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PubMed2001

In vitro effects of isoprinosine and a dipeptide methyl ester on Echinococcus multilocularis protoscoleces.

A protoscoleces/vesicles in vitro maintenance test with assessment of viability by eosin exclusion was used to evaluate the quantitative and qualitative activities of isoprinosine, its active component inosine and the dipeptide methylester L-Phe-Phe-OMe on isolated protoscoleces of Echinococcus multilocularis for 24 and 48 h. Isoprinosine and inosine showed dose- and time-dependent activity, the latter displaying a more rapid effect than the former. A high activity was shown with L-Phe-Phe-OMe, when compared to praziquantel. Ultrastructural alterations were much more striking with L-Phe-Phe-OMe, with an effect similar to that of praziquantel, whereas the chemotherapeutic activity of inosine and isoprinosine appeared to be directed against a metabolic target, with a lethal effect not immediately visible at the ultrastructural level. Thus, the previously reported in vivo activities of these drugs result largely from a direct effect on the parasite.

Animals↗

PubMed2001

Echinococcus multilocularis in carnivores from the Klatovy district of the Czech Republic.

A unique human case of alveolar echinococcosis was described in 1979 from the Klatovy district of the Czech Republic. However, there were no previous epidemiological studies in this area focusing on detection of the source of infection--Echinococcus multilocularis adults producing eggs. During the period June 1997 to April 1999, 29 out of a total of 46 (63.3%) red foxes (Vulpes vulpes) in the Klatovy district and one of four foxes (25.0%) in the Pilsen South district were found to be infected with adult worms of E. multilocularis. No E. multilocularis adults were found in other animals from the Klatovy district (i.e. three specimens of Martes martes, two Martes foina, one Mustela erminea, two Meles meles and one Felis catus f. domestica). An examination of faecal samples from 55 dogs (Canis familiaris) from the Klatovy district resulted in the detection of E. multilocularis DNA in one (1.8%) sample. The present results support the possibility that human alveolar echinococcosis previously described in the Czech Republic had the character of an autochthonous infection. There are also indications of a potential risk of infection to humans.

Animals↗

PubMed1987

Spontaneous death of Echinococcus multilocularis: cases diagnosed serologically (by Em2 ELISA) and clinical significance.

Nine cases of asymptomatic alveolar hydatid disease (AHD) of the liver were diagnosed in 1985 among Eskimos from the endemic region of western Alaska. The patients were identified by screening with the enzyme-linked immunosorbent assay, using purified Echinococcus multilocularis antigen (Em2 ELISA). Five patients, and one diagnosed earlier (1979), were found to have lesions in which the larval E. multilocularis had died spontaneously at an early stage of infection. Viability was assessed histologically; by the avidin-biotin immunohistochemical method; and in vivo through intraperitoneal inoculation of membranes of the larval cestode into red-backed voles, Clethrionomys rutilus. The results were in agreement with the clinical impression, based on findings by computerized tomography and ultrasound scanning, and on the macroscopic appearance of the lesions, that the cestode was dead. Spontaneous death of E. multilocularis in humans has not been previously reported. The findings show that the Em2 ELISA may be positive in patients having lesions of AHD in which the etiologic agent is no longer viable.

Aged↗

PubMedDate not supplied

[Current problems of echinococcosis (Echinococcus multilocularis)].

Alveolar echinococcosis is by far the most dangerous human parasitosis in Central Europe. For the one part this is due to the fact that the parasite is localised in the liver, and for the second part it is due to the infiltrative growth of the larval cestode tissue. The life cycle of echinococcus multilocularis takes place primarily between the fox as final host and mice as intermediate hosts. In this cycle, man is an accidental host and is infected orally via the intake of cestode eggs. The individual channels of infection and their frequency have so far only been subject to speculation. The infection rate of foxes is highest in the range of the Swabian mountain pastures (known as Schwäbische Alb) (up to 55 per cent), but recently values of more than 30% have been registered in the adjacent areas as well. No accurate data are available on the infection rate in humans; seroepidemiological data indicate a prevalence between 0.1 and 0.5 per thousand. IHA, IIFT and ELISA are the most frequently used methods in serodiagnostics of echinococcosis. The specific E. multilocularis antigen should be used instead of the commercially available E. granulosus antigen when examining patients for alveolar echinococcosis. In clinical diagnostics, sonography and especially CT are well tried. If the parasitic infestation is discovered early, radical surgery is the method of choice. Chemotherapy with mebendazole (e.g. Vermox) is still unsatisfactory despite improvements. Possibly an echinococcus therapy of foxes via baits containing praziquantel may be a possibility to minimize the transfer to the human organism.

Animals↗

PubMed1997

[Occurrence of Echinococcus multilocularis Leuckart, 1863, in foxes (Vulpes vulpes) in the Czech Republic].

During the period between December 1994 and December 1996 the authors examined, using the method of helminthological dissection of the intestinal tract in 15 districts of four regions in the Czech Republic a total of 824 foxes from 350 land registers. Echinococcus multilocularis was found in the following regions: South Bohemian (districts Jindrichův Hradec, Ceský Krumlov, Prachatice and Strakonice), Central Bohemian (Benesov district) and North Bohemian (district Teplice and Chomutov). There were 87 positive (10.6%) foxes in 56 places (16%). In some areas of the South Bohemian region examinations of foxes were repeated. It was found that there are places where the prevalence of E. multilocularis varied between 28.6 and 75% (mean 53.7%) but also areas which were repeatedly negative. In 1996 after one-week intervals foxes from the Prachatice district (South Bohemian region) were sent for parasitological examination. During different months of the year the prevalence of E. multilocularis varied between 2.6 and 33.3%. A similar investigation was made during May to October 1996 (except September) also in the district of Ceský Krumlov (South Bohemian region). Tapeworms were found in 7.14 to 66.7% of foxes.

Animals↗

PubMed1998

Echinococcus multilocularis: relationship between susceptibility/resistance and liver fibrogenesis in experimental mice.

To analyze collagen and other matrix protein deposits in experimental alveolar echinococcosis as well as the expression of lysyl oxidase, the enzyme that initiates the first steps in the pyridinoline cross-linking of collagen, and to establish a relationship between resistance/susceptibility to Echinococcus multilocularis larval growth and fibrogenesis, we compared AKR/J mice (susceptible to E. multilocularis infection) with NMRI mice (resistant hosts) in this study. Collagen deposits in the lesions were evaluated using a colorimetric method; the nature of matrix proteins involved in the periparasitic fibrosis and lysyl oxidase expression were assessed using immunostaining on tissue sections. The results obtained in this sequential study confirm that fibrogenesis is an important aspect of the host immune reaction against parasitic development and that both the extent and the course of matrix protein deposition differ in the liver of susceptible and resistant mice, respectively. The long-lasting expression of alpha-actin and lysyl oxidase by host cells in NMRI mice suggests that in this resistant strain, fibrosis was not only more developed but also more highly cross-linked and, thus, less sensitive to collagenases than in susceptible mice. A very strong expression of lysyl oxidase by parasitic cells was observed in both strains of mice; the observation that E. multilocularis itself has a role in lysyl oxidase cross-linking of host collagens can be hypothesized and would be a new example of parasite-host interplay.

Actins↗

PubMed2003

Identification, molecular characterization and expression of the gene encoding the epidermal growth factor receptor orthologue from the fox-tapeworm Echinococcus multilocularis.

Receptor tyrosine kinases (RTKs) are crucially involved in the development of metazoan organisms and possible mediators of cell-cell communication that occurs between eukaryotic parasites and hosts. We have now cloned and characterized the complete complementary DNA (cDNA) molecule encoding a novel receptor tyrosine kinase, EmER, of the human parasite Echinococcus multilocularis. EmER shared significant amino acid sequence homologies with members of the epidermal growth factor (EGF) receptor family of different phylogenetic origin, exhibited a domain structure which is typical for this group of membrane receptors and contained all catalytically important residues at the corresponding positions. Highest homologies were detected between EmER and a previously identified receptor kinase, SER, from the parasitic trematode Schistosoma mansoni. The EmER encoding gene, emer, spans a chromosomal region of 39 kb and is composed of 23 exons. Structural comparisons indicated that emer and the EGF receptor encoding genes from mammals derive from a common ancestor. DNA/DNA hybridization experiments demonstrated that emer is present as a single copy locus in the parasite. Transcriptional analyses on in vitro cultivated parasite larvae revealed that emer is expressed in both the metacestode and the protoscolex stage, although about 10-fold higher transcription levels were detectable for the protoscolex. Northern blot experiments further indicated that emer is expressed as a single 5.2-kb transcript in parasitic larvae during an infection of the intermediate host. These results suggest an involvement of EmER in echinococcal differentiation processes towards the protoscolex stage during natural infections and provide, for the first time, structural information on an epidermal growth factor receptor-like kinase from a cestode.

Amino Acid Sequence↗

PubMed1988

IgE-dependent humoral immune response in Echinococcus multilocularis infection: circulating and basophil-bound specific IgE against Echinococcus antigens in patients with alveolar echinococcosis.

Clinical symptoms of immediate-type hypersensitivity (ITH) and specific IgE against Echinococcus granulosus antigens are frequently present in patients with hydatid cysts. In alveolar echinococcosis (AE) due to E. multilocularis, clinical manifestations related to ITH have never been reported. The IgE-dependent humoral immune response was evaluated in 30 patients with AE. Circulating specific IgE (sIgE) were determined with two different methods of radio-allergo-sorbent test. Serum sIgE were determined sequentially in 18 patients over 15 months. Specific IgE bound to circulating basophils were assessed with two tests in vitro, measuring specific degranulation and histamine release. The respective abilities of E. granulosus and E. multilocularis antigens to reveal bound and circulating IgE antibodies were also assayed. Despite the absence of clinical symptoms of ITH and the frequent lack of circulating sIgE, an immunological response involving IgE was always present in human AE: basophil-bound sIgE were revealed in every patient by histamine release and degranulation tests; these tests were constantly negative in control subjects. Echinococcus granulosus extracts were more effective for detecting circulating sIgE; however E. multilocularis antigenic preparation induced a histamine release significantly higher than E. granulosus extracts. These results suggest that IgE-dependent humoral immune response could play a role in the host-parasite relationship in AE. Moreover, the sensitivity of the tests used to detect basophil-bound sIgE was higher than that of the usual serological tests, and the basophil degranulation test could be used to confirm diagnosis of AE in endemic countries.

Adult↗

PubMed1994

Echinococcus multilocularis: parasite-specific humoral and cellular immune response subsets in mouse strains susceptible (AKR, C57B1/6J) or 'resistant' (C57B1/10) to secondary alveolar echinococcosis.

Parasite-specific humoral and cell-mediated immune responses were investigated in highly susceptible (AKR and C57B1/6J) and relatively resistant (C57B1/10) mice undergoing secondary alveolar echinococcosis (infection with Echinococcus multilocularis metacestode). The parasite-specific proliferative immune response of lymph node cells upon in vitro antigen stimulation remained weak in all three mouse strains. By day 30 p.i., CD4+ lymphoblast cells dominated the total population of blast cells in all three mouse strains. There was, however, an unexpectedly high proportion of CD8+ blast cells; by day 90 p.i., a marked proportional increase in CD8+ cells was seen in susceptible (AKR and C57B1/6J), but not in resistant (C57B1/10) mice. Susceptible, but not resistant mice exhibited a significantly decreased responsiveness of lymph node cells to concanavalin A (Con A) stimulation on day 90 p.i. Analysis of the humoral immune response by ELISA showed that resistance in C57B1/10 mice was associated with the ability of the host to synthesize antibodies to Em2 of the IgG3 and IgG1 isotype. Em2 is a lectin-binding carbohydrate antigen of the laminated layer. In susceptible AKR and C57B1/6J mice, low levels of anti-Em2 antibodies of the IgG2a isotype were detected. Anti-Em2 antibodies of the IgG3/IgG1 isotype, however, were absent. Differences in subclass-specific IgG responses were confirmed by immunoblot analyses. Our findings suggest that differences in antigen recognition (with respect to subsets of humoral and cellular immune components), probably controlled by non-H-2 gene(s), coupled to immune suppression modulated by CD8+ cells and/or respective cytokines, may determine susceptibility or resistance in experimental infection with E. multilocularis.

Animals↗

PubMed1995

The first report of echinococcus multilocularis in domestic dogs in Zhang County of Gansu Province.

In Zhang County of Gansu Province, 13 human cases of alveolar hydatid disease were reported up to 1990. However, the causative organism has not yet been described in animal hosts. In July 1990, postmortem examinations were made on 59 dogs (Canis familiaris) from the Caotan region of Zhang County. Of them, 6 were found to be infected with the adult worms of Echinococcus multilocularis. The morphological characteristics of the adult were compared in detail with those reported from different parts of China. Our adult specimens were closely resembling those of E. multilocularis reported from Ningxia. The role of domestic dogs in the epidemiology of alveolar echinococcosis in Zhang County was discussed.

Animals↗

PubMed2006

[Mixed infection of Echinococcus granulosus and Echinococcus multilocularis in dog].

OBJECTIVE: To identify mixed infection of Echinococcus granulosus and E. multilocularis in a dog from Xinjiang. METHODS: Thirty dogs from the pasture area were dissected and over 10,000 Echinococcus adult worms were found from one dog. Morphological observation revealed possible mixed infection of the two Echinococcus species. Further identification was made by amplification of the target gene DNA fragment (mitochondrial 12S rRNA gene). RESULTS: The adult worms of E. granulosus showed a relatively longer and larger gravid proglottid, its genital pore situated near or below the middle-side of the segment. The uterus was in a sacculate shape with irregular branches and approximately over 200 - 800 eggs in it. Morphology of the adult worms of E. multilocularis was similar to E. granulosus, slightly smaller, consisting of 4 to 5 proglottids. The uterus was not sacculate and with no branch. Its lateral genital pore often situated in the anterior part of the segment. Sequence analysis of mitochondrial 12S rRNA gene showed that amplification with the Eg1f/r primers shared complete identity with E. granulosus G1 genotype (GenBank accession no. AY462129), while that witht the EmH15/17 primers shared complete identity with E. multilocularis (GenBank accession no. AB031351). The presence of both E. granulosus and E. multilocularis was confirmed by microscopy and gene identification. CONCLUSION: Mixed infection of the two species of Echinococcus has been confirmed in the dog by morphological observation and PCR technique.

Animals↗

PubMed2004

Impeded Th1 CD4 memory T cell generation in chronic-persisting liver infection with Echinococcus multilocularis.

Memory T cells of the CD4 lineage coordinate immune responses against pathogens via the antigen-induced secretion of potent effector cytokines. The efficacy of these responses is thought to depend on both the overall number of pathogen-specific memory T cells and the particular array of cytokines that these cells are programmed to secrete. It is unknown to what extent cellular immunity can be induced by Echinococcus multilocularis infection. To examine the immunological memory provided by the adaptive cellular immune system in control of the chronic-persisting infection, peripheral lymphocytes of patients with alveolar echinococcosis (AE) were studied ex vivo. Stimulation of memory cells was performed with E. multilocularis vesicular fluid, purified protein derivative as recall antigen and phytohemagglutinin. Cytomegalovirus latency served as disease control. Frequencies of circulating CD4(+) T cells secreting IFN-gamma, IL-2, tumor necrosis factor-alpha, IL-4, IL-5 and IL-10 were determined by both cytokine flow cytometry and ELISPOT assays. Most strikingly, in chronic AE the frequencies of E. multilocularis antigen-specific cells committed to T(h)1-cytokine production were low (mean 0.5% of CD4(+) T cells). However, an E. multilocularis-specific response of CD4(+) T cells at frequencies of >/=0.1% was detected in the majority of AE patients (68%). Low numbers of cells committed to T(h)1 cytokine secretion were invariably seen in patients with active and inactive disease. Interestingly, the number of specific CD4(+) memory T cells was not increased in cured AE patients after complete surgical removal of the metacestode. Hyporesponsiveness during the chronic helminth infection was E. multilocularis specific. Thus, our results demonstrate that antigen-specific memory function against E. multilocularis is markedly different from that against viral or bacterial pathogens. Whether the antigen-specific cellular hyporesponsiveness with impeded T(h)1 CD4(+) memory T cell generation is a cause or a result of the progressive metacestode activity remains to be determined.

Adult↗

PubMed1990

In vivo morphological damage induced by a new benzimidazole prodrug in Echinococcus multilocularis metacestodes.

Benzimidazoles are the most widely used compounds in chemotherapy of alveolar hydatid disease (AHD), but long-term administration is required to reach detectable plasma levels. N-Methoxycarbonyl N'-2-nitro 4-trifluoromethyl phenyl thiourea (2) was prepared as a potential prodrug of (5-trifluoromethyl-1H-benzimidazole-2-yl)-carbamic acid methyl ester. Biological effects of 2 were evaluated in gerbils (Meriones unguiculatus) against the causative agent of AHD, Echinococcus multilocularis metacestodes, through a transmission electron microscopy study. Significant morphological damage of tegument and protoscolices occurred after an 18-day treatment. The treatment of AHD by using the prodrug form of the benzimidazole deserves further study.

Animals↗

PubMed2003

Identification and molecular characterisation of a gene encoding a member of the insulin receptor family in Echinococcus multilocularis.

Receptor kinases play a key role in the communication of cells with their environment and could be important mediators of the effects of host cytokines on endoparasitic organisms. In this paper we describe, for the first time, the characterisation of a receptor tyrosine kinase of the insulin receptor family from a parasitic helminth. Using a degenerative PCR approach, we identified and completely characterised the 5.5kb coding DNA for an Echinococcus multilocularis factor (EmIR) which displays significant homologies to insulin receptors of different phylogenetic origin. EmIR exhibited a domain structure which is typical for the protein family and contained all catalytically important residues at corresponding positions. One striking difference between EmIR and known insulin receptors was the presence of a 172 amino acid insert in the tyrosine kinase region of, as yet, unknown function. In yeast two-hybrid analyses, the ligand binding domains of the human insulin receptor and of EmIR showed comparable affinity to human insulin. The EmIR encoding chromosomal locus (emir) was characterised and comprised 16.5kb. Southern blot hybridisations demonstrated that emir is present as a single copy locus in E. multilocularis. Furthermore, structural comparisons indicated that emir and the insulin receptor genes from mammals and insects derive from a common ancestor. Based on reverse transcriptase-polymerase chain reaction analyses, emir was found to be expressed in the two larval stages metacestode and protoscolex. EmIR is, therefore, likely to play an important role in echinococcal development and possibly also in the interaction with the mammalian host.

Amino Acid Sequence↗

PubMedDate not supplied

Echinococcus multilocularis: identification and molecular characterization of a Ral-like small GTP-binding protein.

In mammals, Ral (Ras-like) GTPases have been implicated in the regulation of several cellular key processes such as oncogenic transformation, endocytosis, and actin-cytoskeleton dynamics. Here we provide, for the first time, molecular data on a Ral homologue from a parasitic helminth. We have cloned and characterized the complete cDNA molecule and the chromosomal locus encoding a novel GTP binding protein, EmRal, of the human parasite Echinococcus multilocularis. The encoded protein contained all highly conserved amino acid residues of the protein family at corresponding positions and shared significant sequence homologies with human RalA (53% identity) and RalB (54%). Upon heterologous expression of EmRal in Escherichia coli, the recombinant protein was able to bind GTP, thus indicating functionality of the Echinococcus factor. Using an in vitro prenylation assay, the purified protein was shown to be geranylgernylated, but not farnesylated, in both rabbit reticulocyte and Echinococcus cell extracts. The EmRal mRNA was found to be processed via trans-splicing and, using RT-PCR and virtual Northern blot experiments, expression of the factor could be demonstrated for the larval stages metacestode and protoscolex during an infection of the intermediate host. The data presented herein provide a solid basis for further investigations on Ras-Ral signaling mechanisms in Echinococcus.

Amino Acid Sequence↗

PubMed2005

Evaluation of the efficacy of emodepside+praziquantel topical solution against cestode (Dipylidium caninum, Taenia taeniaeformis, and Echinococcus multilocularis) infections in cats.

Emodepside+praziquantel topical solution was developed to provide broad-spectrum anthelmintic activity against gastrointestinal parasites in cats. Eight controlled studies were conducted to evaluate the efficacy of a topical solution of emodepside (3 mg/kg) and praziquantel (12 mg/kg) (Profender, BayerAG, Leverkusen, Germany) against feline infections with three species of cestodes. Studies featured naturally acquired infections of Dipylidium caninum or Taenia taeniaeformis, or experimental infections with Echinococcus multilocularis that were placebo-controlled, randomized and blinded. Cats were euthanatized and necropsied between 2 and 11 days after treatment, depending on the target parasite. The efficacy of emodepside+praziquantel topical solution was 100% against D. caninum and T. taeniaeformis, and 98.5- 100% against E. multilocularis. No significant systemic or local adverse reactions to treatment were noted in cats that received the combination. Topical treatment of cats with emodepside+praziquantel topical solution was safe and highly effective against cestode infections.

Administration, Topical↗

PubMed2000

Resistance/susceptibility to Echinococcus multilocularis infection and cytokine profile in humans. I. Comparison of patients with progressive and abortive lesions.

To clarify the role of Th1- and Th2-type cytokines in the various outcomes of human alveolar echinococcosis (AE), the cytokine immune response of self-cured patients was studied and compared with those of progressive AE patients and healthy subjects. Self-cured patients were divided into two groups according to the following clinical features: subjects who had positive Echinococcus multilocularis serologies and hepatic calcifications typical of AE were classified as 'abortive AE' patients, and those who had positive E. multilocularis serologies but no hepatic lesions or calcifications detectable by ultrasonography were classified as 'positive serology' subjects. Secretions of IL-5, IL-10 and interferon-gamma, and expression of IL-5 mRNA were evaluated in peripheral blood mononuclear cells (PBMC) stimulated in vitro with the mitogen phytohaemagglutinin-C or specific E. multilocularis antigenic preparations. The cytokine profile of abortive AE patients was the opposite of that observed in progressive AE patients. An intermediate profile was observed in positive serology subjects. PBMC from abortive AE patients, whether non-stimulated or stimulated with PHA and antigenic preparations, secreted significantly lower levels of IL-10 than those isolated from progressive AE patients. Our observations seem to confirm the regulatory role of IL-10 in the immunopathology of human AE.

Animals↗

PubMed2003

Interactions between landscape changes and host communities can regulate Echinococcus multilocularis transmission.

An area close to the Qinghai-Tibet plateau region and subject to intensive deforestation contains a large focus of human alveolar echinococcosis while sporadic human cases occur in the Doubs region of eastern France. The current review analyses and compares epidemiological and ecological results obtained in both regions. Analysis of rodent species assemblages within quantified rural landscapes in central China and eastern France shows a significant association between host species for the pathogenic helminth Echinococcus multilocularis, with prevalences of human alveolar echinococcosis and with land area under shrubland or grassland. This suggests that at the regional scale landscape can affect human disease distribution through interaction with small mammal communities and their population dynamics. Lidicker's ROMPA hypothesis helps to explain this association and provides a novel explanation of how landscape changes may result in increased risk of a rodent-borne zoonotic disease.

Animals↗