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PubMed1982

The production of eggs by Echinococcus multilocularis in the laboratory following in vivo and in vitro development.

Adult worms of Echinococcus multilocularis produced eggs in vitro following partial development in the definitive host for 20-21 days and maturation for 8 days in a monophasic or diphasic medium. The technique was shown to be reproducible with morphologically normal eggs produced in two separate experiments. The onset of egg production in vitro was identical to that in vivo although the number of eggs produced by each worm was less in vitro.

Animals↗
PubMed1992

In vitro culture of the strobilar stage of Echinococcus granulosus of sheep and donkey origin from Jordan.

Protoscolices of Echinococcus granulosus isolated from hydatid cysts of sheep and donkeys in Jordan were cultured in vitro using a modified diphasic culture system. Protoscolices from these two sources manifested differences in the mode of development, evagination and growth rates. Protoscolices isolated from sheep cysts grew in vitro in the polyzoic direction up to the three- to four-segmented mature worms, reaching a length of 2.9 mm. In contrast, donkey protoscolices failed to develop beyond the early stages, even after 67 days of culturing. On prolonged culturing, few worms of donkey origin reached the banding and segmentation stages, attaining a maximal length of 1.6 mm at periods ranging between 81 and 114 days of culturing. None of these segmented worms showed genital differentiation or analgen. The evagination and growth rates of protoscolices isolated from donkey liver cysts were compared with those obtained from sheep liver or lung cysts. The most significant difference in these rates occurred at the commencement of the segmentation stage. Differences in the development, growth and evagination rates observed between the donkey and sheep forms may reflect the strain variation of E. granulosus in this country.

Animals↗
PubMed1992

Echinococcus multilocularis: immunological study on the "Em2-positive" laminated layer during in vitro and in vivo post-oncospheral and larval development.

Echinococcus multilocularis oncospheres, primary vesicular cysts, and protoscolices were assessed in vitro and in vivo for their potential to synthesize a PAS-positive laminated layer containing monoclonal antibody (mAb) G11-binding Em2 antigen. The presence of Em2 antigen in developed oncospheres and cysts was subsequently correlated to the potential of in vivo development into a secondary metacestode in recipient host mice, which also responded by anti-Em2 serum antibody formation. In contrast, protoscolices failed to develop the "Em2-positive" layer in vitro under the selected experimental conditions. The failure to develop subsequently in vivo into a secondary metacestode was underlined by a lack of anti-Em2 serum antibody formation by the hosts. We furthermore developed a technique to obtain E. multilocularis clones by inoculating single oncospheres into recipient mice.

Animals↗
PubMedDate not supplied

Activation of human complement by crude and purified antigens from Echinococcus granulosus.

Hydatid cyst fluid of Echinococcus granulosus was found to interact with human complement. This interaction resulted in the depletion of hemolytic complement in normal serum and in the electrophoretic conversion of C3 and factor B. The results provided evidence of an activation of complement via the alternative pathway when hydatid cyst fluid was used in the mixtures. Also purified hydatid antigen preparations were found to interact with complement in vitro; this interaction resulted in the consumption of complement not only via the alternative pathway, but also via the classical pathway.

Animals↗
PubMed1997

Establishment of a primary culture of Echinococcus multilocularis germinal cells.

This study was designed to establish an in vitro primary culture of germinal cells of Echinococcus multilocularis, a parasite that causes alveolar echinococcosis of the liver (AEL). We also investigated the temperature-dependency of the cultured cells. The germinal cells, which originated from a human lesion, were cultured by an original fluid-suspension method at 25 degrees C or 37 degrees C for 4 weeks. Anchorage-dependent and -independent cells were observed by light microscopy, transmission electron microscopy, and immunocytochemistry to confirm their origin. Cell number and viability were examined by immunocytochemistry and mitochondrial exclusion test. The cultured cells were also inoculated into jirds (Meriones unguiculatus) to evaluate metacestode formation. Morphology and immunocytochemistry showed that the cultured cells were typically germinal cells. The cell number declined gradually over the 4-week culture period, but viability remained at 50% at 3 weeks. These findings were not associated with either of the two culture temperatures; moreover, host-associated cells were not noted in the cultured cells at 25 degrees C. The implanted cells formed metacestodes in the jird peritoneal cavity, and their histology demonstrated mature and typical alveolar-type echinococcal cysts. We successfully established an in vitro primary culture of germinal cells. This should contribute to future studies, and, hence, a better outcome for patients with AEL.

Animals↗
PubMed2006

The ability of 67Ga scintigraphy to detect the lesions of Echinococcus multilocularis infection: preliminary results.

AIM: To assess the ability of 67Ga scintigraphy to detect the lesions of Echinococcus multilocularis (EM) infection. MATERIALS AND METHODS: An animal model of EM infection was developed. The infected tissues taken from stock infection were placed into the abdominal cavity of uninfected animals operatively. The success of implantation was controlled 20-25 days after implantation. Five infected and 2 healthy animals were studied. All of the animals were examined by ultrasound before the scintigraphic evaluation. After the injection of 7.4 MBq (200 microCi) 67Ga citrate intravenously, static images from the whole anterior thorax and abdomen were obtained at 24, 48 and 72 hours. Visual and semiquantitative analyses were performed. In semiquantitative analysis, an irregular region of interest was drawn over the thorax as the background, excluding the heart and a second region of interest was drawn over the abdomen, excluding the liver and spleen. Abdomen/ background ratios were calculated using the mean counts. RESULTS: In the visual evaluation, it was noticed that there was considerably increased 67Ga uptake in the abdomens of the infected animals. In infected animals, mean abdomen/background ratios at 48 and 72 hours (3.76 +/- 1.04, 4.13 +/- 0.72, respectively) were increased compared with mean abdomen/background ratios at 24 hours (2.94 +/- 0.77). These increases in abdomen/background ratios were statistically significant at 72 hours (p = 0.04). Between the infected animals and control group, mean abdomen/background ratios were compared, and statistically significant differences were found in the images obtained at 48 and 72 hours. CONCLUSION: Imaging at 72 hours seems to be more suitable imaging time for the diagnosis of alveolar echinococcosis. 67Ga scintigraphy may successfully demonstrate the lesions of EM infection localized intraperitoneally. The method of 67Ga scintigraphy is useful because it is simple, non-invasive and relatively safe.

Animals↗
PubMed2000

In vitro effect of praziquantel and albendazole combination therapy on the larval stage of Echinococcus granulosus.

Protoscolices of Echinococcus granulosus were incubated in vitro with praziquantel (PZ), albendazole (ABZ), or a combination of both (PZ + ABZ). PZ and ABZ displayed slower protoscolicidal activity when applied separately than when used in combination. Despite the low PZ + ABZ concentrations used, protoscolex viability dropped rapidly (within 15 days). At this time, cysts did not develop following their inoculation into mice. The ultrastructural changes induced in the protoscolices by PZ + ABZ were (a) the loss of sucker concavity, (b tegumental contraction of the soma region, (c) the formation of digitiform tegumental extensions, (d) destruction of the tegument, and (e) the degeneration of parenchyma cells as reflected by the presence of numerous lamellar bodies. The PZ + ABZ treatment was effective only against small cysts, which had collapsed at 10 days postinoculation (p.i.). This treatment caused the following alterations: (a) loss of cyst turgidity at 6 days p.i.; (b) separation of the laminated and germinal layers; (c) loss of microtriches; (d) the appearance of numerous lipid droplets in the inner region of the germinal layer, (e) vacuolation of the cyton cytoplasm; and (f) the formation of abundant autophagosomes, which finally led to loss of the integrity of the germinal layer.

Albendazole↗
PubMed1997

Interconverting gating modes of a nonselective cation channel from the tapeworm Echinococcus granulosus reconstituted on planar lipid bilayers.

A 107-pS (symmetrical 150 mM KCl), nonselective cation channel was reconstituted from a microsomal membrane fraction of the larval stage of the tapeworm Echinococcus granulosus. Most of the time, it displayed a high open probability (>>0.95) irrespective of either the applied voltage, Ca2+, Ba2+, or tetraethylammonium concentration. Nevertheless, in contrast with this "leaklike" behavior, less frequently this "all-the-time-open" channel reversibly entered two different kinetic modes. One of them was characterized by lower Po values and some voltage sensitivity (V(1/2) congruent with 129 mV, and an equilibrium constant for channel closing changing e-fold per 63-mV change) the kinetic analysis revealing that it resulted from the appearance of voltage-sensitivity in the mean closed times and a sixfold increase in the equilibrium constant for channel closing at 0 mV. The other mode was characterized by a very fast open-close activity leading to poorly resolved current levels and a Po around 0.6-0.7 which, occasionally and in a voltage-sensitive manner, entered a long-lived nonconducting state. However, the rare nature of these mode-shifting transitions precluded a more detailed analysis of their kinetics. The conductive properties of the channel were not affected by these switches. Model gating alone does not seem to ensure any physiological role of this channel and, instead, some other channel changes must occur if this phenomenon were to be of regulatory importance in vivo. Thus, mode-shifting might constitute an alternative target for channel activity modulation also in tapeworms.

Animals↗
PubMed2005

Contrast-enhanced CT and MRI findings of atypical hepatic Echinococcus alveolaris infestation.

Diagnosis of liver infestation by Echinococcus alveolaris (EA) is based on serological and radiological findings. In this report, we present a 15-year-old girl with atypical hepatic EA infestation showing central punctate calcifications and contrast enhancement on the portal and late phases of CT and MRI. CT showed a hypodense mass involving more than half of the liver with prominent central calcifications. MRI revealed hypointense signal of the infiltrative mass on both T1- and T2-weighted images. Contrast enhancement is a unique finding in hepatic EA infestation that may cause difficulties with diagnosis. MRI may provide invaluable information in the diagnosis of EA infestation of the liver, either by disclosing the infiltrative pattern of infestation without significant effect to vascular structures, or by the signal characteristics.

Adolescent↗
PubMed2002

An intact laminated layer is important for the establishment of secondary Echinococcus multilocularis infection.

Echinococcus multilocularis causes alveolar echinococcosis primarily in rodents, but also in humans where it represents one of the most lethal helmintic infections. We used a susceptible mouse (C57BL/6) model to demonstrate failure in controlling secondary infection with the E. multilocularis metacestode, even when performed at the lowest possible infection dose. This was achieved by intraperitoneal or intrahepatic inoculation of a single parasite vesicle. In secondary infections, the primary physical barrier between the parasite and the host is constituted by the acellular laminated layer (LL), which is predominantly composed of high-molecular-weight glycans and surrounds the entire metacestode. Only those metacestode structures which exhibited an intact LL were successful in establishing infection, whereas metacestodes which were punctured - thus exhibiting an opened LL and thereby an accessible germinal layer - were no longer infective. Conversely, both types of vesicle survived in vivo maintenance, as assessed by RT-PCR based upon II/3 gene expression. In consequence, the encapsulating LL appears to be one of the key factors that mediates survival and successful proliferation of the parasite metacestode in vivo.

Animals↗
PubMed2002

Characterization of the immune response induced by a carbohydrate enriched fraction from Echinococcus granulosus protoscoleces in patients with cystic hydatid disease.

The serological and cellular immune response against a carbohydrate-enriched fraction of Echinococcus granulosus (E4+) was studied in a group of patients with cystic hydatid disease (CHD). The profile of IgG subclass against E4+ and the native non-enriched extract (protoscolex somatic antigen, PSA) were compared. A relatively higher ratio of IgG2:IgG4 was induced by E4+ than by PSA (12.4 vs 3.6 respectively). Serological data were associated with clinical parameters dealing with the outcome of disease. The expression of IgG1 and IgG4 subclasses against both of the antigens was associated with the progression of the disease. Interestingly, anti-PSA IgG4 was the only variable that was specifically associated with the stage of cysts (P < 0.04). Similar levels of IL-13 were evoked by peripheral blood mononuclear cells from patients with CHD when incubated with either of the antigens, while higher levels of IL-10 were obtained in supernatants from cells stimulated with E4+ (P < 0.029) than from those stimulated by PSA. Considerable amounts of IL-10 (median 60 pg/ml) were obtained in the supernatants of cells from healthy individuals incubated with E4+. Our results suggest a putative role for E4+ in immunoregulation during the course of infection with E. granulosus in humans.

Adult↗
PubMed2003

Analysis of cDNAs coding for immunologically dominant antigens from an oncosphere-specific cDNA library of Echinococcus multilocularis.

A cDNA library based on mRNA from oncospheres of Echinococcus multilocularis was constructed and screened with an oncosphere-specific rabbit serum. cDNA sequences of three clones that were isolated out of this library are discussed: one codes for a serpin-like proteinase inhibitor, the first isolated from cestodes. Two other clones code for dominant oncosphere antigens and represent homologues of known genes: one is known from several taeniid cestodes as a protective antigen containing fibronectin III domains, the second is related to genes of small heat shock proteins. It contains an internal duplication that might be specific for platyhelminths.

Amino Acid Sequence↗
PubMed2003

A model for the transmission of Echinococcus multilocularis in Hokkaido, Japan.

A mathematical model for Echinococcus multilocularis transmission would be useful for estimating its prevalence and determining control strategies. We propose a mathematical model which quantitatively describes the transmission of E. multilocularis in Hokkaido, Japan. The model takes into account the influence of the dynamics of both the definitive and the intermediate host populations, which show large scale seasonal variation as they are wild animals. The simulations based on the model clarify the mechanism of seasonal transmission of E. multilocularis quantitatively, notwithstanding a lack of seasonal prevalence data. At present, human alveolar echinococcosis is prevalent throughout the mainland of Hokkaido. The risk of being infected in the human population has been investigated by analyzing the seasonal fluctuation in parasite egg dispersion in the environment. This is necessary for the planning of more suitable preventive measures against E. multilocularis.

Animals↗
PubMed2003

Detection of Echinococcus multilocularis antigens in faeces by ELISA.

Faecal samples deriving from 391 animals belonging to nine species (polecats, badgers, martens, weasels, rats, dogs, cats, red foxes, raccoon-dogs) were examined by capture ELISA for the presence of the Echinococcus multilocularis coproantigen. The main claim of our studies is the reliable detection of E. multilocularis coproantigens, mainly in the faeces of foxes, dogs and cats. For the first time in coproantigen detection we used a "double-sandwich" ELISA. The main advantage of this method is the higher specificity and better differentiation of positive and negative faecal samples, in comparison with sandwich ELISA. The overall specificity of double-sandwich ELISA was 95.1% with only 16 of 327 E. multilocularis-free animals giving false-positive results. The E. multilocularis coproantigen was detected by double-sandwich ELISA in 37.5% of examined red foxes and in 8.0% of examined raccoon-dogs, compared with a prevalence of just 29.8% in red foxes and 8.0% in raccoon-dogs, as determined by parasitological techniques.

Animals↗
PubMed2004

Long-term in vitro cultivation of Echinococcus multilocularis metacestodes under axenic conditions.

We report here on the development of an in vitro system for the long-term cultivation of Echinococcus multilocularis larvae under axenic conditions. In the absence of feeder cells from the host, long-term survival of the parasite depended strictly on low oxygen conditions and the presence of reducing agents in the medium. Host serum supported survival of the parasite but the growth of metacestode vesicles and differentiation towards the protoscolex stage only occurred in the presence of culture medium that was preconditioned by hepatoma cells or several other immortal cell lines. On the basis of this in vitro system, future analyses on the identification of host-derived growth factors for E. multilocularis will be greatly facilitated.

Animals↗
PubMed2004

The Tibetan hare Lepus oiostolus: a novel intermediate host for Echinococcus multilocularis.

Multicystic hydatids have been found in the livers of hares ( Lepus oiostolus) examined from the Qinghai-Tibet plateau of China. In this study, the causative species was definitively identified as Echinococcus multilocularis by mitochondrial DNA sequencing. This is the first confirmation of larval E. multilocularis from hares. The hydatids contained protoscolices, suggesting that the hare may contribute to the transmission of E. multilocularis on the Tibetan plateau.

Animals↗
PubMed2004

An in vitro model to evaluate the cytokine response in Echinococcus infections.

With the aim of proposing an alternative model to animal experimentation, we investigated cytokine production in response to antigens in an in vitro system. This is a co-culture system of healthy human leukocytes and enterocyte-like Caco-2 cells. The antigens tested, EgA31, EgTrp, and FABP1, are candidates for vaccines in infections caused by Echinococcus spp. in the gut. All three have previously been described in the protoscolex stage and belong to protein families which confer protective immunity against several helminths. In this study, we evaluate the Th1/Th2 profile (Th1: IL-12, IFN-gamma; Th2: IL-6, IL-10) in response to protoscoleces, EgA31 and the mixture of EgA31, EgTrp and FABP1. No cytokine production was detected in response to protoscoleces. Neither IFN-gamma nor IL-6, but a significant IL-10 and IL-12 concentration was detected in response to both types of antigens. These findings suggest that EgA31 and the mixture EgA31/EgTrp/FABP1 generated an immunogenic response associated with a mixed Th1/Th2 cytokine.

Animals↗