Search

This article analyzes data from scientific publications (mainly reviews) concerning the link between human neurocysticercosis and epilepsy. Along with data from our own studies on experimental hippocampal sclerosis induced by a Taenia crassiceps metacestode factor in mice, it explores the connection between mechanisms that likely favor the development of epilepsy in cases of human neurocysticercosis. The data from both sources suggest the idea that the T. solium metacestode factor causes hippocampal sclerosis and later epilepsy in humans with neurocysticercosis.

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

PURPOSE OF REVIEW Infection with the larval (metacestode) stage of Echinococcus multilocularis causes alveolar echinococcosis (AE), a serious hepatic disorder. The parasite has increased its infection extensity in wildlife and domestic dogs, mainly due to urbanization and spatial extension of wildlife hosts in Europe, Asia as well as North America, resulting in emerging infection risk for humans. RECENT FINDINGS In hyperendemic areas such as Kyrgyzstan and China, ecological and socioeconomic changes have been associated with the unpredictable increase of AE cases. In North America, the appearance of the European-like genotype is of concern. In Europe, the annual increase of human case numbers reached a plateau even in hyperendemic situations. Therefore, we conclude that most of the exposed individuals are resistant to parasite invasion and/or to disease development. Thus, AE develops in a few healthy individuals, but preferentially in immunosuppressed patients. SUMMARY In the future, improved diagnostic strategies will allow more precise estimations of transmission routes including the role of food, water and direct dog contact, which should yield improved public health recommendations. Finally, understanding protective innate and acquired immune mechanisms as well as parasite-driven immune-evasion processes will be essential to develop curative therapies in nonoperable patients and, futuristically, appropriate vaccines.

Neurocysticercosis (NCC) is a parasitic infection caused by Cysticercus cellulosae, the metacestode of pork tapeworm ( Taenia solium). NCC is one of the most common public health problems worldwide. We present a patient harboring a bilobed ring-enhancing lesion with a presumed diagnosis of brain metastasis, who returned to the USA after traveling to an endemic region. The diagnosis of NCC was established based on a characteristic resonance of succinate on proton magnetic resonance spectroscopy. Also, higher mean diffusivity and lower fractional anisotropy along with relative cerebral blood volume were observed from the lesion compared to contralateral normal brain regions. Multiparametric analysis may improve the differential diagnosis of ring-enhancing intracranial lesions such as NCC.

Background Cysticercosis caused by cysticercus tenuicollis is a metacestode infection that affects several species of ungulates. It is caused by the larval stage of Taenia hydatigena, an intestinal tapeworm in dogs and wild canids. In the intermediate host, the mature cysticerci are usually found in the omentum, mesentery, and peritoneum, and less frequently in the pleura and pericardium. The migrating larvae can be found mostly in the liver parenchyma causing traumatic hepatitis in young animals. Most infections are chronic and asymptomatic, and are diagnosed at the abattoir. The acute form of infection is unusual in sheep and reports of death in lambs are rare. Methods In March 2018, fifteen female lambs presented anorexia, weakness, lethargy, and death secondary to acute visceral cysticercosis. Twelve of them underwent hepatic ultrasonography. Examinations were performed on standing or left lateral recumbent animals. Results Livers of affected animals presented rounded margins and a thickened, irregular and hyperechoic surface. Hepatic parenchyma appeared to be wholly or partially affected by lesions characterized by heterogeneous areas crossed by numerous, irregular, anechoic tracts ranging from 1 to 2 cm in length and 0.1 to 0.2 cm in width. Superficial and intraparenchymal cystic structures were also visualized. The presence of lesions was confirmed by anatomopathological examination, and T. hydatigena cysticerci was identified by morphological and molecular characterization of isolates. Conclusions Our results highlighted that hepatic ultrasonography is effective for an intra-vitam diagnosis of acute cysticercosis in lambs.

Alveolar echinococcosis (AE) is a life-threatening parasitic disease caused by the zoonotic cestode Echinococcus multilocularis. Our goals were to confirm infection, identify species, and analyze biogeographical origin of metacestode tissues from a suspected human AE case in Saskatchewan, Canada. We conducted polymerase chain reaction (PCR) targeting the nad1 mitochondrial gene for E. multilocularis and the rrnS ribosomal RNA gene for E. granulosus and conducted haplotype analysis at the nad2 locus. Our analysis confirmed AE and indicated that sequences matched infected Saskatchewan coyotes and European E3/E4 haplotypes. The patient had no travel history outside North America. This suggests autochthonous transmission of a European-type strain.

Background Cysticercus tenuicollis (larvae of Taenia hydatigena) is a frequent cosmopolitan endoparasite of ruminants. The infestation by this parasite is underestimated since it is neither zoonotic nor inducing high economic losses in the sheep sector. Methods This study aimed at estimating different parasitological indicators, Cysticercus tenuicollis infestation in small ruminants using a slaughterhouse‐based survey in Northeast Tunisia. A total number of 3,692 sheep and 78 goats were examined in the slaughterhouse of Menzel Temime. Results The overall prevalence was estimated to be 2.8 (106/3692) and 8.9% (7/78) in sheep and goats, respectively. The abundance in these two species was 0.24 and 0.05 and the intensity 1.97 and 2.85, respectively. In goats, all the cysts were found in the mesentery, whilst, in sheep, the majority were in mesentery (96%) but also on the liver in 2% of the cases and in both organs (2%). There were only cysts with aqueous liquid with predominantly middle‐sized cysts (1 to 3 cm) corresponding to 63.15 and 70.34% in sheep and goats, respectively. Conclusion Since, in small ruminants, the infestation by C. tenuicollis indicators are not high, low cost control measures should be implemented to eliminate this parasite in Northeast Tunisia., The authors estimated the infestation prevalence of sheep and goats by Cysticercus tenuicollis. The overall prevalence was estimated to 2.8 (106/3692) and 8.9% (7/78) in sheep and goats, respectively. The abundance in these two species was 0.24 and 0.05 and the intensity was 1.97 and 2.85, respectively.

The cestode Taenia hydatigena uses canids, primarily dogs, as definitive hosts, while the metacestode larval stage cysticercus infects a range of intermediate hosts, including domestic animals such as goats, sheep, and pigs. Cysticercosis due to T. hydatigena has large veterinary and economic drawbacks. Like other taeniids, e.g., Echinococcus, intraspecific variation is found among the members of the genus Taenia. In Africa, few studies are available on the epidemiology and distribution of T. hydatigena, and even fewer studies are available on its genetic variation. In this study, we molecularly identified 11 cysticerci from sheep in Sudan and demonstrated the genetic variation based on the NADH dehydrogenase subunit 1 (nad1) and cytochrome c oxidase subunit 1 (cox1) mitochondrial genes. The isolates were correctly identified as T. hydatigena with more than 99% similarity to those in the GenBank database. Low diversity indices and insignificant neutrality indices were observed, with 3 and 2 haplotypes for the nad1 and cox1 genes, respectively. The results suggest the presence of unique T. hydatigena haplotypes in Sudan, as haplotypes with 100% similarity were not found in the GenBank database. With few available studies on the genetic variation of T. hydatigena in Africa, this report represents the first insights into the genetic variation of T. hydatigena in Sudan and constitutes useful data.

Cystic echinococcosis (CE) is a zoonotic parasitic disease caused by infection with the larvae of Echinococcus granulosus sensu lato (s.l.) cluster. It is urgent to identify novel drug targets and develop new drug candidates against CE. Glucose transporter 1 (GLUT1) is mainly responsible for the transmembrane transport of glucose to maintain its constant cellular availability and is a recent research hotspot as a drug target in various diseases. However, the role of GLUT1 in E. granulosus s.l. (EgGLUT1) was unknown. In this study, we cloned a conserved GLUT1 homology gene (named EgGLUT1-ss) from E. granulosus sensu stricto (s.s.) and found EgGLUT1-ss was crucial for glucose uptake and viability by the protoscoleces of E. granulosus s.s. WZB117, a GLUT1 inhibitor, inhibited glucose uptake by E. granulosus s.s. and the viability of the metacestode in vitro. In addition, WZB117 showed significant therapeutic activity in E. granulosus s.s.-infected mice: a 10 mg/kg dose of WZB117 significantly reduced the number and weight of parasite cysts (P < 0.05) as efficiently as the reference drug, albendazole. Our results demonstrate that EgGLUT1-ss is crucial for glucose uptake by the protoscoleces of E. granulosus s.s., and its inhibitor WZB117 has a therapeutic effect on CE.

The metacestode stage of Echinococcus granulosus can cause cystic echinococcosis (CE), which still widely occurs around the world. Since the early 1970s, benzimidazoles have been shown to inhibit the growth of cysts and used to treat CE. However, benzimidazoles are still ineffective in 20%-40% of cases. In order to explore the new agents against CE, we have investigated the therapeutic effect of the recombinant adenoviral vector expressing mouse IL-28B (rAd-mIL-28B) on protoscoleces-infected mice. In our study, we successfully established the model mice which infected with protoscoleces intraperitoneally. At 18 weeks post-infection, the mice received rAd-mIL-28B (1×107 PFU) weekly by intramuscular injection for 6 weeks. Compared with the untreated control (13.1 ± 2.2 g), there was a significant reduction in cysts wet weight in rAd-mIL-28B group (8.3 ± 3.5 g) (P, Author summary Echinococcosis is a chronic zoonotic parasitic disease, which is caused by the larval stage of Echinococcus granulosus (E. granulosus) and Echinococcus multiocularis (E. multiocularis). This disease is still widely prevalent in the world and seriously endangers human health and life, causing heavy burdens and economic losses to agriculture and animal husbandry. China is also one of the high incidence areas of the disease. At present, the preferred treatment is surgical excision of the parasitic mass, but patients with multiple cysts appear in multiple organs have to receive drug therapy. The approved chemotherapeutic drugs in clinic, such as albendazole (ABZ) and mebendazole, often do not work expectedly with a high rate of recurrence. Therefore, it is urgent to develop the new anti-echinococcal drug. IL-28B is an important member of type-III IFNs, which is equipped with the capacity of anti-viral activity and anti-tumor. In our previous study, we found that IL-28B could inhibit the proliferation of cervical cancer cells via down-regulating Treg cells in mice. This study mainly studied the therapeutic effect of rAd-mIL-28B on E. granulosus-infected mice, and the results showed that rAd-mIL-28B could relieve the parasitic burden and inhibit Treg cells meanwhile improve the Th1 and Th17 immune responses. It may contribute to another choice for the anti-echinococcal treatment.

A typical multivesiculated metacestode tissue has been found in the liver of a European brown hare (Lepus europaeus) originating from a northern area of Switzerland. In this study, the causative species was identified as Echinococcus multilocularis by appropriate histological and molecular analyses and corresponding DNA sequencing. This is the first confirmation of larval E. multilocularis from hares in central Europe. The metacestode tissue contained protoscolices, suggesting that the hare may contribute to the transmission of E. multilocularis in Switzerland.

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.

International audience; Assessing the genetic diversity of the parasite Echinococcus multilocularis provides key information about the temporal and spatial strain flow in a given area. Previous studies indicated that a historical endemic area conventionally presents a relatively high genetic diversity, whereas peripheral or newly endemic areas exhibit a more restricted variability of the parasite. The Swiss plateau region is part of the European historically endemic area, and the genetic diversity has already been investigated by assessing either human metacestode isolates or adult worms from foxes. To date, there have been no studies covering the whole geographical area affected by the parasite. The aim of the present study was to make use of the domestic pig to investigate the genetic diversity of E. multilocularis in relation to spatial distribution. A total of 55 E. multilocularis-induced hepatic lesions from slaughtered pigs from Switzerland were studied using EmsB microsatellite analyzes, and findings were compared to already published data (originating from human, primate, foxes, and rodent samples). A total of 12 EmsB profiles were described among the domestic pigs, some of them presenting a clear spatial organization in the Swiss plateau, with three of the main profiles geographically separated. One of the 12 EmsB profiles has been newly identified for Switzerland in this study, while the other 11 profiles had been previously described in other Swiss E. multilocularis isolates from other hosts. Overall, a total of 18 EmsB profiles have so far been described within the Swiss endemic area. Six profiles appeared only among human, primate, rodent, and fox samples. Based on a richness and diversity accumulation analysis, the sampling efficiency for the whole studied area has now been improved considerably by compilation of 178 E. multilocularis specimens obtained from four different intermediate and one definitive host species in Switzerland.

Coenurosis is a serious parasitic disease of herbivorous animals caused by the metacestode of Taenia multiceps (Coenurus cerebralis). Accordingly, a significant amount of research is currently dedicated to the development of appropriate antigens for use in rapid and accurate coenurosis diagnosis kits. In the present study, antigen B (AgB) and thioredoxin peroxidase (TPx) from T. multiceps were cloned and expressed using a prokaryotic system, molecular characterization of Tm-AgB was determined by bioinformatical analyses. The serological diagnostic potentials of rTm-AgB and rTm-TPx were evaluated by indirect ELISA and compared with those of previously reported rTm-AnxB2, rTm-HSP70, and rTm-GST. The results showed that Tm-AgB is a specific lipoprotein of cestodes with good thermal stability. The ELISA assay showed that rTm-AgB exhibited a sensitivity of 95.8% and a specificity of 87.5%, indicating its strong potential for serological diagnosis of T. multiceps.

Albendazole, as the main anti-echinococcal benzimidazole, has demonstrated safe and effective therapeutic outcomes in the treatment of echinococcosis. The emergence of resistance or reduced response to albendazole sulfoxide (ABZ_SOX) and other benzimidazoles have been demonstrated in several parasitic helminths of medical and veterinary importance. As the genetic makeup and miRNA profile of helminths affects their response to albendazole sulfoxide, the present study was conducted to investigate the expression of miRNAs in different developmental stages of Echinococcus granulosus exposed to albendazole sulfoxide in vitro. Different developmental stages of the helminth were obtained from in vitro cultured E. granulosus in monophasic and diphasic media. In both ABZ-SOX-treated and control parasites miRNAs were extracted from microcysts, intact protoscoleces and strobilated worms with one and three segments. Expression of two miRNAs, let-7 and miR-61 was evaluated using RT-qPCR for each stage. Results of the present study revealed significant differential expression of both let-7 and miR-61 at different drug concentrations. A significant difference of let-7 expression was observed between the strobilated and metacestode stages of E. granulosus exposed to ABZ-SOX. In the treated protoscoleces, let-7 expression was significantly reduced in the presence of ABZ-SOX at 1000 μg/ml concentration. In contrast higher expression levels were documented in the segmented worms. In the microcysts exposed to different drug concentrations a significant decline of miR-61 expression was demonstrated. Also, a significant increase in expression of miR-61 was observed in one proglottid worms as well as the protoscoleces. Under high drug concentration or long-term exposure of the protoscoleces to ABZ-SOX significantly higher miR-61 expression was observed compared to the controls. Our findings suggested that under in vitro benzimidazole exposure the expression of two E. granulosus miRNAs were significantly affected in the microcyst stage. This study presents the first evidence of the nature of benzimidazole effects on miRNA expression in platyhelminths.

Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico (CNPq) Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil / University of S?o Paulo. Faculty of Veterinary Medicine and Animal Science. Department of Pathology. S?o Paulo, SP, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. University of S?o Paulo. Faculty of Veterinary Medicine and Animal Science. Department of Pathology. S?o Paulo, SP, Brazil. Minist?rio da Sa?de. Secretaria de Vigil?ncia em Sa?de. Instituto Evandro Chagas. Ananindeua, PA, Brasil. We describe a case of microscopic liver lesion caused by larval cestode in a frugivorous bat (Molossus molossus) from the Western Brazilian Amazon. Histopathological analysis of liver indicated the occurrence of metacestode associated with multifocal histiocytic response and the phylogenetic analysis of Cox-I and 18S rDNA genes indicated that the parasite belonged to the family Paruterinidae. This is the first identification of cestodes of the family Paruterinidae in bats and may suggest a broad range of paruterinid tapeworm hosts to be investigated in the Brazilian Amazon.

The essential oil (EO) of Thymus capitatus, seven fractions (F1–F7) obtained from silica gel chromatography, and several pure EO components were evaluated with respect to in vitro activities against Echinococcus multilocularis metacestodes and germinal layer (GL) cells. Attempts to evaluate physical damage in metacestodes by phosphoglucose isomerase (PGI) assay failed because EO and F1–F7 interfered with the PGI-activity measurements. A metacestode viability assay based on Alamar Blue, as well as transmission electron microscopy, demonstrated that exposure to EO, F2 and F4 impaired metacestode viability. F2 and F4 exhibited higher toxicity against metacestodes than against mammalian cells, whereas EO was as toxic to mammalian cells as to the parasite. However, none of these fractions exhibited notable activity against isolated E. multilocularis GL cells. Analysis by gas chromatography-mass spectrometry showed that carvacrol was the major component of the EO (82.4%), as well as of the fractions F3 (94.4%), F4 (98.1%) and F5 (90.7%). Other major components of EO were β-caryophyllene, limonene, thymol and eugenol. However, exposure of metacestodes to these components was ineffective. Thus, fractions F2 and F4 of T. capitatus EO contain potent anti-echinococcal compounds, but the activities of these two fractions are most likely based on synergistic effects between several major and minor constituents.

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.

Canids, particularly dogs, constitute the major source of cystic echinococcosis (CE) infection to humans, with the majority of cases being caused byEchinococcus granulosus(G1 genotype). Canine echinococcosis is an asymptomatic disease caused by adult tapeworms ofE. granulosussensu lato (s.l.). Information on the population structure and genetic variation of adultE. granulosusis limited. Using sequenced data of the mitochondrial cytochromecoxidase subunit 1 (cox1) we examined the genetic diversity and population structure of adult tapeworms ofE. granulosus(G1 genotype) from canid definitive hosts originating from various geographical regions and compared it to that reported for the larval metacestode stage from sheep and human hosts.Echinococcus granulosus(s.s) was identified from adult tapeworm isolates from Kenya, Libya, Tunisia, Australia, China, Kazakhstan, United Kingdom and Peru, including the first known molecular confirmation from Gaza and the Falkland Islands. Haplotype analysis showed a star-shaped network with a centrally positioned common haplotype previously described for the metacestode stage from sheep and humans, and the neutrality indices indicated population expansion. Low Fst values suggested that populations of adultE. granulosuswere not genetically differentiated. Haplotype and nucleotide diversities forE. granulosusisolates from sheep and human origin were twice as high as those reported from canid hosts. This may be related to self-fertilization ofE. granulosusand/or to the longevity of the parasite in the respective intermediate and definitive hosts. Improved nuclear single loci are required to investigate the discrepancies in genetic variation seen in this study.

A total of 13 metacestodes were collected from the lung and parietal pleura from a red brocket deer (Mazama americana) from the Peruvian Amazon. All metacestodes were identified as cysticerci of Taenia omissa by morphological and molecular analyzes. Cytochrome c oxidase subunit 1 sequences from the new isolate T. omissa had more than 96.8% identity with other Peruvian isolates of the species previously sequenced. Lower similarities (93.8-95.8%) were verified between Peruvian and Canadian isolates. This finding adds a new intermediate host for T. omissa and also expands its geographical distribution.

Cysticercosis is a disease caused by the metacestode of the parasite Taenia solium (T. solium). In humans, the most severe complication of the disease is neurocysticercosis. The drug of choice to treat this disease is albendazole; however, the bioavailability and efficacy of the drug are variable. Therefore, new molecules with therapeutic effects against this and other parasitic infections caused by helminths must be developed. Naphthoquinones are naphthalene-derived compounds that possess antibacterial, antifungal, antitumoral, and antiparasitic properties. The aim of this work was to evaluate the in vitro anti-helminthic effect of 2-[(3-chlorophenylamino)phenylmethyl]-3-hydroxy-1,4-naphthoquinone, isolated from a natural source and then synthesized (naphthoquinone 4a), using an experimental model of murine cysticercosis caused by Taenia crassiceps (T. crassiceps). This compound causes paralysis in the cysticerci membrane from day 3 of the in vitro treatment. Additionally, it induces changes in the shape, size, and appearance of the cysticerci and a decrease in the reproduction rate. In conclusion, naphthoquinone 4a has in vitro cysticidal activity on T. crassiceps cysticerci depending on the duration of the treatment and the concentration of the compound. Therefore, it is a promising drug candidate to be used in T. crassiceps and possibly T. solium infections.

Background Alveolar echinococcosis (AE) is caused by metacestode larva of the tapeworm Echinococcus multilocularis. AE diagnostics currently rely on imaging techniques supported by serology, but unequivocal detection of AE is difficult. Although polymerase chain reaction (PCR)-based methods to detect tapeworm DNA in biopsies have been suggested for several species, no validated protocol adhering to accepted guidelines has so far been presented for AE diagnostics. We herein established a PCR protocol for metacestode biopsies and technically evaluated the method using isolated parasite DNA and cells, biopsies of clinically relevant material, and formalin fixed paraffin-embedded (FFPE) human tissue blocks. We compared the results with an immunochemical (IHC) approach using the monoclonal antibody Em2G11 specific for the antigen Em2 of E. mulitlocularis. Methodology/Principal findings Based on tapeworm 12S rDNA sequences we established and validated a PCR protocol for robust detection of as little as 50 parasite cells per specimen and report 127 cases of positive identification of Echinococcus species in samples from humans and animals. For further validation, we analyzed 45 liver, heart, brain, and soft tissue samples as well as cytological probes of aspirates of FFPE-material from 18 patients with clinically confirmed AE. Of each patient we analyzed (i) fully viable lesions with laminated layer; (ii) tissue with mAbEm2G11-positive small particles of E. multilocularis (spems); (iii) mAbEm2G11-negative tissue adjacent to the main lesion; and (iv) lymph node tissue with mAbEm2G11-positive spems. To identify the areas for the PCR-based approach, we performed IHC-staining with the monoclonal antibody Em2G11. Micro-dissected tissue of these areas was then used for PCR-analysis. 9 of 15 analyzed samples with viable E. multilocularis lesions with laminated layer were positive by PCR. Of this group, all samples preserved for less than 6 years (6/6) were tested positive. 11 of 15 samples of spems and 7 of 9 samples of the control group mAbEm2G11-negative tissue were negative by PCR. We further show that all probes from lymph nodes with spems are PCR negative. Conclusions/Significance We present a sensitive PCR method for the detection of E. multilocularis in human tissue, particularly in fresh biopsy material and tissue blocks stored for less than 5 years. While the diagnostic sensitivity of material containing only spems was higher using IHC, PCR detection was possible in IHC negative liver tissue and in patients with negative serology. Our results support the view that spems do not contain parasitic DNA or viable cells of the parasite. spems thus most probably do not directly contribute to metastasis formation during AE., Author summary Alveolar echinococcosis (AE) is a potentially lethal zoonosis during which metacestode larval tissue of the tapeworm Echinococcus multilocularis grows like a malignant tumor, infiltrating the human liver. Current AE diagnosis, which mainly relies on imaging techniques (ultrasound, computed tomography, magnetic resonance tomography) supported by serology, is not always suitable to unequivocally identify an infection. In this work the authors present a PCR-based approach to detect the parasite in biopsy material taken from patients. The method was first validated for routine laboratory use employing isolated parasite DNA as well as defined numbers of parasite cells in clinically relevant settings. The method was then verified under clinically relevant settings using pathology samples from patients with defined AE. Particularly in cases when these samples had been stored for less than 6 years, the PCR methodology was highly suitable to identify the infection, even when serology was negative. The authors show that small particles of E. multilocularis (spems), containing surface material of the parasite and often located in the draining adjacent lymph nodes, are PCR negative and, thus, most probably do not contain viable parasite cells. The authors conclude that in cases of doubtful serology and imaging, a combination of immunochemical staining methods with PCR are reliable means to unequivocally detect the infection.

Hydatigera taeniaeformis formerly referred to as Taenia taeniaeformis is a cestode of cats (definitive hosts) and rodents (intermediate hosts). The prevalence of the metacestode larval stage has been reported in rodents in many parts of the world even though the genetic polymorphisms or intraspecies variation is still understudied. Here, we report a prevalence of 22.09% (38/172) from an urban rodent population in Pakistan and a nucleotide diversity (cox1) of 0.00463 among the population. Infection was higher in male (27.85%) and adult (32.29%) rats than female and sub-adult/young rats. Interestingly, The median-joining network and phylogenetic construction comprising isolates from China, Japan, Kenya, Laos, Malaysia, Senegal, the United Arab Emirates, and countries in Europe demonstrated that Pakistani H. taeniaeformis are closer to Asian and African population than those of European origin. The results of the study will add-in preliminary data for H. taeniaeformis and will also contribute to understand the global molecular epidemiology and population structure of H. taeniaeformis.

Cystic echinococcosis (CE) is a neglected tropical disease, caused by metacestode (larval) form of the Echinococcus granulosus sensu lato (sl) in humans. MicroRNAs (miRNAs) are small, stable, tissue-specific RNA molecules encoded by the genome that are not translated into proteins. Circulating miRNA expression profiles vary in health and disease. The aim of this study is to determine the altered cellular pathways in CE by comparing the miRNA profiles of controls and CE patients with active or inactive cysts. Following abdominal ultrasonography (US) examination, 20 patients diagnosed with active CE (CE1, CE2, CE3a and CE3b) or inactive CE (CE4 and CE5) and three healthy controls were included in the study. The expression profiles of 372 biologically relevant human miRNAs were investigated in serum samples from CE patients and healthy controls with miScript miRNA HC PCR Array. Compared with the control group, expression of 6 miRNAs (hsa-miR-4659a-5p, hsa-miR-4518, hsa-miR-3977, hsa-miR-4692, hsa-miR-181b-3p, hsa-miR-4491) and one miRNA (hsa-miR-4687-5p) were found to be downregulated in CE patients with active and inactive cysts, respectively (p < 0.05). For downregulated miRNAs in this study, predicted targets were found to be associated mainly with cell proliferation, apoptosis, cell-cell interactions and cell cycle regulation. Further studies in this direction may elucidate the pathogenesis of human CE and the relationship between CE and other pathologies.

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

Cystic echinococcosis (CE) diagnosis by means of serological assays is hampered by the presence of parasites closely related to Echinococcus granulosus sensu lato (s.l.), responsible of the zoonotic disease and with which share cross-reacting antigens. Thus, improvements on the characterization of Echinococcus specific antigens expressed in the larval stage are required, in order to provide useful information for the development of immunological assays for the serodiagnosis of CE in sheep. Here, the proteome of the hydatid cyst fluids (HFs) of Echinococcus granulosus (hydatid fluid, EgHF) and other ovine parasites cyst fluids (CFs), Taenia hydatigena (ThCF) and Taenia multiceps (TmCF) were analyzed by a shotgun proteomic approach. Parasite and host protein profiles in the three types of cyst fluids were characterized and compared. Among the identified proteins, differential parasitic markers with serodiagnostic potential, due to their well-known immunoreactivity in human, included Ag5, AgB proteins, 8-kDa glycoproteins, hydatid disease diagnostic antigen P29 and major egg antigen P40. In particular, seven proteoforms of AgB and 8-kDa glycoprotein resulted to be the most promising diagnostic biomarkers, as they might predict CE in ovine and discriminate between different types of parasites.

The neglected zoonotic disease alveolar echinococcosis (AE) is caused by the metacestode stage of the tapeworm parasite Echinococcus multilocularis. MicroRNAs (miRNAs) are small non-coding RNAs with a major role in regulating gene expression in key biological processes. We analyzed the expression profile of E. multilocularis miRNAs throughout metacestode development in vitro, determined the spatial expression of miR-71 in metacestodes cultured in vitro and predicted miRNA targets. Small cDNA libraries from different samples of E. multilocularis were sequenced. We confirmed the expression of 37 miRNAs in E. multilocularis being some of them absent in the host, such as miR-71. We found a few miRNAs highly expressed in all life cycle stages and conditions analyzed, whereas most miRNAs showed very low expression. The most expressed miRNAs were miR-71, miR-9, let-7, miR-10, miR-4989 and miR-1. The high expression of these miRNAs was conserved in other tapeworms, suggesting essential roles in development, survival, or host-parasite interaction. We found highly regulated miRNAs during the different transitions or cultured conditions analyzed, which might suggest a role in the regulation of developmental timing, host-parasite interaction, and/or in maintaining the unique developmental features of each developmental stage or condition. We determined that miR-71 is expressed in germinative cells and in other cell types of the germinal layer in E. multilocularis metacestodes cultured in vitro. MiRNA target prediction of the most highly expressed miRNAs and in silico functional analysis suggested conserved and essential roles for these miRNAs in parasite biology. We found relevant targets potentially involved in development, cell growth and death, lifespan regulation, transcription, signal transduction and cell motility. The evolutionary conservation and expression analyses of E. multilocularis miRNAs throughout metacestode development along with the in silico functional analyses of their predicted targets might help to identify selective therapeutic targets for treatment and control of AE., Author summary Alveolar echinococcosis (AE) is a zoonotic disease caused by the metacestode stage of the helminth parasite Echinococcus multilocularis. Current treatment requires surgery and/or prolonged drug therapy. Thus, novel strategies for the treatment of AE are needed. MicroRNAs (miRNAs), a class of small ~22-nucleotide (nt) non-coding RNAs with a major role in regulating gene expression, have been suggested as potential therapeutic targets for treatment and control of helminth parasite infections. In this work, we analyzed the expression profile of E. multilocularis miRNAs throughout metacestode development in vitro. We predicted functional roles for highly expressed miRNAs and found that they could be involved in essential roles for survival and development in the host. We determined that E. multilocularis miR-71, a highly expressed miRNA that is absent in the human host, is expressed in germinative cells and in other cell types of the germinal layer in E. multilocularis metacestodes cultured in vitro. Germinative cells are a relevant cell type to target for anti-echinococcosis drug development. MiRNAs that are absent in the human host, involved in essential functions, highly expressed and/or expressed in germinative cells in E. multilocularis metacestodes may represent selective therapeutic targets for treatment and control of AE.

Taeniosis-cysticercosis caused by Taenia crassiceps (Zeder, 1800) is a useful experimental model for biomedical research, in substitution of Taenia solium Linnaeus, 1758, studied during decades to develop effective vaccination, novel anti-helminthic drugs and diagnostic tools. Cysticercosis in mouse (Mus musculus Linnaeus) is achieved by the larval subculturing of the Wake Forest University (WFU) strain of T. crassiceps. Golden hamster, Mesocricetus auratus (Waterhouse), has been shown to be the most suitable host for adult forms of parasite in experimental taeniosis. Metacestodes of T. crassiceps WFU multiply by budding without restrictions once inoculated into the mouse, while the number of tapeworms developed from these larvae in hamsters remains highly variable. Three objectives have been proposed to improve the infection of T. crassiceps WFU in hamsters: (1) to re-evaluate the need of immune suppression; (2) to investigate the advantage of infecting hamsters with metacestodes with in vitro protruded scolices; and (3) to compare a number of tapeworms developed from metacestodes subcultured in hamsters against those proliferated in mice. Our results demonstrated that when the evagination of murine metacestodes was high, the number of T. crassiceps WFU adults obtained from hamsters was also high. Immunosuppressive treatment remains relevant for this experimental rodent model. The hamster-to-hamster cysticercosis-taeniosis by T. crassiceps overcame the mouse-to-hamster model in the yield of adult specimens. In vitro scolex evagination and metacestode asexual proliferation in hamsters place this rodent model by T. crassiceps WFU as the most affordable experimental models with taeniids.

Purpose To investigate the immuno-histological evidences in viable and non-viable hydatid cysts obtained from naturally infected camels. Methods A cohort study (February 2018–December 2019), a total of 15 hydatidosis-infected camels from slaughter houses in Cairo were involved. Specimens were investigated for parasite viability, liver histological changes, IL-17A cytokine immunohistochemical expressions in the adventitial layer, and the anti-nuclear antibodies (ANAs) immunofluorescent expression in the metacestode's structures. Real-Time Quantitative -Morphocytometry and SPSS were utilized. Results Multi-focal lesions and high viability were found in 60% of the cases. Overall accumulation of collagen associated the parasite establishment that involved infiltrations of mononuclear cells with significantly increased IL-17A expression. Interestingly, the ANAs appeared to have a role in the immune-defense against the metacestode showing different patterns. ANAs production correlated with IL-17A expression and the viability of the parasite. Conclusion IL-17A responses in hydatidosis is associated with collagen deposition and ANA production as a sort of anti-parasite immunity in a viability dependent manner.

E. granulosus is a cestode that causes Cystic Echinococcosis (CE), a zoonotic disease with worldwide presence. The immune response generated by the host against the metacestode induces a permissive Th2 response, as opposed to pro-inflammatory Th1 response. In this view, mixed Th2 and regulatory responses allow parasite survival. Overall, larval Echinococcus infections induce strong regulatory responses. Fasciola hepatica, another common helminth parasite, represents a major infection in cattle. Co-infection with different parasite species in the same host, polyparasitism, is a common occurrence involving E. granulosus and F. hepatica in cattle. 'While it is known that infection with F. hepatica also triggers a polarized Th2/Treg immune response, little is reported regarding effects on the systemic immune response of this example of polyparasitism. F. hepatica also triggers immune responses polarized to the Th2/ Treg spectrum. Serum samples from 107 animals were analyzed, and were divided according to their infection status and Echinococcal cysts fertility. Cytokines were measured utilizing a Milliplex Magnetic Bead Panel to detect IFN-γ, IL-1, IL-2, IL-4, IL-6, IL-10, IL-12 and IL-18. Cattle infected only with F. hepatica had the highest concentration of every cytokine analyzed, with both 4.24 and 3.34-fold increases in IL-10 and IL-4, respectively, compared to control animals, followed by E. granulosus and F. hepatica co-infected animals with two-fold increase in IL-10 and IL-4, compared to control animals, suggesting that E. granulosus co-infection dampens the cattle Th2/Treg immune response against F. hepatica. When considering Echinococcal cyst fertility and systemic cytokine concentrations, fertile cysts had higher IFN-γ, IL-6 and IL-18 concentrations, while infertile cysts had higher IL-10 concentrations. These results show that E. granulosus co-infection lowers Th1 and Th2 cytokine serological concentration when compared to F. hepatica infection alone. E. granulosus infections show no difference in IFN-γ, IL-1, IL-2, IL-6 and IL-18 levels compared with control animals, highlighting the immune evasion mechanisms of this cestode.

Alveolar echinococcosis (AE) is a severe disease caused by the larval stage of the tapeworm Echinococcus multilocularis. Current chemotherapeutic treatment options based on benzimidazoles are of limited effectiveness, which underlines the need to find new antiechinococcosis drugs. Metformin is an antihyperglycemic and antiproliferative agent that shows activity against the related parasite Echinococcus granulosus. Hence, we assessed the in vitro and in vivo effects of the drug on E. multilocularis. Metformin exerted significant dose-dependent killing effects on in vitro cultured parasite stem cells and protoscoleces and significantly reduced the dedifferentiation of protoscoleces into metacestodes. Likewise, oral administration of metformin (50 mg/kg of body weight/day for 8 weeks) was effective in achieving a significant reduction of parasite weight in a secondary murine AE model. Our results revealed mitochondrial membrane depolarization, activation of Em-AMPK, suppression of Em-TOR, and overexpression of Em-Atg8 in the germinal layer of metformin-treated metacestode vesicles. The opposite effects on the level of active Em-TOR in response to exogenous insulin and rapamycin suggest that Em-TOR is part of the parasite’s insulin signaling pathway. Finally, the presence of the key lysosomal pathway components, through which metformin reportedly acts, was confirmed in the parasite by in silico assays. Taken together, these results introduce metformin as a promising candidate for AE treatment. Although our study highlights the importance of those direct mechanisms by which metformin reduces parasite viability, it does not necessarily preclude any additional systemic effects of the drug that might reduce parasite growth in vivo.

Polyparasitism occurs when animals harbour multiple parasites concomitantly. It is a common occurrence but is generally understudied in wild and domestic animals. Fasciola hepatica and Echinococcus granulosus, which are helminths of ungulates, frequently coinfect cattle. The effects of this particular type of polyparasitism are not well documented. The metacestode of Echinococcus granulosus is surrounded by the adventitial layer, which constitutes the host immune response to the parasite. This layer in cattle is produced by a granulomatous reaction and is involved in echinococcal cyst (EC) fertility. Due to the systemic immune-modulating abilities of Fasciola hepatica, coinfection possibly generates a favourable environment for EC growth. A total of 203 Echinococcus granulosus sensu stricto cysts were found in 82 cattle, of which 42 ECs were found in 31 animals coinfected with Fasciola hepatica. The overall infection intensity was 3 cysts per animal. Coinfection with Fasciola hepatica decreased the mean infection intensity to 1.4 cysts per animal. Regarding EC size, coinfection resulted in smaller ECs (15.91 vs 22.09 mm), especially for infertile lung cysts. The adventitial layer of ECs in coinfected animals lacked lymphoid follicles and palisading macrophages, which are generally hallmarks of the granulomatous immune response. The ECs in coinfected animals had organized laminated layers, whereas those in animals without coinfection did not. Although coinfection was not statistically associated with EC fertility, we did not find fertile cysts in the livers of coinfected animals. We concluded that coinfection with Fasciola hepatica and Echinococcus granulosus has a detrimental effect on ECs, particularly infertile cysts.

Introduction: The E. multilocularis laminated layer (LL) is a heavily glycosylated parasitic structure that plays an important role in protecting the larval stage (metacestode) of this parasite from physiological and immunological host reactions. We elaborated an experimental design with the idea to modify the (glycan) surface of the LL by a targeted digestion. This should allow the host defense to more easily recognize and attack (or kill) the parasite by immune-mediated effects. Methods: Experimentally, E. multilocularis (clone H95) metacestodes were cultured in vitro with or without addition of α1-3,4,6-galactosidase or β1-3-galactosidase in the medium. Morphological changes were subsequently measured by microscopy at different time points. Parasites were then recovered at day 5 and reinjected into mice for assessing their viability and infectious status. For finally recovered parasites, the respective load was assessed ex vivo by wet weight measurement, and host-related PD1 and IL-10 levels were determined as the key immunoregulators by using flow cytometry. Results: Our experiments demonstrated that the parasite vesicular structure can be directly destroyed by adding galactosidases into the in vitro culture system, resulting in the fact that the parasite metacestode vesicles could not anymore infect and develop in mice after this glycan digestion. Moreover, when compared to the mice inoculated with E. multilocularis metacestode without galactosidases, PD1 expression was upregulated in CD4+ Teffs from mice inoculated with E. multilocularis metacestode pretreated with β1-3-galactosidase, with a lower IL-10 secretion from CD4+ Teffs; there was no difference of PD1 and IL-10 expression levels regarding CD4+ Teff from mice inoculated with E. multilocularis metacestode pretreated with α1-3,4,6-galac­tosidase. Discussion: We raised our hypothesis that this “aborting” effect may be linked to an altered PD1 and IL-10 response fine-tuning between immunopathology and immune protection. These findings justify a continuation of these experiments upon therapeutical in vivo administration of the enzymes.

Background Alveolar (AE) and cystic echinococcosis (CE) in humans are caused by the metacestode of the tapeworms Echinococcus multilocularis and Echinococcus granulosus sensu lato (s.l.). Immunohistochemistry with the monoclonal antibodies (mAb) Em2G11, specific for AE, and the mAb EmG3, specific for AE and CE, is an important pillar of the histological diagnosis of these two infections. Our aim was to further evaluate mAb EmG3 in a diagnostic setting and to analyze in detail the localization, distribution, and impact of small particles of Echinococcus multilocularis (spems) and small particles of Echinococcus granulosus s.l. (spegs) on lymph nodes. Methodology/principal findings We evaluated the mAb EmG3 in a cohort of formalin-fixed, paraffin embedded (FFPE) specimens of AE (n = 360) and CE (n = 178). These samples originated from 156 AE-patients and 77 CE-patients. mAb EmG3 showed a specific staining of the metacestode stadium of E. multilocularis and E. granulosus s.l. and had a higher sensitivity for spems than mAb Em2G11. Furthermore, we detected spegs in the surrounding host tissue and in almost all tested lymph nodes (39/41) of infected patients. 38/47 lymph nodes of AE showed a positive reaction for spems with mAb EmG3, whereas 29/47 tested positive when stained with mAb Em2G11. Spegs were detected in the germinal centers, co-located with CD23-positive follicular dendritic cells, and were present in the sinuses. Likewise, lymph nodes with spems and spegs in AE and CE were significantly enlarged in size in comparison to the control group. Conclusions/significance mAb EmG3 is specific for AE and CE and is a valuable tool in the histological diagnosis of echinococcosis. Based on the observed staining patterns, we hypothesize that the interaction between parasite and host is not restricted to the main lesion since spegs are detected in lymph nodes. Moreover, in AE the number of spems-affected lymph nodes is higher than previously assumed. The enlargement of lymph nodes with spems and spegs points to an immunological interaction with the small immunogenic particles (spems and spegs) of Echinococcus spp., Author summary Echinococcosis is a life-threatening disease in humans that is caused by the larval stages of the tapeworms Echinococcus multilocularis and Echinococcus granulosus s.l.. For the histological diagnosis of alveolar echinococcosis (AE) and cystic echinococcosis (CE) the two monoclonal antibodies (mAb) Em2G11, specific for AE, and the mAb EmG3, specific for AE and CE are available. We have analyzed a large cohort of formalin-fixed, paraffin embedded human tissue (FFPE) specimens of AE (n = 360) and CE (n = 178) using these two antibodies. We show that the mAb EmG3 has a specific staining pattern of the metacestode stadium of E. multilocularis and E. granulosus s.l. while staining of the larval state of E. multilocularis is limited to the mAb Em2G11. We further identified so called “small particles of Echinococcus multilocularis” (spems) and “small particles of Echinococcus granulosus” (spegs) not only in the main lesion of AE and CE, but also in the adjacent tissue and in the vast majority of lymph nodes analyzed draining the main lesion. The lymph nodes with these microparticles are enlarged as compared to a control group. We conclude that immunochemistry with these two antibodies is a valuable adjunct for the final histological diagnosis of AE und CE. The frequent detection of microparticles of E. multilocularis and E. granulosus in lymph nodes of patients with AE and CE argue for a larger interface of interaction of the parasite with the host`s immune system than previously assumed and point to further mechanisms of these infections in humans which may be the basis for immunological reactions in the host.

Introduction: Although Cysticercus tenuicollis is one of the most economic and veterinary important parasite in Iraq, scanty molecular characterization exists for this helminth. This study aimed to determine the prevalence and molecular description of C. tenuicollis isolates from sheep in Kalar district of Iraq. Methodology: A total of 2,906 slaughtered sheep were examined post-mortem. Up to 20 samples of C. tenuicollis was extracted and amplified using mitochondrial COX1 gene. Results: The overall prevalence rate was 6.88%, and female sheep recorded higher rate of infection (24.35%) than male (6.16%) with significant difference (p

Neurocysticercosis (NCC) is infection by cestode Taenia solium/pork tapeworm. Sero-diagnosis of NCC is still a challenge. Radiological imaging (CT/MRI) are cost intensive, requires technical expertise and resource intensive. Hence, its availability is restricted in endemic zone. Existing Enzyme electro immune transfer blot (EITB) antigens are difficult to make and is not standardized for endemic population. Therefore, there is a definite need for easy and reliable EITB tool.T. solium metacestode were harvested from naturally infected swine post slaughter. The cyst fluid/vesicular fluid was aspirated and processed with ultracentrifugation and immune blot was performed with this antigen.A total of 406 cases [rural 256 (NCC 78, seizures other than NCC 108 and healthy controls 70); urban 150 (NCC 41, seizures other than NCC 59 and healthy controls 50)] were enrolled. Positive EITB (detection of band50 kDa) was significantly associated with NCC patients of urban population only (p 0.001) but not in rural populations (p = 0.292). However, identification of 15 kDa band had significant association with NCC both in urban and rural populations with overall sensitivity of 91.5% and specificity of 91.6%. Presence of 35 kDa band was associated with multiple NCC (p 0.001). The study shows that 15 kDa reactive band on EITB is highly sensitive and specific for diagnosis of NCC in endemic population.Presence of 35 kDa band on EITB was associated with infection by multiple cysticerci. The observations demand purification of cyst fluid antigens to develop simple and easy to execute test in field studies.

Echinococcosis are considered to be potentially lethal zoonotic diseases that cause serious damage to hosts. The metacestode of Echinococcus multilocularis and E. granulosus can result in causing the alveolar and cystic echinococcoses, respectively. Recent studies have shown that non-coding RNAs are widely expressed in Echinococcus spp. and hosts. In this review, the two main types of non-coding RNAs—long non-coding RNAs (lncRNAs) and microRNAs (miRNAs)—and the wide-scale involvement of these molecules in these parasites and their hosts were discussed. The expression pattern of miRNAs in Echinococcus spp. is species- and developmental stage-specific. Furthermore, common miRNAs were detected in three Echinococcus spp. and their intermediate hosts. Here, we primarily focus on recent insights from transcriptome studies, the expression patterns of miRNAs and lncRNAs, and miRNA-related databases and techniques that are used to investigate miRNAs in Echinococcus and echinococcosis. This review provides new avenues for screening therapeutic and diagnostic markers.

The current chemotherapeutical treatment against alveolar echinococcosis relies exclusively on benzimidazoles, which are not parasiticidal and can induce severe toxicity. There are no alternative treatment options. To identify novel drugs with activity against Echinococcus multilocularis metacestodes, researchers have studied potentially interesting drug targets (e.g. the parasite's energy metabolism), and/or adopted drug repurposing approaches by undertaking whole organism screenings. We here focus on drug screening approaches, which utilize an in vitro screening cascade that includes assessment of the drug-induced physical damage of metacestodes, the impact on metacestode viability and the viability of isolated parasite stem cells, structure-activity relationship (SAR) analysis of compound derivatives, and the mode of action. Finally, once in vitro data are indicative for a therapeutic window, the efficacy of selected compounds is assessed in experimentally infected mice. Using this screening cascade, we found that the anti-malarial mefloquine was active against E. multilocularis metacestodes in vitro and in vivo. To shed more light into the mode of action of mefloquine, SAR analysis on mefloquine analogues was performed. E. multilocularis ferritin was identified as a mefloquine-binding protein, but its precise role as a drug target remains to be elucidated. In mice that were infected either intraperitoneally with metacestodes or orally with eggs, oral treatment with mefloquine led to a significant reduction of parasite growth compared to the standard treatment with albendazole. However, mefloquine was not acting parasiticidally. Assessment of mefloquine plasma concentrations in treated mice showed that levels were reached which are close to serum concentrations that are achieved in humans during long-term malaria prophylaxis. Mefloquine might be applied in human AE patients as a salvage treatment. Future studies should focus on other repurposed anti-infective compounds (MMV665807, niclosamide, atovaquone), which showed stronger in vitro activity against E. multilocularis than mefloquine., Graphical abstract Image 1, Highlights • Anti-infective drugs are repurposed for identification of novel drugs against AE. • Mefloquine is active against E. multilocularis in vitro and in mice. • The energy metabolism of E. multilocularis is a target for future drug development. • MMV665807 and niclosamide are highly active against E. multilocularis in vitro.

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.

Alveolar echinococcosis (AE) is a zoonotic parasitosis caused by larvae of the fox tapeworm, Echinococcus multilocularis. E. multilocularis is distributed widely in the Northern hemisphere, causing serious health problems in various animals and humans. E. multilocularis, like other cestodes, lacks a digestive tract and absorbs essential nutrients, including glucose, across the syncytial tegument on its external surface. Therefore, it is hypothesized that E. multilocularis uses glucose transporters on its surface similar to a closely-related species, Taenia solium. Based on this hypothesis, we cloned and characterized glucose transporter homologues from E. multilocularis. As a result, we obtained full-length sequences of 2 putative glucose transporter genes (EmGLUT1 and EmGLUT2) from E. multilocularis. In silico analysis predicted that these were classified in the solute carrier family 2 group. Functional expression analysis using Xenopus oocytes demonstrated clear uptake of 2-deoxy-D-glucose (2-DG) by EmGLUT1, but not by EmGLUT2 in this experimental system. EmGLUT1 was shown to have relatively high glucose transport activity. Further analyses using the Xenopus oocyte system revealed that 2-DG uptake of EmGLUT1 did not depend on the presence or concentration of Na+ nor H+, respectively. Immunoblot analyses using cultured metacestode, ex vivo protoscolex, and adult worm samples demonstrated that both EmGLUTs were stably expressed during each developmental stage of the parasite. Based on the above-mentioned findings, we conclude that EmGLUT1 is a simple facilitated glucose transporter and possibly plays an important role in glucose uptake by E. multilocularis throughout its life cycle.

Tapeworms of the family Gryporhynchidae are endoparasites of fish-eating birds distributed worldwide. Currently the family contains 16 genera classified on the basis of the morphology of the rostellar apparatus, rostellar hooks and strobilar anatomy. However, the phylogenetic relationships among the genera are still unknown. In this study, sequences of the near complete 18S (SSU) and 28S (LSU) from rDNA of 13 species of gryporhynchids (adult specimens) representing eight genera (Cyclustera,Dendrouterina,Glossocercus, Gryporhynchidae gen. sp.,Neovalipora,Paradilepis,Parvitaenia,Valipora) and one species of metacestode from fish (Neovalipora) were generated. Additionally, sequences of metacestodes of the generaAmirthalingamia,Neogryporhynchus,Paradilepis,ParvitaeniaandValiporafrom Africa recently added to the GenBank database were analysed. Phylogenetic relationships were inferred using maximum-likelihood (ML) and Bayesian inference of each (SSU and LSU) dataset. The phylogenetic analyses indicated that the family Gryporhynchidae is a well-supported monophyletic group within the Cyclophyllidea. The trees inferred with SSU and LSU datasets had similar topologies and suggested that the generaGlossocercus(two species sequenced) andParadilepis(four spp.) are monophyletic. In contrast,Dendrouterina,ParvitaeniaandValiporaare paraphyletic, suggesting that the species composition of these genera should be critically reviewed. Interestingly, species of the genera that use the same groups of definitive hosts such as herons (Ardeidae), cormorants (Phalacrocoracidae) and ibis (Threskiornithidae) are together in the phylogenetic tree, even though they differ markedly from each other in some morphological characters, especially shape and size of rostellar hooks.

Cestodes are emerging agents of severe opportunistic infections among immunocompromised patients. We describe the first case of human infection, with the recently-proposed genus Versteria causing an invasive, tumor-like hepatic infection with regional and distant extension in a 53-year-old female kidney transplant recipient from Atlantic Canada.

Alveolar echinococcosis (AE) is a zoonotic disease that is deadly if left untreated. AE is caused by the larval metacestode stage of the cestode Echinococcus multilocularis. Better knowledge on the host-parasite interface could yield novel targets for improvement of the treatment against AE. We analyzed culture media incubated with in vitro grown E. multilocularis metacestodes by 1H nuclear magnetic resonance spectroscopy to identify the unknown metabolic footprint of the parasite. Moreover, we quantitatively analyzed all amino acids, acetate, glucose, lactate, and succinate in time-course experiments using liquid chromatography and enzymatic assays. The E. multilocularis metacestodes consumed glucose and, surprisingly, threonine and produced succinate, acetate, and alanine as major fermentation products. The metabolic composition of vesicle fluid (VF) from in vitro grown E. multilocularis metacestodes was different from parasite-incubated culture medium with respect to the abundance, but not the spectrum, of metabolites, and some metabolites, in particular amino acids, accumulated in the VF. Overall, this study presents the first characterization of the in vitro metabolic footprint of E. multilocularis metacestodes and VF composition, and it provides the basis for analyses of potentially targetable pathways for future drug development.

Cysticercosis is a worldwide parasitic disease of humans and pigs principally caused by infection with the larvae of the pork tapeworm Taenia solium. Through the use of the recently-made-available T. solium genome, we identified a gene within a novel 1448 bp ORF that theoretically encodes for a 433 amino acid-long protein and predicted to be an α-enolase closely related to enolases of other flatworms. Additional bioinformatic analyses revealed a putative plasminogen-binding region on this protein, suggesting a potential role for this protein in pathogenesis. On this basis, we isolated the mRNA encoding for this presumptive enolase from T. solium metacestodes and reverse-transcribed it into cDNA before subsequently cloning and expressing it in both E. coli (rEnoTs) and insect cells (rEnoTsBac), in a 6xHis tagged manner. The molecular weights of these two recombinant proteins were ∼48 and ∼50 kDa, respectively, with the differences likely attributable to differential glycosylation. We used spectrophotometric assays to confirm the enolase nature of rEnoTs as well as to measure its enzymatic activity. The resulting estimates of specific activity (60.000 U/mg) and Km (0.091 mM) are quite similar to the catalytic characteristics of enolases of other flatworms. rEnoTs also exhibited high immunogenicity, eliciting a strong polyclonal antibody response in immunized rabbits. We subsequently employed rEnoTsBac for use in an ELISA aimed at discriminating between healthy pigs and those infected with T. solium. This diagnostic assay exhibited a sensitivity of 88.4% (95% CI, 74.92%–96.11%) and a specificity of 83.7% (95% CI: 69.29%–93.19%). In conclusion, this study reports on and enzymatically characterizes a novel enolase from T. solium metacestode, and shows a potential use as an immunodiagnostic for porcine cysticercosis.

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.

A captive-born adult female Nilgiri langur (Semnopithecus johnii) developed an edematous swelling of the left thigh and a firm mass around the right ankle joint. The animal also suffered from lethargy and anorexia and was euthanized because of poor general condition. Necropsy revealed that the skeletal muscle of the left thigh had been replaced by a multilocular cystic mass containing numerous sand-grain–sized whitish structures. Small cysts were also present in the lung and the myocardium. The mass of the right ankle joint was histologically consistent with a myxosarcoma. In contrast, the cystic masses from the left thigh, the lung, and the myocardium represented metacestode tissue with evidence of numerous larval cestodes consistent with cysticerci. Cysticerci showed morphological characteristics of Cysticercus longicollis, the larval form of Taenia crassiceps, which was confirmed by genetic analysis. This is the first documented case of a Taenia crassiceps cysticercosis in an Old World monke...