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PubMed1993

Suppression of T-cell proliferation by CD8+ T cells induced in the presence of protoscolices of Echinococcus multilocularis in vitro.

Immunoregulatory influences of protoscolices (PSC) of Echinococcus multilocularis on murine T-lymphocyte functions have been examined in an in vitro system. Proliferative responses of spleen cells stimulated with concanavalin A (ConA) or anti-CD3 monoclonal antibodies were depressed by the addition of PSC. In the presence of PSC, both interleukin-2 (IL-2) production and IL-2 receptor (IL-2R) expression by lymphocytes stimulated with ConA were significantly reduced. However, exogenous IL-2 reconstituted both the ConA-stimulated proliferative responses and IL-2R expression. These findings suggest that PSC of E. multilocularis can suppress lymphoid cell responses via influences on IL-2 production. Indeed, addition of CD(8+)-enriched cells from cultures stimulated with ConA plus PSC to fresh spleen cells showed marked suppression of the ConA responses. IL-2 production as well as IL-2R expression on the spleen cells so treated were suppressed. These findings reveal a suppressive immunologic function induced by E. multilocularis PSC that involves inhibition of IL-2 production and reduction of IL-2R expression. The PSC-induced CD8+ cells appear to play a key role in the suppressive regulation of host immune responses against E. multilocularis.

Animals↗
PubMed1996

Inhibition of human peripheral blood mononuclear cell proliferative response by glycosphingolipids from metacestodes of Echinococcus multilocularis.

The effect on human peripheral blood mononuclear cells (PBMCs) of neutral glycosphingolipids extracted from metacestodes of the parasite Echinococcus multilocularis was investigated. Neutral glycosphingolipids inhibited [3H]thymidine uptake by human PBMCs upon stimulation by mitogens such as phytohemagglutinin A and pokeweed mitogen or by allogeneic Burkitt B cells. This effect was dose dependent and was related to a decrease in interleukin 2 (IL-2) synthesis, the expression of IL-2 receptors (CD25) being unmodified. Addition of exogenous recombinant IL-2 restored the cell proliferation. Partial inhibition of immunoglobulin G (IgG), IgA, and IgM synthesis was observed in the supernatant of cell culture in association with the inhibitory effect. Identification of active subfractions contained in the neutral glycosphingolipid fraction was also studied in relation to cell viability. The free ceramide fraction had an inhibitory effect, in part related to cell lysis, particularly at high concentration, while the monogalactosylceramides had a paradoxical effect: as an activator at low concentrations and as an inhibitor at high concentrations, with limited cell survival. The immunogenic neutral glycosphingolipids containing at least two carbohydrate residues, all having a structure based on Gal beta 1-->6Gal, were inhibitors of PBMC proliferation and showed good cell survival. These results suggest that parasite neutral glycosphingolipids may play an immunologically relevant role in alveolar hydatid disease.

Animals↗
PubMed1997

Expression and immunogenicity of an Echinococcus granulosus fatty acid-binding protein in live attenuated Salmonella vaccine strains.

Fatty acid-binding proteins (FABPs) are candidate molecules for vaccines against several parasitic platyhelminths. A FABP from the cestode Echinococcus granulosus (EgDf1) was expressed in Salmonella vaccine strains as a C-terminal fusion to fragment C of tetanus toxin (TetC) by using expression vector pTECH. The fusion protein was equally expressed in several attenuated vaccine strains derived from bacteria with different genetic backgrounds and different attenuating mutations. Single-dose immunization experiments with the aroA Salmonella typhimurium strain SL3261 carrying the pTECH-EgDf1 construct were conducted with mice, using both the intravenous and the oral routes. Surprisingly, the antibody response to EgDf1 and the antigen-specific cytokine production in spleen cells were stronger in mice immunized orally. Furthermore, immune mouse sera strongly reacted with fixed sections of the worm's larval stage. Analysis of the isotype distribution of the specific anti-EgDf1 antibodies showed similar production of immunoglobulin G1 (IgG1) and IgG2a together with specific IgA antibodies. In addition, stimulation of spleen cells from mice immunized with the different constructs with either Salmonella lysate, TetC, or EgDf1 showed that, together with Th1-related cytokines (gamma interferon and interleukin 2 [IL-2]), significant levels of a Th2 cytokine (IL-5) were produced specifically, indicating a Th2 component to the response to the Salmonella carrier and to the recombinant antigens. Salmonellae expressing the TetC-rEgDfl fusion are currently under evaluation as potential vaccines against E. granulosus.

Animals↗
PubMed2001

Modulation of human immune response by Echinococcus granulosus antigen B and its possible role in evading host defenses.

By directly suppressing the function of certain immune cell subsets and by stimulating other cell populations related to immunopathology, parasite-derived substances play an important role in the chronic establishment of parasitic disease. Our objective was twofold: (i) to investigate further the role of Echinococcus granulosus antigen B (AgB) in the human early inflammatory response by determining its effect on polymorphonuclear cell (PMN) random migration, chemotaxis, and oxidative metabolism and (ii) to determine its action in acquired immunity by evaluating AgB and sheep hydatid fluid (SHF)-driven Th1 (gamma interferon [IFN-gamma] and interleukin 12 [IL-12]) and Th2 (IL-4 and IL-13) cytokine production by peripheral blood mononuclear cells (PBMC) from 40 patients who had cured or stable or progressive cystic echinococcosis. AgB significantly inhibited PMN recruitment but left their random migration and oxidative metabolism unchanged. Patients' PBMC stimulated with AgB produced IL-4 and IL-13 but did not produce IL-12. They also produced significantly lower IFN-gamma concentrations than did PBMC stimulated with SHF (P = 10(-5)). AgB skewed the Th1/Th2 cytokine ratios towards a preferentially immunopathology-associated Th2 polarization, predominantly in patients with progressive disease. AgB-stimulated patients' PBMC also proliferated less than SHF-stimulated PBMC (P = 9 x 10(-3)). In vitro Th2 cytokine production was reflected in vivo by elevated specific immunoglobulin E (IgE) and IgG4 antibodies binding to AgB. These findings confirm that AgB plays a role in the escape from early immunity by inhibiting PMN chemotaxis. They also add new information on the host-parasite relationship, suggesting that AgB exploits the activation of T helper cells by eliciting a nonprotective Th2 cell response.

Adult↗
PubMed2001

Major carbohydrate antigen of Echinococcus multilocularis induces an immunoglobulin G response independent of alphabeta+ CD4+ T cells.

Echinococcus multilocularis causes alveolar echinococcosis, one of the most lethal helminthic (accidental) infections in humans, as the life cycle predominantly includes wildlife rodents as intermediate hosts. The physical barrier between the proliferating parasitic metacestode and the host tissue is the acellular laminated layer (LL), which is characterized by its rich high-molecular-weight polysaccharide composition. Conversely to a crude protein-rich vesicular fluid antigen, a major carbohydrate antigen of the LL--the Em2(G11) antigen--did not stimulate murine T-cell proliferation in vitro. In fact, the persistent metacestode growth and antigenic stimulation induced a Th2 shift in vivo following conventional infection by intraperitoneal inoculation of 100 metacestode vesicles into C57/BL6 mice. Concurrently, the expression of Th1 cytokines (interleukin-2 and gamma interferon) remained persistently low until the late stage of chronic infection. In comparison to a recombinant proteinic II/3 antigen, the specific immunoglobulin G (IgG) response against the Em2(G11) antigen (including all IgG isotypes) maintained persistently low avidity. Furthermore, the Em2(G11) antigen induced a specific IgM and IgG response in T-cell-deficient athymic nude, TCRbeta(-/-), major histocompatibility complex class II (MHCII)(-/-)(CD4-deficient), and CD40(-/-) mice. The Em2(G11)-specific IgG synthesized in nude TCRbeta(-/-) and MHCII(-/-) mice was predominantly of the IgG3 and IgG2a isotypes and of the IgG3 and IgG2b isotypes in CD40(-/-) mice. This finding suggested that in vivo, the IgG response to major carbohydrate antigen Em2(G11) of E. multilocularis could take place independently of alphabeta+ CD4+ T cells and in the absence of CD40-CD40 ligand interactions; thus, the Em2(G11) antigen of the acellular LL represents a T-cell-independent antigen. Functionally, the encapsulating LL, and especially its major carbohydrate antigen, Em2(G11), seems to be one of the key factors in the parasite's survival strategy and acts by modulating the host immune response by virtue of its T-cell-independent nature.

Animals↗
PubMed1998

Detection of Echinococcus multilocularis in the definitive host: coprodiagnosis by PCR as an alternative to necropsy.

Recently, extensions of the range of Echinococcus multilocularis in Europe and North America and drastic increases in fox populations in Europe put an increasing proportion of the human population at risk of alveolar echinococcosis. To obtain data on the local infection pressure, studies of the prevalence of the parasite in the animals that transmit the parasite, foxes, dogs, and cats, are urgently required. Such investigations, however, have been hampered by the need for necropsy of the host animal to specifically diagnose infection with the parasite. In this study, a nested PCR and an improved method for DNA extraction were developed to allow the sensitive and specific diagnosis of E. multilocularis infections directly from diluted fecal samples from foxes. The target sequence for amplification is part of the E. multilocularis mitochondrial 12S rRNA gene. The specificity of the method was 100% when it was tested against 18 isolates (metacestodes and adult worms) of 11 cestode species, including E. granulosus. The sensitivity of the method was evaluated by adding egg suspensions and individual eggs to samples of diluted feces from uninfected foxes. The presence of one egg was sufficient to give a specific signal. To confirm the PCR results, an internal probe which hybridized only with E. multilocularis amplification products but not with the DNA of other cestodes was constructed. In order to investigate the applicability of this method for epidemiological studies, 250 wild foxes from a area in southern Germany where echinococcosis is highly endemic were examined by both necropsy and PCR of rectal contents. The sensitivity correlated with the parasites' number and stage of maturity. It ranged from 100% (>1,000 gravid worms) to 70% (<10 nongravid worms). On the basis of positive PCR results for 165 foxes, the sensitivity of the traditional and widely used necropsy method was found to be not higher than 76%. We therefore present this PCR system as an alternative method for the routine diagnosis of E. multilocularis in carnivores.

Animals↗
PubMed1989

Comparison of albendazole, mebendazole and praziquantel chemotherapy of Echinococcus multilocularis in a gerbil model.

The efficacy of albendazole (50 mg/kg/d), mebendazole (50 mg/kg/d) and praziquantel (500 mg/kg/d) against established intraperitoneal infections of Echinococcus multilocularis in gerbils was compared by monitoring parasite weight and making ultrastructural observations on treated and untreated material. Praziquantel was the most active protoscolicidal agent, reducing protoscolex viability to less than 2%, although it did not inhibit cyst growth. Albendazole was the most effective agent in reducing cyst growth and was, when compared with other regimes significantly more effective than mebendazole (p less than 0.05), praziquantel (p less than 0.01) or untreated controls (p less than 0.01).

Albendazole↗
PubMed1991

Use of sentinel lambs for early monitoring of the South Powys Hydatidosis Control Scheme: prevalence of Echinococcus granulosus and some other helminths.

Sentinel lambs were used to identify young Echinococcus granulosus infections in sheep, to provide an early indication of the progress of the South Powys Hydatidosis Control Scheme. Four sentinel lambs were purchased on each of 60 farms, from inside and outside the control area; they were examined when approximately six, 10 and 15 months of age. Gross examination, thin slicing of organs and histological examination of the lesions in the viscera revealed no E granulosus hydatid cysts in lambs born within the control area, whereas 25 per cent of the 15-month-old lambs from outside the area harboured E granulosus cysts (less than 1 to 2 mm in diameter). Lambs from E granulosus infected farms had significantly higher anti-E granulosus ELISA antibody titres than lambs from uninfected farms. It was concluded that within one year of beginning to treat dogs with praziquantel every six weeks the transmission of E granulosus to sheep had ceased. In contrast, this treatment did not prevent infections with Taenia hydatigena or T ovis; an examination of the 240 lambs revealed T hydatigena in 33.3 per cent of them, Tovis in 4.2 per cent, Dictyocaulus filaria in 12.1 per cent and Meullerius capillaris in 49.2 per cent.

Animals↗
PubMed1995

Echinococcus granulosus infections of dogs in the Durazno region of Uruguay.

The prevalence and distribution of Echinococcus granulosus in domestic dogs was examined in three dog populations in the Durazno region of Uruguay. The prevalence was 19.7 per cent in 704 dogs successfully purged with arecoline hydrobromide. Higher prevalences were detected in dogs from the rural area (30.0 per cent) and the village of La Paloma (25.9 per cent) than in the town of Sarandi del Yi (7.9 per cent). The frequency distribution of E granulosus was overdispersed (k, the negative binomial parameter = 0.08), with only a few animals harbouring heavy infections. The results of a questionnaire showed that the prevalence was greatest in male dogs, in dogs that were not kennelled, in dogs that had access to fields and in dogs that were not dosed with praziquantel. Dogs that were given raw sheep offal by their owners were no more likely to be parasitised than other dogs; this may reflect the inaccuracy of the owners' replies, or that the dogs were being infected outside their home.

Animals↗
PubMed1981

The karyotype of Echinococcus multilocularis (Cestoda: Taeniidae).

The chromosomes of Echinococcus multilocularis Leuckart, 1863 have been described, based on components of cells from the adult stage and the larval stage of the cestode. Chromosomal complements of 2n = 18, n = 9 were confirmed. The fundamental number of chromosomal arms were 24. Karyograms are presented, in which the 18 elements are grouped according to arm-ratios and size. An idiogram of G-bands is included.

Animals↗
PubMed1990

Perioperative prophylactic chemotherapy of Echinococcus granulosus: determination of minimum effective length of albendazole therapy in in vitro protoscolex culture.

Protoscoleces of Echinococcus granulosus were cultured in vitro in 500, 250 or 100 micrograms/l albendazole sulphoxide for 1,3,7,10,14d and then 'rescued' (R) into drug-free medium for the remainder of the culture period. Successful minimum lengths of therapy were much longer than for praziquantel, and only at 500 micrograms/l was the 10dR treatment as effective as continuous therapy for 28d. Treatment with 100 micrograms/l both in continuous culture and in the 'R' experiments was ineffective over a 35d period. The results are compared with those from similar experiments using praziquantel.

Albendazole↗
PubMed1992

Diagnostic value of a synthetic peptide derived from Echinococcus granulosus recombinant protein.

A specific monoclonal antibody (MAb; EG 02 154/12) directed against a protein epitope of Echinococcus granulosus antigen 5 was used to screen a cDNA library constructed from E. granulosus protoscoleces RNA. One clone designated Eg14 was selected and shown to code for an amino acid sequence partially homologous to that of the clone Eg6 previously identified with the same MAb. Hydrophobic cluster analysis showed that both recombinant antigens may adopt a similar alpha-helical organization and share a common conformational epitope. A synthetic peptide (89-122) mimicking the conformational site of Eg6 and Eg14 was constructed and demonstrated to be able to inhibit binding of the MAb and human hydatid sera to the Eg6 fusion protein (FP6) or to native hydatid antigens. To assess the diagnostic value of the peptide 89-122, we tested sera from patients infected with different parasites for their antibody reactivity with this peptide in ELISA. A high binding sensitivity and specificity of IgG-A-M antibodies were obtained with E. granulosus-infected patient sera. Moreover, the peptide 89-122 was found to be specifically recognized by IgE antibodies from patients with hydatid disease. These results indicate the particular interest of this synthetic peptide as a standardized antigen in diagnosis and treatment surveillance of hydatidosis.

Amino Acid Sequence↗
PubMed2005

Detection of Echinococcus multilocularis infection in a colony of cynomolgus monkeys (Macaca fascicularis) using serology and ultrasonography.

Five animals in a colony of cynomolgus monkeys (Macaca fascicularis) died or were euthanatized because of alveolar echinococcosis, during a period of 5 years. The remainder of the colony was screened for possible infection with Echinococcus multilocularis, using serology and ultrasonography. A total of 46 animals out of a group of 55 were examined. The presence of anti-Em2 antibodies analyzed with enzyme-linked immunosorbent assay was demonstrated in 3 monkeys. In 2 of these 3 monkeys, multilocular structures compatible with metacestodal cysts in the liver were identified, using ultrasonography. The presence of alveolar echinococcosis was subsequently confirmed at postmortem examination in 1 animal. The other animals are still alive. Two other monkeys were negative in the serological examination but had cystic structures in the liver, which were identified as bile duct cysts at postmortem examination in 1 animal. The other monkey is still alive. These findings suggest that serology for antibodies against the Em2 antigen may represent a useful method in identifying animals that might be infected with E. multilocularis and are therefore at risk of developing fatal alveolar echinococcosis.

Animals↗
PubMed2002

Epidemiology and strain characteristics of Echinococcus granulosus in the Benghazi area of eastern Libya.

The incidence of surgically confirmed cystic echinococcosis in eastern Libya was estimated to be at least 4.2 cases/100,000, with significantly more female cases than male. The prevalences of infection with Echinococcus granulosus among 1087 sheep, 881 goats, 428 camels and 614 cattle from the same region, determined postmortem in abattoirs, were 20%, 3.4%, 13.6% and 11%, respectively. Infection in the livestock was age-dependent and, generally, the female animals were more often infected than the male. The measurements of rostellar hooks on protoscoleces collected from sheep and cattle were similar but significantly different from the corresponding measurements of parasites of human or camel origin. However, when a portion of the cytochrome c-oxidase subunit I (cox1) gene from each of 30 protoscolex samples from Libya (12 from cattle, three from humans, five from camels and 10 from sheep) was sequenced, the sequences were all found to be identical to that published for the common sheep strain of E. granulosus.

Adolescent↗
PubMed2004

Using morphometry of the larval rostellar hooks to distinguish Iranian strains of Echinococcus granulosus.

Human cystic echinococcosis is widely distributed throughout Iran, where sheep, cattle, goats, camels and other animals act as intermediate hosts. It appears that morphometry of the rostellar hooks on the protoscoleces of the causative parasite, Echinococcus granulosus, can be used to separate the strains or variants of the parasite to be found in humans and livestock in Iran. Multivariate statistical procedures, including principal-component analysis and discriminant-function analysis (DFA), were used to explore the morphometric data obtained from the larval hooks of Iranian samples of E. granulosus. Although five physical variables were initially considered, the results of the PCA indicated that just two factors (based on the length of hooks and number of hooks) accounted for 91.6% of the variance observed in the hook measurements. The results of the DFA allowed the correct classification of all the samples. The presence in Iran of the sheep and camel strains of the parasite was confirmed. The sheep strain was the most common variant encountered, being recovered from sheep, cattle, goats, humans and occasionally camels. Most of the metacestode samples from camels and a few of those from cattle (five of 30), sheep (one of 27) and humans (two of 23) were found to be of the camel strain. These results indicate that larval-hook morphology is a valid method for identifying E. granulosus strains in Iran, and one that is potentially useful for epidemiological studies.

Animals↗
PubMed1994

Echinococcus multilocularis: effect of size of inoculum and route of infection on metastasis, amyloidogenesis and alveolar hydatid cysts mass in mice.

Alveolar hydatid disease (AHD) in mice, caused by Echinococcus multilocularis, is characterized by restrictive and metastasizing progressive growth phases. In experimentally induced infections, neither inoculum size (5, 50 or 250 viable cysts) nor the route (intraperitoneal/subcutaneous) of infection altered the course of disease as measured by the size of the larval cyst mass (LCM) produced. Spleen weight and amyloid deposition were also shown to be independent of the route or size of inoculum. Inoculation of a soluble parasite protein extract (AHC-EXT) induced amyloid deposition, with a dose-dependent threshold. These results support our postulate that soluble component(s) of the LCM are the major factor in the pathogenesis of AHD.

Amyloid↗
PubMed1998

Coproantigen detection in a survey of Echinococcus multilocularis infection among red foxes, Vulpes vulpes schrencki, in Hokkaido, Japan.

Detection of Echinococcus coproantigen using sandwich enzyme-linked immunosorbent assay (sELISA) was performed on fecal samples of red foxes in Hokkaido, Japan. Fecal samples were collected around fox dens in 1990 and 1992. The antibodies used for sELISA recognize heat-resistant antigens, thus all fecal samples were heated to render it safe for handling before examination. Detection of taeniid egg in fox feces collected was considered as an indication of E. multilocularis infection. In fecal samples collected in 1990 and 1992, coproantigen positive results out of taeniid-egg positive cases were 38/40 (95.0%) and 95/97 (97.9%), respectively. In addition, coproantigen was detected regardless of fecal condition when collected from the field, suggesting that the antigens detected by this method are quite stable. These results suggest that detection of coproantigen is useful for field surveys of foxes naturally infected with E. multilocularis.

Animals↗
PubMed2005

Outbreak of larval Echinococcus multilocularis infection in Japanese monkey (Macaca fuscata) in a zoo, Hokkaido: western blotting patterns in the infected monkeys.

A high prevalence of larval Echinococcus multilocularis (Em) infection was found in zoo primates in Hokkaido, Japan. In October 1997, a Japanese monkey (Macaca fuscata) died and histopathologically diagnosed as alveolar hydatidosis. Serum samples were collected from the remaining Japanese monkeys and examined for antibodies against Em by enzyme-linked immunosorbent assay and western blotting. Serological tests showed 12 more animals of the remaining 57 monkeys were possibly infected. Ultrasonography revealed that nine of these 12 animals had a cystic lesion in the liver. The band patterns of western blotting in the monkeys were very similar to those in human.

Animals↗