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Cystic Echinococcosis (CE) is a neglected zoonotic disease caused by the metacestode form of Echinococcus granulosus sensu lato. Non-invasive imaging techniques, especially ultrasound, are primarily used for CE diagnosis. MicroRNAs (miRNAs) are small, non-coding RNA molecules that act as post-transcriptional regulators in various biological processes. After identification of parasite-derived miRNAs, these miRNAs are considered to be potential biomarkers for diagnosis and follow-up. The focus of this research is to compare the expression profiles of certain parasite-derived miRNAs in CE patients with active and inactive cysts as well as healthy controls. Parasite-derived miRNAs, egr-let-7-5p, egr-miR-71a-5p, and egr-miR-9-5p, of inactive CE patients were found to be differentially expressed with 3.74-, 2.72-, and 20.78-fold change (p

Hydatid disease has a great impact on public health, causing high morbidity and mortality. Main lines of treatment include surgery, which mostly requires the installation of a scolicidal agent into hydatid cysts to prevent dissemination. Alternatively, medical treatment involves the use of benzimidazole drugs; however, the results are not satisfactory, and new drug compounds are urgently needed. Fluralaner is a potent inhibitor of GABA-gated chloride channels and L-glutamate-gated chloride channels (GluCls) providing immediate and persistent flea, tick and mite control in dogs after a single oral dose. Researches previously identified different genes encoding ion channels in Echinococcus granulosus, making ion channel inhibitors a promising target for treating hydatid disease. Thus, the present study aimed to evaluate the effect of fluralaner on protoscolices and metacestode layers. Parasite materials (Protoscolices, Metacestodes layers) were exposed to different concentrations of the drug ranging from “12.5–100 ug/ml” and examined for viability after 1, 6 and 24 h. Morphological and ultrastructural alterations were recorded by both light and electron microscopies. Immunohistochemical staining confirmed caspase-3 activation as an indicator of apoptosis- induced therapy. The treated protoscolices and metacestode layers showed loss of the viability, the formation of vacuoles and lipid droplets, separation of the germinal layer, and damage in the laminated layer; apoptosis was prominent after treatment. These findings revealed that fluralaner has a potent scolicidal activity and suggested its therapeutic potential against hydatid disease. Further evaluations for animals and human use in the treatment and prevention of hydatid disease are needed.

BackgroundAlveolar echinococcosis (AE) is a chronic zoonosis caused by the larval form ofEchinococcus multilocularis(E. multilocularis). Current chemotherapy against AE has relied on albendazole and mebendazole, which only exhibit parasitostatic and not parasiticidal efficacy. Therefore, novel compounds for the treatment of this disease are needed.MethodsPhosphoglucose isomerase (PGI) assays were used for compound screening of seven neonicotinoids. The anti-parasitic effects of thiacloprid were then evaluated onE. multilocularismetacestode vesicles, germinal cells and protoscoleces in vitro. Human foreskin fibroblasts (HFF) and Reuber rat hepatoma (RH) cells were used to assess cytotoxicity. Glucose consumption inE. multilocularisprotoscoleces and germinal cells was assessed by measuring uptake of 2-deoxyglucose (2-DG). Molecular docking was used to evaluate the potential binding sites of thiacloprid to acetylcholine receptors. In vivo efficacy of thiacloprid was evaluated in mice by secondary infection withE. multilocularis. In addition, ELISA and flow cytometry were used to evaluate the effects of cytokines and T lymphocyte subsets after thiacloprid treatment. Furthermore, collagen deposition and degradation in the host lesion microenvironment were evaluated.ResultsWe found that thiacloprid is the most promising compound, with an IC50of 4.54 ± 1.10 μM and 2.89 ± 0.34 μM, respectively, against in vitro-culturedE. multilocularismetacestodes and germinal cells. Thiacloprid was less toxic for HFF and RH mammalian cell lines than for metacestodes. In addition, thiacloprid inhibited the acetylcholinesterase activity in protoscoleces, metacestodes and germinal cells. Thiacloprid inhibited glucose consumption by protoscoleces and germinal cells. Subsequently, transmission electron microscopy revealed that treatment with thiacloprid damaged the germinal layer. In vivo, metacestode weight was significantly reduced following oral administration of thiacloprid at 15 and 30 mg/kg. The level of CD4+T lymphocytes in metacestodes and spleen increased after thiacloprid treatment. Anti-echinococcosis-related cytokines (IL-2, IL-4, IL-10) were significantly increased. Furthermore, thiacloprid inhibited the expression of matrix metalloproteinases (MMPs 1, 3, 9, 13) and promoted collagen deposition in the host lesion microenvironment.ConclusionsThe results demonstrated that thiacloprid had parasiticidal activity againstE. multilocularisin vitro and in vivo, and could be used as a novel lead compound for the treatment of AE.Graphical abstract

Background Cystic echinococcosis (CE)/hydatidosis is an important neglected parasitic zoonotic disease caused by the metacestode of Echinococcus granulosus s.l. The present study was designed to identify the pulmonary CE species/genotypes in isolated human underwent to surgery in our center in Southern Iran. Methods The study population of this study were all patients in Fars province who were admitted to Namazi Hospitals for pulmonary hydatid cyst surgery. Thoracic surgery was performed in the thoracic ward and the cyst/s was removed by open surgery via posterolateral or lateral thoracotomy. DNA was extracted from the germinal layer or the protoscoleces. PCR technique was performed using the cytochrome C oxidase subunit1 (cox1) gene, and the products were sequenced. Results A total of 32 pulmonary hydatid cyst samples were collected from 9 (28%) female and 23 (72%) male aged from 4 to 74 years old. A total of 18(56%) cyst/s were in the left lobe and 14 (44%) cysts in the right lobe. Sequence analysis of the cysts showed that 24 samples (75%) were E. granulosus s.s (G1-G3) genotype and 8 (25%) were E. canadensis (G6/G7) genotype. Conclusion E.granulosus s.s genotype was the most prevalent genotype followed by E. canadensis (G6/G7) genotype. There was no significant statistical correlation between cysts’ size, location, genotype strain, and patients’ age and gender.

Cystic echinococcosis is a zoonotic disease caused by the metacestode of Echinococcus granulosus sensu lato. The disease is characterized by the development of cystic structures inside viscera of the intermediate host, mainly liver and lungs. These cysts are formed by three layers: germinal, laminated, and adventitial layer, the latter being the local host immune response. Metacestodes that develop protoscoleces, the infective stage to the definitive host, are termed fertile, whereas cysts that do not produce protoscoleces are termed non-fertile. Sheep usually harbor fertile cysts while cattle usually harbor non-fertile cysts. Adventitial layers with fibrotic resolution are associated to fertile cysts, whereas a granulomatous reaction is associated with non-fertile cysts. The aim of this study was to analyze cellular distribution in the adventitial layer of fertile and non-fertile E. granulosus sensu stricto cysts found in liver and lungs of cattle and sheep. A total of 418 cysts were analyzed, 203 from cattle (8 fertile and 195 non-fertile) and 215 from sheep (64 fertile and 151 non-fertile). Fertile cysts from cattle showed mixed patterns of response, with fibrotic resolution and presence of granulomatous response in direct contact with the laminated layer, while sheep fertile cysts always displayed fibrotic resolution next to the laminated layer. Cattle non-fertile cysts display a granulomatous reaction in direct contact with the laminated layer, whereas sheep non-fertile cysts display a granulomatous reaction, but in direct contact with the fibrotic resolution. This shows that cattle and sheep cystic echinococcosis cysts have distinct local immune response patterns, which are associated to metacestode fertility.

The article examines the presence of metacestodes on the liver capsule in two wild boars and on the liver capsule and mediastinum in moose. Cysticerci were identified as Taenia hydatigena metacestodes by morphological features—size and shape of rostellar hooks and molecular analysis of the partial sequences of cytochrome c oxidase subunit 1 gene. Molecular analysis revealed similarities between the metacestodes isolated from the wild mammals in the present study to T. hydatigena in European wolves. It is possible that infection of moose and wild boar with T. hydatigena metacestode is related to the growth in the wolf population in Europe. Electronic supplementary material The online version of this article (10.1007/s10393-019-01392-9) contains supplementary material, which is available to authorized users.

Taenia multiceps, one of the most widely distributed zoonotic tapeworm parasites, is able to parasitize the small intestine of canids. The metacestode of T.multiceps is fatal to ruminants and causes important economic losses in livestock. However, molecular characteristics of T.multiceps and coenurus in China are still unclear. In this study, 36 goat isolates of the coenurus stage and 18 dog isolates of the adult stage of T.multiceps were obtained from three geographical areas in China and the isolated parasite above were analyzed by amplifying the partial of cytochrome coxidase subunit 1(pcox1), 12S ribosomal RNA (12S rRNA) from mitochondrial DNA (mtDNA) regions and an internal transcribed spacer (ITS) of ribosomal DNA (rDNA). These DNA sequences obtained from T.multiceps and coenurus were employed to evaluate the nucleotide diversity and confirm the relationship between T.multiceps and coenurus. Sequences variation were 0–1.4%, 0–1.5%, 0–4.2% for pcox1, 12S rRNA and ITS, respectively, among T.multiceps and coenurus isolates obtained in this study. In Sichuan province, sequence variations for Coenurus cerebralis isolated from Yaan city were 0–1.4% for pcox1, 0–1.0% for 12S rRNA and 0–2.1% for ITS. In Hunan province, variations were 0–1.0%, 0–1.5% and 0–3.3% for corresponding genes for non-coenurus cerebralis isolated from Changsha city, while variations of T.multiceps isolates from Xiangxi autonomous prefecture were 0–1.0%, 0–1.1% and 0–3.4% for pcox1, 12S rRNA and ITS, respectively. Phylogenetic analysis based on pcox1 sequences indicated that all cerebral and noncerebral metacestodes belong to T.multiceps. These results provide reference values for future molecular epidemiological and biological study on T.multiceps in dogs and intermediate hosts.

Porcine cysticercosis, caused by metacestodes of Taenia solium is an important emerging zoonotic disease with public health and economic significance. Pigs acquire the disease through consumption of Taenia solium eggs excreted by human tapeworm carriers. The present study was conducted to investigate the prevalence of porcine cysticercosis in Nagpur and Mumbai region of Maharashtra, India by P/M examination of carcasses followed by histopathology of affected organs in infected animals and molecular identification of cysts for confirmation. Out of 1000 pigs examined during slaughter, three pigs were found to be heavily affected with T. solium cysts giving a prevalence of 0.3%. Histological section of brain in infected animals revealed marked vascular congestion of meninges, mild neuronal degeneration, perivascular cuffing and gliosis while the liver showed the infiltration of mononuclear cell, predominantly eosinophils throughout the parenchyma. Some degree of calcification was observed in the cysts lodged in liver while calcification was not evident in case of cysts lodged in brain, tongue, diaphragm and skeletal muscle. Molecular identification by PCR using two sets of oligonucleotide primers against LSU rRNA gene and Mt-Cox1 gene of T. solium confirms the cysts to be that of T. solium. The molecular diagnostics methods have been considered for validation in conjunction with P/M inspections, parasitological and histopathological examinations. The study confirms the presence of porcine cysticercosis in the two regions and demands proper sanitary measures to minimize the risk of infection from zoonoses and food safety point of view.

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Background Serotonin is a phylogenetically ancient molecule that is widely distributed in most metazoans, including flatworms. In addition to its role as a neurotransmitter, serotonin acts as a morphogen and regulates developmental processes. Although several studies have focused on the serotonergic nervous system in parasitic flatworms, little is known on the role of serotonin in flatworm development. Methods To study the effects of serotonin on proliferation and development of the cestode Echinococcus multilocularis, we cloned the genes encoding the E. multilocularis serotonin transporter (SERT) and tryptophan hydroxylase (TPH), analyzed gene expression by transcriptome analysis and whole mount in situ hybridization (WMISH) and performed cell culture experiments. Results We first characterized orthologues encoding the SERT and TPH, the rate-limiting enzyme in serotonin biosynthesis. WMISH and transcriptomic analyses indicated that the genes for both SERT and TPH are expressed in the parasite nervous system. Long-term treatment of parasite stem cell cultures with serotonin stimulated development towards the parasite metacestode stage. Mature metacestode vesicles treated with serotonin showed increased rates of incorporation of the thymidine analogue 5-ethynyl-2′-deoxyuridine (EdU), indicating stimulated cell proliferation. In contrast, treatment with the selective serotonin reuptake inhibitor paroxetine strongly affected the viability of parasite cells. Paroxetine also caused structural damage in metacestode vesicles, suggesting that serotonin transport is crucial for the integrity of parasite vesicles. Conclusions Our results indicate that serotonin plays an important role in E. multilocularis development and proliferation, providing evidence that the E. multilocularis SERT and TPH are expressed in the nervous system of the protoscolex. Our results further suggest that the E. multilocularis SERT has a secondary role outside the nervous system that is essential for parasite integrity and survival. Since serotonin stimulated E. multilocularis metacestode development and proliferation, serotonin might also contribute to the formation and growth of the parasite in the liver. Graphical Abstract

Despite the wide distribution of wild boar populations in Italy and the increase of its diffusion in urbanized areas, only one case report has described the occurrence of Echinococcus granulosus s.l. in a wild boar from Marche (Central Italy). The present study investigated the presence of E. granulosus sensu lato with an epidemiological survey on wild boars from central Italy that had been killed during hunting season. Seven hundred sixty-five (765) adult wild boars were examined during the 2016-2017 hunting season. Of these animals, 1.0% (8/765) were positive to E. granulosus s.l. with a fertility of 0.3% (2/765), and 2.9% animals (22/765) were positive for the metacestode stage of Taenia hydatigena (Cysticercus tenuicollis), while 0.5% (4/765) showed mixed infection (E. granulosus s.l. + T. hydatigena). Sixteen hydatids were found, of which 12.5% were fertile, 37.5% were sterile, 31.3% were caseous, and 18.8% were calcified. Eight hydatids (two fertile and six sterile cysts) were molecularly characterized by analysis of the mitochondrial gene, cytochrome c oxidase subunit 1 (cox1), and the NADH dehydrogenase subunit 1 (ND1). Hydatids found in wild boars were characterized as E. granulosus sensu stricto (G1 genotype). The present survey represents the first epidemiological study on cystic echinococcosis in wild boar in Italy which highlights the need for more extensive epidemiological investigations to determine the causal factors, economic impact, and public health importance of the disease in this livestock-wildlife setting.

Cysticercus tenuicollis is the metacestode of canine tapeworm Taenia hydatigena, which has been reported in domestic and wild ruminants and is causing veterinary and economic losses in the meat industry. This study was conducted to determine the sequence variation in the mitochondrial cytochrome c oxidase subunit 1 (coxl) gene in 20 isolates of T. hydatigena metacestodes (cysticercus tenuicollis) collected from northern West Bank in Palestine. Nine haplotypes were detected, with one prevailing (55%). The total haplotype diversity (0.705) and the total nucleotide diversity (0.0045) displayed low genetic diversity among our isolates. Haplotype analysis showed a star-shaped network with a centrally positioned common haplotype. The Tajima’s D, and Fu and Li’s statistics in cysticercus tenuicollis population of this region showed a negative value, indicating deviations from neutrality and both suggested recent population expansion for the population. The findings of this study would greatly help to implement control and preventive measures for T. hydatigena larvae infection in Palestine.

Cystic echinococcosis (CE), which is caused by the metacestode of Echinococcus granulosus, is one of the most important zoonoses affecting humans. Benzimidazoles (in particular albendazole) and praziquantel (PZQ) are effective against CE, but poor water solubility of these agents often leads to inadequate results. Here, we evaluate the effects of chitosan-albendazole (ChABZ) and chitosan-praziquantel (ChPZQ) nanoparticles as a new formulation on hydatid cysts both in vitro and in vivo. Developed microcysts in culture were treated with different concentrations of ChABZ and ChPZQ nanoparticles (either alone or in combination), and ABZ + PZQ suspension. The viability rate of microcysts was used to evaluate the drug efficacies. In addition, the prophylactic and therapeutic effects of the drugs were studied on infected DBA/2 mice. Transmission electron microscopy was used to observe the ultra-structural changes. The viability rate of microcysts and differences in cyst weights were compared by ANOVA, and the cyst numbers were compared using the Kruskal-Wallis test. The combination of ChABZ + ChPZQ nanoparticles was more effective than the ABZ + PZQ suspension in vitro (p

Coenurus cerebralis, the larval stage (metacestode or coenurus) of Taenia multiceps, parasitizes sheep, goats, and other ruminants and causes coenurosis. In this study, we isolated and characterized complementary DNAs that encode lactate dehydrogenase A (Tm-LDHA) and B (Tm-LDHB) from the transcriptome of T. multiceps and expressed recombinant Tm-LDHB (rTm-LDHB) in Escherichia coli. Bioinformatic analysis showed that both Tm-LDH genes (LDHA and LDHB) contain a 996-bp open reading frame and encode a protein of 331 amino acids. After determination of the immunogenicity of the recombinant Tm-LDHB, an indirect enzyme-linked immunosorbent assay (ELISA) was developed for preliminary evaluation of the serodiagnostic potential of rTm-LDHB in goats. However, the rTm-LDHB-based indirect ELISA developed here exhibited specificity of only 71.42% (10/14) and sensitivity of 1:3200 in detection of goats infected with T. multiceps in the field. This study is the first to describe LDHA and LDHB of T. multiceps; meanwhile, our results indicate that rTm-LDHB is not a specific antigen candidate for immunodiagnosis of T. multiceps infection in goats.

Background Taenia multiceps is a harmful tapeworm and its larval form (coenurus cerebralis) is the causative agent of coenurosis, a disease affecting the health of herbivores, resulting in great economic loss to animal husbandry. Heat-shock proteins (HSPs), expressed in all prokaryotes and eukaryotes, act as molecular chaperones and can affect pathogenicity. Methods Herein, cDNAs of T. multiceps genes Tm-HSP60 and Tm-p36 were cloned and molecularly characterised by bioinformatics analyses. The immunogenicity and immunoreactivity of recombinant rTm-HSP60 and rTm-p36 proteins were investigated by immunoblotting and indirect ELISA was established to evaluate their serodiagnostic potential. Tissue localisation and transcriptional level at different life stages of T. multiceps were determined by immunohistochemical and quantitative real-time PCR analyses. Result The 533 residue rTm-HSP60 and the 314 residue rTm-p36 proteins share typical highly conserved features of HSPs. Tm-p36 shares structural characteristics with metazoan small HSPs, with two N-terminal α-crystallin domains. Compared with Tm-p36, Tm-HSP60 displayed stronger immunogenicity, and the indirect ELISA based on rTm-HSP60 exhibited a sensitivity of 83.3% and a specificity of 87.5%, while rTm-p36 was not suitable to develop indirect ELISA. Tm-HSP60 was widely distributed in all stages of T. multiceps, albeit at relatively low levels, while Tm-p36 was specifically distributed in the protoscolex and oncosphere. Conclusions The sequence, structural and functional analyses of these two HSPs indicates that they may play important roles in the life-cycle of T. multiceps as molecular chaperones. Tm-HSP60 displayed stronger immunogenicity compare to Tm-p36, and has the potential for antibody detection. Tm-p36 was strongly associated with the activation of oncospheres and has potential interest for vaccination. Electronic supplementary material The online version of this article (10.1186/s13071-019-3352-8) contains supplementary material, which is available to authorized users.

International audience; Echinococcosis is a zoonotic infection caused by cestode species of the genus Echinococcus, with limited treatment options. It is urgent to develop new anti-hydatid agent. In this paper, we reported anacardic acid (AA), a natural product isolated from the Brazilian cashew-nut shell liquid, which presented a high activity against metacestodes of Echinococcus multilocularis (E. multilocularis) and Echinococcus granulosus sensu stricto (E. granulosus s.s.) in vitro and in vivo. AA exerted a better efficacy on E. granulosus s.s. protoscoleces and E. multilocularis metacestodes than that of albendazole (ABZ) and dihydroartemisinin (DHA) in vitro, and an inhibition on the growth of Echinococcus metacestode as effective as ABZ in vivo. Moreover, we also found that one of the mechanisms of AA against Echinococcus could be the suppression of angiogenesis on/in the metacestode mass through inhibiting vascular endothelial growth factor (VEGF)—induced signalling pathways. This work finds that AA is a new promising potential candidate drug for echinococcosis treatment.

Fish constitute a major component of diet for the people of Northeast India and they are extensively used as a protein-rich food for human consumption. The present studies incorporate the spectrum, composition and diversity of the parasitic species in freshwater fishes in Meghalaya, Northeast India, with a view to identifying the species recovered by morphological criteria based on light microscopy. The collection sites included sixteen foci from eleven districts of Meghalaya for parasites occurring in the common food fishes (Cypriniformes, Channiformes, Silurformes, Symbranchiformes and Anguilliformes). The helminth parasite spectrum recovered from the various piscine host species in the study area comprised of a total of 19 taxa: 2 monogenean, 8 trematode (4 adult and 4 metacercarial stages), 12 cestode (11 adult and a metacestode stage), 6 nematode (3 adult and 3 larval stages) and a single acanthocephalan species. A checklist of the parasite species with short remarks for each is provided herein.

Bovine cysticercosis is detected during the routine post mortem examination of carcasses by visual inspection (knife and eye method). However, the sensitivity of this procedure is several times lower than immunoassays, even when it is performed by qualified professionals. In the present study, a new generation capture antigens were screened from a phage display peptide library using antibodies from Taenia saginata-infected animals. Eight phage clones were selected, and one, Tsag 3 (VHTSIRPRCQPRAITPR), produced similar results to the T. saginata metacestode crude antigen (TsCa) when used as a capture antigen in an ELISA. The phage-displayed peptides competed with TsCa for binding sites, reducing the reactivity by approximately 30 %. Alanine scanning indicated that proline, arginine, and serine are important residues for antibody binding. Tsag 1 (HFYQITWLPNTFPAR), the most frequent affinity-selected clone, and Tsag 6 (YRWPSTPSASRQATL) shared similarity with highly conserved proteins from the Taeniidae family with known immunogenicity. Due to their epitopic or mimotopic properties, these affinity-selected phages could contribute to the rational design of an ante mortem immunodiagnosis method for bovine cysticercosis, as well as an epitope-based vaccine to interrupt the taeniosis/cysticercosis complex.

Present study was undertaken to estimate the prevalence, cyst viability, organ distribution and economic importance of major metacestodes in sheep and goats at Dessie municipal abattoir during the period between November 2011 and March 2012. Visceral organs in 930 slaughtered small ruminants (510 sheep and 420 goats) were examined during the study period, and economic losses were estimated based on the condemnation of organs infested by metacestodes. Out of 510 randomly selected slaughtered sheep, 45.69, 8.43 and 9.02 % were infected with Cysticercus tenuicollis, Cysticercus ovis and hydatid cysts, and of 420 randomly selected slaughtered goats, 72.38, 8.57, 1.90 % were infected with C. tenuicollis, C. ovis and hydatid cysts, respectively. In slaughtered sheep, the major organs infested with C. tenuicollis, C. ovis and hydatid cysts were the omentum (33.92 %), heart (6.78 %) and lung (4.31 %), respectively. Similarly in goats, the omentum (34.52 %), heart (1.90 %) and lung (0.95 %) were the most infested organs with C. tenuicollis, C. ovis and hydatid cysts, respectively. The only major risk factor observed was for C. tenuicollis prevalence which varied significantly with age both in sheep (P

Neurocysticercosis (NC) is one of the most important diseases caused by parasites affecting the central nervous system. We fractionated by ion-exchange chromatography using diethylaminoethyl (DEAE)-sepharose resin the total saline extract (S) from Taenia solium metacestodes and evaluated obtained fractions (DEAE S1 and DEAE S2) by enzyme-linked immunosorbent assay (ELISA, n = 123) and immunoblotting (IB, n = 22) to detect human NC in serum. Diagnostic parameters were established by ROC and TG ROC curves for ELISA tests. IB was qualitatively analyzed. S and DEAE S1 presented sensitivity of 87. 5 % and DEAE S2 90 %. The best specificity was observed for DEAE S2 (90.4 %). In IB, using DEAE S2 samples from NC patients presented bands of 20–25, 43–45, 55–50, 60–66, 82, 89, and 140 kDa. The great diagnostic parameters reached by DEAE S2 suggest the potential applicability of this fraction in NC immunodiagnosis.

Background Coenurosis is caused by coenurus, the metacestode of Taenia multiceps, which mainly parasitizes the brain and spinal cord of cattle, sheep and goats. To date, no widely-approved methods are available to identify early coenurus infection. Methods In this study, we identified a full-length cDNA that encodes GP50 (TmGP50) from the transcriptome of T. multiceps, and then cloned and expressed in E. coli. The native proteins in adult stage and coenurus were located via immunofluorescence assays, while the potential of recombinant TmGP50 protein (rTmGP50) for indirect ELISA-based serodiagnostics was assessed using native goat sera. In addition, we orally infected 20 goats with mature T. multiceps eggs. Praziquantel (10%) was given to 10 of the goats 45 days post-infection (p.i.). Blood samples were collected for 17 weeks p.i. from the 20 goats and anti-rTmGP50 antibodies were evaluated using the indirect ELISA established here. Results The TmGP50 contains an 897 bp open reading frame, in which signal sequence resides in 1 ~ 48 sites and mature polypeptide consists of 282 amino acid residues. Immunofluorescence staining showed that native TmGP50 was localized to the microthrix and parenchymatous zone of the adult parasite and coenurus, and the coenurus cystic wall. The indirect ELISA based on rTmGP50 exhibited a sensitivity of 95.0% and a specificity of 92.6% when detecting GP50 antibodies in sera of naturally infected goats and sheep. In goats experimentally infected with T. multiceps, anti-TmGP50 antibody was detectable from 2 to 17 weeks p.i. in the control group, while the antibody fell below the cut-off value about 3 weeks after praziquantel treatment. Conclusion Our results indicate that recombinant TmGP50 is a suitable early diagnostic antigen for coenurus infection in goats.

Summary We report a rare case of a cardiac hydatid cyst that was incidentally found during routine work up for a redo-CABG and was picked up on echocardiography and confirmed by magnetic resonance imaging and, after successful removal, further confirmed by histopathology. The report emphasizes the importance of early and urgent surgery for such cardiac hydatid cysts whenever discovered to prevent fatal and unexpected death. Cardiac hydatidosis is a most infrequent type, in comparison with hydatidosis of the liver (65%) and lung (25%). Learning points: † Hydatidosis or cystic echinococcosis is caused by infection with the metacestode stage of the tapeworm Echinococcus (family Taeniidae). The adult tapeworm is usually found in dogs or other canines; the tapeworm eggs are expelled in the animal’s feces and humans become infected after ingestion of the eggs. The initial phase of primary infection is asymptomatic. † Cardiac hydatidosis is extremely rare, more commonly the liver and lungs are affected. † Morbidity from heart echinococcosis in men is three times higher than that in women. Solitary cysts occur in almost 60% of the cases; the most frequent location is the ventricular myocardium and they are usually subepicardially located, hence they rarely rupture in the pericardial space. The left ventricle is damaged twofold to threefold more frequently than the right one. † The diagnosis of echinococcosis in heart can be divided into two steps: detection of the cyst and its identification as echinococcus. It is based on serological reactions, echocardiography, X-ray, computerized tomography, and/or magnetic resonance imaging. † The most dangerous complication of cardiac echinococcosis is cyst perforation. After cyst perforation three quarters of the patients die from septic shock or embolic complications. † It is very important to understand that chemotherapy may lead to cyst death, and destruction of its wall and result in cyst rupture. Therefore, no germicide must be administered before surgical removal.

In this work, we studied a recombinant mu-class glutathione transferase of 25.5 kDa from Taenia solium metacestode (rTs25GST1-1) that follows Michaelis–Menten kinetics with 1-chloro-2,4-dinitrobenzene (CDNB). The kinetic parameters obtained for rTs25GST1-1 with CDNB and GSH were V max = 12.04 μmol/min/mg and Km = 1.38 mM, and V max = 10.20 μmol/min/mg and Km = 0.90, respectively. The optimal activity was found at pH 8 in the 37–40 °C temperature range. Circular dichroism studies for rTs25GST1-1 at different pH showed that it maintains a typical α-helix structure between pH 6.5–7.5, but loses it between pH 8 and 8.5. Thermal CD assays showed rTs25GST1-1 barely changed its secondary structure. Unfolding/refolding assays showed that rTs25GST1-1 retained its structure up to 40 °C without loss of its activity. Additionally, exposure of rTs25GST1-1 to cumene hydroperoxide did not produce significant changes in its structure and only affected 50 % of its activity.

Poultry farms in and around Namakkal with a history of tapeworm infection were surveyed for the presence of beetles which could act as intermediate host for the tapeworms. Beetles collected from different poultry farms with suspected tapeworm infection were examined for the presence of metacestode stage of the parasite. A total of 1,880 beetles were collected from 12 poultry farms with suspected tapeworm infection to study the vector potentiality. Out of these, 205 beetles (10.9 %) from nine farms were found to harbour cysticercoids. The percentage of cysticercoid infection in beetles was 8.24, 10.34 and 16.66 % respectively in three different surveys. The beetles harbouring the cysticercoids were identified as Opatroides frater, which may be a natural intermediate host for Raillietina cesticillus. Infection free young chicks (4 weeks old) were experimentally infected with specific number of cysticercoids and prepatent period of tapeworms was found to be between 12 and 13 days. Gravid segments were expelled between 3 and 4 p.m. consistently. The results of this study would help to formulate suitable control measures against the above tapeworm infection.

The cytostatic drugs Vincristine (VCR), Navelbine (NAV), and Methotrexate (MTX) were evaluated for their growth inhibitory potential against metacestodes of Echinococcus multilocularis (Em) by in vitro and in vivo assays. In vitro cultures of E. multilocularis were exposed to IC 90, IC 80, and IC 5 concentrations of VCR, NAV, or MTX for 1 week, then parasite tissue cultures were kept for 1 week without drug exposure in vitro, and thereafter, metacestode tissues were injected intra-peritoneally into Meriones unguiculatus. Metacestode growth was monitored for several months post-infection (p.i.) by body weight control, magnetic resonance imaging (MRI), and autopsy at 5 months p.i. Weight monitoring of infected M. unguiculatus did not provide conclusive evidence for Em-metacestode growth, while MRI could detect growing Em-metacestode in the MTX-treated group at 8 weeks (p.i.), whereas metacestodes exposed to VCR and NAV were at 17 weeks (p.i.) detectable. MRI disclosed progressive and massive growth of Em-metacestode in the VCR- and MTX-exposed groups, while the NAV-pretreated Em-metacestodes' volume did not exceed 4 cm(3). At autopsy, Em-metacestodes of less than 4 cm(3) were found in infected M. unguiculatus, which was not detected by MRI. In summary, the cytostatic drugs Methotrexate, Navelbine, and Vincristine--as applied in the present work--did not show parasitocidal or clear parasitostatic effects on metacestodes of E. multilocularis. While parasite growth in vivo was inhibited in NAV- and VCR-pretreated Em-metacestodes, MTX pretreatment seemed to enhance parasite proliferation. Magnetic resonance imaging appears suitable to monitor in vivo the effects of drugs on growth progression and regression only of larger Em-metacestode tissues.

Control of cellular redox homeostasis is a central issue for all living organisms. Glutathione and thioredoxin enzymatic redox systems are the usual mean used to achieve such a control. However, parasitic platyhelminths studied to date possess a nicotinamide adenine dinucleotide phosphate-dependent thioredoxin-glutathione reductase (TGR) as the sole redox control system. Thus, TGR is considered as a potential therapeutic target of parasitic platyhelminths, and based on this assumption, the gold compound auranofin is a potent inhibitor of TGR. The aim of this research was to investigate the effect of auranofin on metacestode (cysticerci) of Taenia crassiceps in culture. Accordingly, the time course for viability and respiration of cysticerci in culture was evaluated in the presence of this compound. After 4 h at 10 microM auranofin, 90% of cysticerci were alive, but respiration activity had declined by 50%. After 12 h, neither survivors nor respiration was detected; a LD(50) for auranofin of 3.8 microM was calculated. Interestingly, crude extracts of cysticerci pretreated with 3 microM auranofin nearly nil TGR activity (IC(50) = 0.6 microM). Zymography for TGR in polyacrylamide gel electrophoresis was conducted because the previously mentioned extracts clearly showed a dose-response inactivation of TGR toward auranofin. The killing of cysticerci by this gold compound is most likely related with TGR inactivation. Therefore, further research on the suitability of auranofin as a therapeutic tool in the treatment of cysticercosis in animals and humans is sustained.

In many helminthic infections, eotaxin, a CC-chemokine, triggers the mobilization of eosinophils, thus, contributing to an elevated blood and periparasitic eosinophil level. Following an experimental intraperitoneal infection of C57BL6 mice with Echinococcus multilocularis metacestodes, however, we observed the absence of eosinophils in the peritoneal cavity and a low number of such cells in the blood of infected animals. Therefore, we carried out an explorative study to address the question why eosinophilia did not occur especially in the peritoneal cavity of such secondarily AE-infected mice. In an in vitro assay, we showed that metacestode antigens (in vitro generated vesicle fluid and E/S products) were able to proteolytically digest eotaxin. This effect was confirmed with semiquantitative Western blotting, which demonstrated a decreasing intensity of remaining eotaxin signals. Proteolysis of eotaxin was, thus, dose-dependent and proportional to the time of incubation with the metacestode antigens. Using appropriate inhibitors, the respective protease was identified as a cysteine protease, which required the presence of Ca(++) as co-enzyme. A chromatographic fractionation procedure by successive separation of VF molecules using a superpose column and subsequently a MonoQ column mounted on an FPLC system allowed to yield a fraction, referred to us as fraction 6; containing the enriched cysteine protease, this fraction will be used for further molecular studies. Eotaxin inactivation by VF and E/S products may contribute to explain the absence of eosinophils within the peritoneal cavity of AE-secondary infected mice. Absent eosinophils, thus, may be a part of a series of events that maintain a low level of inflammation displayed within the peritoneal cavity of experimentally infected mice.

Background Shepherd and stray dogs are thought to represent the primary definitive hosts of Coenurosis by Taenia multiceps, due to their feeding habits which translate into high chances of coming into contact with infected intermediate hosts. Nonetheless, little attention has been paid to the role of the red fox (Vulpes vulpes) in the epidemiology of coenurosis. In fact a knowledge gap exists on the role played by red foxes in the epidemiology of Taenia multiceps and the capability of this parasite to produce fertile and viable eggs in this wild canid, i.e. on the occurrence of a sylvatic cycle. This study investigates the role of the red fox (Vulpes vulpes) in the epidemiology of T. multiceps and related metacestodoses. Methods The small intestine of 63 red foxes was macroscopically examined for the presence of cestodes. Adult parasites were identified morphologically as being T. multiceps. Tapeworm eggs were counted and stored at 4 °C in physiological saline solution prior to experimental infection of four sheep and one goat. Sheep were inoculated orally on Day 0 with 3000 (sheep 1), 5000 (sheep 2 and 3) or 7000 eggs (sheep 4), while the goat was infected with 5000 eggs of T. multiceps. The animals were followed-up regularly by MRI and underwent surgical treatment between days 180 to day 240 post infection. Collected coenuri were identified using morphological and molecular methods. Results A total of 6.3 % of red foxes were found infected with T. multiceps and the eggs obtained from the worms were determined to have a viability of 45.4 %. Two of the challenged sheep and the goat developed disease compatible with T. multiceps. Morphometrical features of the cysts were consistent with those of T. multiceps; nucleotide amplification and sequencing of mitochondrial genes (i.e., cox1 and Nd1) from the metacestode material confirmed the identification. Conclusions The present study is the first to provide evidence of the role of the red fox as a competent definitive host for T. multiceps, thus changing the epidemiological scenarios of infections by this cestode.

The purpose of this study was to determine the Th1 and Th2 cytokine responses induced by Taenia solium metacestode antigens in mice and correlate them with the immune responses elicited in vivo. To assess this aim, mice were inoculated with metacestode antigens. RNA was obtained from spleen cells of immunized or control mice incubated with metacestode antigens and used to determine the cytokine profile. Peripheral blood eosinophilia was measured daily in each mouse and specific serum antibody levels were determined. Results showed that metacestode antigens induce the synthesis of IL-4, IL-5 and IFN-gamma mRNAs in spleen cells. They also induced peripheral blood eosinophilia and elicited specific IgE and IgG antibodies, especially IgG1. Three antigens were recognized by all IgG subclasses and by IgE (104, 88 and 7 kDa), and a 57-kDa protein was recognized by IgG1, IgG2a, IgG2b, and IgE. IgG1 and IgG2b recognized 52, 30 and 20 kDa antigens. Immune responses elicited in vivo and the cytokine profile showed good correlation.

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.

The efficacy of a combination of ivermectin (IVM) and albendazole (ABZ) used as a chemoprophylactic treatment and as treatment of secondary hydatidosis is described for the first time. IVM treatment alone was not effective against Echinococcus granulosus, either when the protoscolices were recently inoculated or when they had developed to the metacestode stage. However, the efficacy of IVM and ABZ when used in combination as a prophylactic treatment was 95.72% and 87% with respect to the number and the wet weight of cysts, respectively. These results were higher than in the treatment of secondary hydatidosis, which were 44.8% and 45.26%, respectively. The ultrastructural changes in the germinal layer of the cysts after the treatments are also described.

The cyst differentiation of protoscolices inoculated in NMRI mice was studied in detail. Although the laminated layer began its formation at 21 days post-infection (p.i.), we could not identify this structure in all parasites until 34 days p.i.. At 34 days p.i., all protoscolices were transformed into the metacestode stage. Subsequently, the efficacy of praziquantel (PZ) was studied in different stages of development of the parasite. We have found a different PZ susceptibility in both earlier stages and well-developed cyst. PZ treatment exhibited a high efficacy at the beginning of cyst differentiation and a zero efficacy in the mature metacestode when the cystic layers were totally developed. The relationship between the tegumental ultrastructural changes occurring during the vesicular evolution of protoscolices in NMRI mice and the efficacy of PZ treatment is discussed in the present study.

Background: Taenia solium taeniasis/cysticercosis is a zoonotic helminth infection mainly found in rural regions of Africa, Asia and Latin America. In endemic areas, diagnosis of cysticercosis largely depends on serology, but these methods have their drawbacks and require improvement. This implies better knowledge of the proteins secreted and excreted by the parasite. In a previous study, we used a custom protein database containing protein sequences from related helminths to identify T. solium metacestode excretion/secretion proteins. An alternative or complementary approach would be to use expressed sequence tags combined with BLAST and protein mapping to supercontigs of Echinococcus granulosus, a closely related cestode. In this study, we evaluate this approach and compare the results to those obtained in the previous study. Findings. We report 297 proteins organized in 106 protein groups based on homology. Additional classification was done using Gene Ontology information on biological process and molecular function. Of the 106 protein groups, 58 groups were newly identified, while 48 groups confirmed previous findings. Blast2GO analysis revealed that the majority of the proteins were involved in catalytic activities and binding. Conclusions: In this study, we used translated expressed sequence tags combined with BLAST and mapping strategies to both confirm and complement previous research. Our findings are comparable to recent studies on other helminth genera like Echinococcus, Schistosoma and Clonorchis, indicating similarities between helminth excretion/secretion proteomes. © 2013 Victor et al.; licensee BioMed Central Ltd.

Subcutaneous implantation of Taenia solium metacestodes in mice induces an inflammatory reaction made up mainly of neutrophils and eosinophils after 12 days. Administration of a small RNA-peptide (metacestode factor, MF) purified from T. solium metacestodes significantly reduces the inflammatory site in both size and composition, yielding a very low number of eosinophils. The metacestodes implanted in control mice were completely destroyed and their remnants were surrounded by an intense inflammation predominantly made up of neutrophils and eosinophils. In contrast, metacestodes implanted in mice treated with MF showed apparently intact suckers, rostellum, hooks, and tegument. Inhibition of inflammation around the parasites was also observed in mice immunized with T. solium metacestode antigens and inoculated simultaneously with MF. Mice immunized only with T. solium metacestode antigens produced a granulomatous process around metacestodes that destroyed most of the large metacestode structures: suckers, rostellum, hooks, and tegument-wall tissues. Furthermore, treatment of mice with MF or implanted metacestodes decreased the antibody (P

The immunoreactivity of Taenia saginata and T. crassiceps metacestode neutral glyco(sphingo)lipids towards IgG antibodies derived from the sera of calves with experimental cysticercosis has been established. The glyco(sphingo)lipids are separable by normal-phase HPTLC (high-performance thin-layer chromatography) into groups of increasing sugar-chain length (lipid/ceramide mono-, di-, tri-, tetra- andtetrasaccharides), with those corresponding to three and four hexoses being the main immunoreactive components (HPTLC immunostaining). In ELISA (enzyme-linked immunosorbent assay), reverse-phase HPTLC-isolated T. crassiceps metacestode glyco(sphingo)lipids equivalent to tri- and tetrahexoside allowed a discrimination between non-infected and infected calves (at least 80 metacestodes recovered). The formation of IgG antibodies was correlated with the infection, not with other non-specific inducing factors, as seen by the differential humoral response detected in experimentally infected (T. saginata) calves before and after Praziquantel treatment (HPTLC immunostaining and ELISA).

Immunohistochemistry was used to examine the type of lymphocytes in muscle taeniosis-cysticercosis in naturally infected pigs. The inflammatory response studied was classified into lesions of grades 1, 3, and 5. In grade 1, with a minimal inflammatory infiltrate consisting of eosinophils and a few mononuclear cells, the immunostaining showed more CD4+ cells than CD8+ cells and IgM cells. In grade 3, when the granulomatous reaction was not yet well developed and the destruction of the parasite began, CD4+ and Ig M+ were the predominant cells, although CD8+ cells showed a notable increase. In grade 5, with a few parasitic structures surrounded by an extensive granulomatous infiltrate, lymphocyte subsets were decreased in number and did not show differences from grade 1 except for IgM+ cells, which remained increased. The organization of an active inflammatory response against the metacestode of Taenia solium in pigs includes the sequential participation of CD4+, CD8+ and IgM+ lymphocytes.

The nervous system of Cysticercus cellulosae, the metacestode stage of the tapeworm Taenia solium, was delineated using histochemical methods for the localization of the enzyme markers; nonspecific esterase and acetylcholinesterase. The main features of the nervous system include a pair of cerebral ganglia, a circumcerebral nerve ring, a rostellar nerve ring, and anterior and posterior nerves and their branches. The posterior nerves form a subtegumental network in the strobila and the bladder wall. A nerve network around excretory tubules could also be demonstrated, suggesting neuronal control of excretion in the metacestode. No sheath was observed around the nervous system. The morphological features described suggest "cephalization" of the nervous system in this parasite.

Taenia saginata cysticercosis in cattle. Acta vet. scand. 1991, 32, 233–241. – Serum IgG response of cattle with cysticercosis caused by Taenia saginata was studied in an enzyme-linked immunosorbent assay (ELISA) where a T. saginata metacestode surface extract was used as antigen. In experimentally infected calves, a sharp rise in specific antibody levels was found 3–4 weeks after the infection followed by a slow decrease. Preinfection levels were reached 2.5 years after infection. The serological level of detection corresponded to about 25 cysts. The ELISA was employed in cattle herds where cysticercosis outbreaks had occurred and also in supposedly uninfected herds. Significantly increased antibody levels were found in the herds with massive cysticercosis cases. The test was not adapted for individual diagnosis as some animals of the uninfected herds, especially within the older age groups, had elevated antibody values. The ELISA was, however, useful in the investigation of outbreaks to determine the extent and pattern of the infection in the herd. The rate of decline in antibody levels in these herds was studied by follow up sampling. The increased antibody levels in the infected herds were also reflected in colostrum-fed calves. This observation was employed to estimate the time of infection.

An enzyme-linked immunosorbent assay (ELISA) was adapted for the serological differential diagnosis of cystic or alveolar echinococcosis in man caused by Echinococcus granulosus or E. multilocularis respectively. By affinity chromatography using rabbit anti hydatid fluid IgG coupled covalently to CNBr-Sepharose 4B a protein fraction (Em 1) containing shared antigens of both parasites could be isolated from an extract of E. multilocularis metacestode tissue. From the same source another antigen fraction (Em 2) with a high degree of specificity for E. multilocularis was prepared by immunosorption. Antigen Em 1 was equally sensitive for the detection of antibodies against E. granulosus and E. multilocularis, whereas antigen fraction Em 2 appeared to be more specific for E. multilocularis. A correct serological differential diagnosis was achieved in 95% of 57 confirmed cases of human cystic or alveolar echinococcosis by the simultaneous use of both antigen fractions in the ELISA and by comparison of their reactivities.

The antigenicity and specificity of crude antigens collected during the in vitro maintenance of Taenia hydatigena and T. ovis, excretory/secretory (ES) antigens, were assessed in a peroxidase microenzyme-linked immunosorbent assay (ELISA), using sera from lambs given experimental monospecific infections with T. hydatigena, T. ovis, Echinococcus granulosus or Fasciola hepatica. ES antigens of larval cysts of T. ovis and T. hydatigena were less reactive than those of adult or oncosphere stages. Strong interspecific cros-reactions occurred between all antigen preparations, and these antigens offered no better specificity than crude somatic extracts. IgG1 was the major immunoglobulin detected in sera from lambs experimentally infected with T. ovis or T. hydatigena using antigens prepared from sonicated oncospheres. Discrete peaks of anti-oncospheral antibodies were detected following initial and challenge infections with eggs (whether the homologous or heterologous species), when sera were assayed with a PBS sonicate or an ES antigen from oncospheres. However, when oncospheres solubilised with sodium deoxycholate were used, the antibody response was prolonged and resembled that reported previously when somatic extracts of adult and metacestode stages were used as antigen. The results showed that oncospheres share antigens in common with other life-cycle stages, but also support the notion that they may possess some unique stage-specific antigenic determinants.

IN a previous article1 evidence was reported supporting the hypothesis that at least two immune responses occur against larval taeniid tapeworms in the intermediate host; and that these responses can be elicited by a single parenteral injection of homologous activated embryos in previously unexposed animals. The initial response appears to occur before the penetration of the hexacanth embryo through the intestinal barrier, and the later response occurs at the site of election against the growing metacestode.

THE asexually proliferating larvae of the cestode Echinococcus multilocularis have an intriguing resemblance to malignant tumours. Indeed, until 1856 when Virchow demonstrated the parasitic nature of these cyst masses in man, such lesions were commonly diagnosed as carcinomas of the liver1. In the laboratory, infections with this parasite can be propagated vegetatively by transplanting the metacestode into the peritoneal cavity of susceptible rodent intermediate hosts2. The protoscolices transform into small cysts which grow and metastasise by peripheral herniation and the detachment of minute vesicles3.