cited by 8 publications

References 3 publications

Isolation and Characterization of a Secretory Component ofEchinococcus multilocularisMetacestodes Potentially Involved in Modulating the Host-Parasite Interface

Isolation and Characterization of a Secretory Component ofEchinococcus multilocularisMetacestodes Potentially Involved in Modulating the Host-Parasite Interface

Echinococcus multilocularis Alkaline Phosphatase as a Marker for Metacestode Damage Induced by In Vitro Drug Treatment with Albendazole Sulfoxide and Albendazole Sulfone

Unique precipitation and exocytosis of a calcium salt of myo‐inositol hexakisphosphate in larval Echinococcus granulosus

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