Stem cell proliferation and differentiation during larval metamorphosis of the model tapeworm Hymenolepis microstoma.

Background: Tapeworm larvae cause important diseases in humans and domestic animals. During infection, the first larval stage undergoes a metamorphosis where tissues are formed de novo from a population of stem cells called germinative cells. This process is difficult to study for human pathogens, as these larvae are infectious and difficult to obtain in the laboratory. Methods: In this work, we analyzed cell proliferation and differentiation during larval metamorphosis in the model tapeworm Hymenolepis microstoma, by in vivo labelling of proliferating cells with the thymidine analogue 5-ethynyl-2'-deoxyuridine (EdU), tracing their differentiation with a suite of specific molecular markers for different cell types. Results: Proliferating cells are very abundant and fast-cycling during early metamorphosis: the total number of cells duplicates every ten hours, and the length of G2 is only 75 minutes. New tegumental, muscle and nerve cells differentiate from this pool of proliferating germinative cells, and these processes are very fast, as differentiation markers for neurons and muscle cells appear within 24 hours after exiting the cell cycle, and fusion of new cells to the tegumental syncytium can be detected after only 4 hours. Tegumental and muscle cells appear from early stages of metamorphosis (24 to 48 hours post-infection); in contrast, most markers for differentiating neurons appear later, and the detection of synapsin and neuropeptides correlates with scolex retraction. Finally, we identified populations of proliferating cells that express conserved genes associated with neuronal progenitors and precursors, suggesting the existence of tissue-specific lineages among germinative cells. Discussion: These results provide for the first time a comprehensive view of the development of new tissues during tapeworm larval metamorphosis, providing a framework for similar studies in human and veterinary pathogens.

High Prevalence of Hymenolepis (Rodentolepis) nana in Amateur Breeding Facilities of Chinchillas (Chinchilla lanigera) and Sugar Gliders (Petaurus breviceps) from Italy.

PURPOSE: Hymenolepis (Rodentolepis) nana is an enteric tapeworm globally widespread in wild and captive rodents. The survey was carried out in three chinchilla breeding facilities and in one sugar glider breeding facility in Central and Southern Italy. METHODS: One hundred and four chinchilla fecal pools and 40 sugar glider fecal pools were collected from cages housing breeding pairs or females with their offspring. Fecal samples were examined with a qualitative and quantitative copromicroscopical approach. Hymenolepis eggs were identified based on morphological and morphometrical features, and the average number of eggs per gram (EPG) of feces was estimated. RESULTS: Hymenolepis nana eggs were detected in 39.42% of chinchilla samples with an average of 16.33 EPG; whereas, all of the sugar glider samples tested positive with an average of 454.18 EPG. Neither helminth eggs nor protozoan cysts/oocysts were detected in any of the breeding facilities. CONCLUSION: The results clearly show a diffuse parasitism in both chinchillas and sugar gliders, and should be taken into consideration given the rising popularity of these two pet species combined with the zoonotic complications presented by H. nana.

Gallic acid and Catechin induce morphological alterations on the zoonotic parasite Hymenolepis diminuta.

Tapeworm infections cause insidious and irreversible effects in the infected individuals and some of them have already shown resistance to available drugs. A search for alternative treatment is urgently required. Phenolic compounds are amongst the most researched natural substances for their medicinal use. The present study aims to determine anthelmintic efficacy of two polyphenols Gallic acid and Catechin against the zoonotic rat tapeworm Hymenolepis diminuta. Both compounds are potent anti-oxidants and play major roles in combating pathogens, while their anthelmintic property according to our knowledge is yet to be explored. The parasite model H. diminuta was procured from intestine of infected rats raised in our laboratory. Two sets of parasites were treated in vitro with 5, 10, 20 and 40 mg/ml concentrations of each Gallic Acid and Catechin separately, another set of parasites were treated with standard dose of Praziquantel in RPMI 1640, while still another set of worms were kept in RPMI 1640 at 37 ± 10C with 1% Dimethyl sulfoxide as control. Motility and structural alterations were the parameters assessed for anthelmintic efficacy of the compounds. After paralysis the worms were processed for morphological, histological, and ultrastructural study and observed under light and electron microscope. Dose-dependent efficacy was observed in both compounds. Shrinkage of suckers, deformed proglottids and architectural alteration of the tegument were observed throughout the body of treated parasites compared to control. Although in terms of time taken for paralysis and mortality Gallic acid was more effective than Catechin, the degree of morphological aberrations caused were almost similar, except histological alteration was more in Catechin treated worms than in Gallic acid. Nevertheless, both Gallic acid and Catechin are suggested to possess anthelmintic efficacy besides other health benefits but extended studies are required to compare their efficacy.

Hymenolepis diminuta, phylogenetic analyses of nuclear rRNA + ITS and mitochondrial cox1 and its infections in non-human primates.

Hymenolepis diminuta is a tapeworm commonly found worldwide in small rodents such as rats with occasional reports in other definitive hosts such as primates including chimpanzees and humans. It has not been reported in African green monkey (AGM, Chlorocebus sabaeus), and the parasite's molecular phenotype and phylogeny remain primitively sketchy. The aims of the current study were to determine if H. diminuta infected AGMs, to molecularly characterize H. diminuta and to review its infection in non-human primates. Feces of AGMs were examined visually for adult helminths and microscopically for eggs using centrifugation flotation. Total DNA extracted from eggs was amplified by PCR followed by DNA sequencing of targeted sequences of nuclear rRNA + internal transcribed spacers (ITS) and mitochondrial cox1. Phylogenetic analyses were performed. The DNA sequences of both nuclear rRNA + ITS and mitochondrial cox1 showed more than 98% and 99% identity to the known sequences respectively. Hymenolepis diminuta has been reported in various non-human primates with the highest prevalence of 38.5% in the white-headed capuchin monkey. The study presented here confirms that this tapeworm is capable of infecting various species of non-human primates with the first report of infections in AGM. Phylogenetic analyses of rRNA + ITS and mitochondrial cox1 demonstrated three separated clades I, II and III with the newly described AGM1 isolate belonging to the clade I. Whether these differences are at species level remains to be confirmed.

Trans-splicing in the cestode Hymenolepis microstoma is constitutive across the life cycle and depends on gene structure and composition.

Spliced leader (SL) trans-splicing is a key process during mRNA maturation of many eukaryotes, in which a short sequence (SL) is transferred from a precursor SL-RNA into the 5' region of an immature mRNA. This mechanism is present in flatworms, in which it is known to participate in the resolution of polycistronic transcripts. However, most trans-spliced transcripts are not part of operons, and it is not clear if this process may participate in additional regulatory mechanisms in this group. In this work, we present a comprehensive analysis of SL trans-splicing in the model cestode Hymenolepis microstoma. We identified four different SL-RNAs which are indiscriminately trans-spliced to 622 gene models. SL trans-splicing is enriched in constitutively expressed genes and does not appear to be regulated throughout the life cycle. Operons represented at least 20% of all detected trans-spliced gene models, showed conservation to those of the cestode Echinococcus multilocularis, and included complex loci such as an alternative operon (processed as either a single gene through cis-splicing or as two genes of a polycistron). Most insertion sites were identified in the 5' untranslated region (UTR) of monocistronic genes. These genes frequently contained introns in the 5' UTR, in which trans-splicing used the same acceptor sites as cis-splicing. These results suggest that, unlike other eukaryotes, trans-splicing is associated with internal intronic promoters in the 5' UTR, resulting in transcripts with strong splicing acceptor sites without competing cis-donor sites, pointing towards a simple mechanism driving the evolution of novel SL insertion sites.

Hymenolepidid cestodes: Diversity, morphological and molecular characterization of a new species, and phylogeny of parasitic species of rodents from North and South America.

Rodents are hosts of a wide diversity of cestodes. Fifteen genera included in the family Hymenolepididae parasitize rodents, and only four of these genera have been recorded from the Neotropical region. The purpose of this paper is to update species of Hymenolepididae from rodents, describe a new species of Hymenolepis based on morphological and molecular characterization (ITS1 rDNA and cox1 mtDNA), comparing the features among the species from North and South American rodents, and provide phylogenetic inferences of Hymenolepididae from rodents based on sequences available in the GenBank. Rodents were collected in the Parque Provincial Ernesto Tornquist, Buenos Aires, Argentina. Hymenolepis ivanovae n. sp. differs from other Hymenolepis species registered from North and South American rodents by body size, scolex, suckers, cirrus sac, cirrus, testes, and eggs, among others. Comparative morphometric data for Hymenolepis species from North and South American rodents is provided. Molecular analyses place H. ivanovae n. sp. within the genus Hymenolepis with strong support, and show it close to species of zoonotic importance. The new species is the first species of Hymenolepis described from Sigmodontinae rodents.

Detection of enteroparasites in foliar vegetables commercialized in street- and supermarkets in Aparecida de Goiânia, Goiás, Brazil / Detecção de enteroparasitas em hortaliças comercializadas em feiras e supermercados em Aparecida de Goiânia, Goiás, Brasil

Foliar vegetables contaminated with fecal residues are an important route of transmission of intestinal parasites to humans. The aim of this study was to evaluate the presence of parasitic forms of protozoa and helminths on lettuces (Lactuca sativa) and collard greens (Brassica oleracea) sold in street- and supermarkets in the city of Aparecida de Goiânia, Goiás, Brazil. A total of 30 samples of each vegetable (15 samples from each supermarkets and street markets) was analyzed. All samples were processed by spontaneous sedimentation method and centrifugal flotation. In 45% of the samples, immature forms of intestinal parasites were identified with 66.7% helminths eggs and 33.3% protozoan cysts or oocysts. Significantly more lettuce samples were contaminated with eggs, cysts or oocyst of at least one parasite than collard green samples (U=216; Z=-3.45; P <0.001). The parasitic forms were identified morphologically up to the family level with eggs of Ancylostomatidae, Strongyloididae, Ascarididae and Taeniidae, or oocysts of Eimeriidae, to the genus with Cystoisospora sp. and Toxocara sp., and to the species level with Cystoisospora canis, Dipylidium caninum and Hymenolepis nana. The presence of these infective agents in lettuce and collard green from both street- and supermarkets highlights the high risk of spreading parasites by eating raw vegetables sold in Aparecida de Goiânia.

Hortaliças contaminadas com resíduos fecais são importantes vias de transmissão de parasitas intestinais ao homem. O objetivo deste estudo foi avaliar a presença de formas parasitárias de protozoários e helmintos em alface (Lactuca sativa) e couve (Brassica oleracea) vendidas em feiras e supermercados na cidade de Aparecida de Goiânia, Goiás, Brasil. Um total de 30 amostras de cada vegetal (15 amostras de supermercados e 15 de feiras livres) foi analisado. Todas as amostras foram processadas pelos métodos de sedimentação espontânea e centrífugo-flutuação. Em 45% das amostras foram identificadas formas imaturas de parasitas intestinais sendo 66,7% ovos de helmintos e 33,3% de cistos ou oocistos de protozoários. Significativamente, mais amostras de alface estavam contaminadas com ovos, cistos ou oocistos de pelo menos um parasita do que as amostras de couve (U = 216; Z = -3,45; P <0,001). As formas parasitárias foram identificadas morfologicamente ao nível de família com ovos de Ancylostomatidae, Strongyloididae, Ascarididae e Taeniidae ou oocistos de Eimeriidae, dos gêneros Cystoisospora sp. e Toxocara sp., e ao nível de espécies com Cystoisospora canis, Dipylidium caninum e Hymenolepis nana. A presença desses agentes infecciosos em alface e couve, provenientes tanto de feiras quanto de supermercados, ressalta o alto risco de veiculação de parasitas pela ingestão de hortaliças cruas comercializadas em Aparecida de Goiânia.

Prevalence of intestinal parasitic diseases in school children of rural areas of district Lower Dir, Pakistan / Prevalência de doenças parasitárias intestinais em escolares de áreas rurais do distrito de Lower Dir, Paquistão

Present study was conducted among school children to recognize the prevalence of IPIs in rural communities of district Dir Lower, Pakistan. A sum of 324 samples of stool were collected (210 boys and 114 girls). Used direct smear method and formol ether sedimentation concentration technique for processing the samples. The result shows that 82% (n=266) were found infected comprised 64.8% male and 35.1% females. Children of the age group 10 to 12 years were found extremely infected 94.2% while 4-6 year age group were having minimum ratio of infection 72%. Current study shows mono parasitism in 50.6% of the students while 22.2% were infected with 2 species and 7.40% were infected with three species of parasites. Seven species of intestinal parasites were reported include Ascaris lumbricoid in male (n=122) 58.0% and in female (n=65) 57.0% followed by Hook worm (n=88) 41.9% and (n=44) 38.5%; Tania saginata (n=44) 20.9% and (n=24) 21.0%; Entrobius vermicularis (n=32) 15.2% and (n=16) 14.0%; Trichuris trichura (n=25) 11.9% and (n=22) 19.2%; Hymenolepis nana (n=24) 11.4% and (n=18) 15.7% and Entameoba histolytica (n=16) 7.61% and (n=14) 12.2% in male and females respectively. The study indicates that most occurring intestinal parasite in the current study were Ascaris lumbricoides 58.0% (n=122) followed by hookworms 41.9% (n=88). Male students were more infected than females in the present study.

O presente estudo foi conduzido entre crianças em idade escolar para reconhecer a prevalência de IPIs em comunidades rurais do distrito de Dir Lower, Paquistão. Foram coletadas 324 amostras de fezes (210 meninos e 114 meninas). Método de esfregaço direto usado e técnica de concentração de sedimentação de formol éter para processar as amostras. O resultado mostra que 82% (n = 266) dos infectados eram 64,8% do sexo masculino e 35,1% do feminino. Crianças da faixa etária de 10 a 12 anos foram encontradas extremamente infectadas 94,2%, enquanto a faixa etária de 4 a 6 anos apresentava proporção mínima de infecção de 72%. O estudo atual mostra monoparasitismo em 50,6% dos alunos, enquanto 22,2% estavam infectados com 2 espécies e 7,40% estavam infectados com três espécies de parasitas. Sete espécies de parasitas intestinais foram relatadas incluindo Ascaris lumbricoid em machos (n = 122) 58,0% e em fêmeas (n = 65) 57,0% seguido por anzol (n = 88) 41,9% e (n = 44) 38,5%; Tania saginata (n = 44) 20,9% e (n = 24) 21,0%; Entrobius vermicularis (n = 32) 15,2% e (n = 16) 14,0%; Trichuris trichura (n = 25) 11,9% e (n = 22) 19,2%; Hymenolepis nana (n = 24) 11,4% e (n = 18) 15,7% e Entameoba histolytica (n = 16) 7,61% e (n = 14) 12,2% em homens e mulheres, respectivamente. O estudo indica que os parasitas intestinais que mais ocorreram no presente estudo foram Ascaris lumbricoides 58,0% (n = 122), seguidos por ancilóstomos 41,9% (n = 88). Estudantes do sexo masculino foram mais infectados do que do sexo feminino no presente estudo.

Characterization of a new type of neuronal 5-HT G- protein coupled receptor in the cestode nervous system.

Cestodes are platyhelminth parasites with a wide range of hosts that cause neglected diseases. Neurotransmitter signaling is of critical importance for these parasites which lack circulatory, respiratory and digestive systems. For example, serotonin (5-HT) and serotonergic G-protein coupled receptors (5-HT GPCRs) play major roles in cestode motility, development and reproduction. In previous work, we deorphanized a group of 5-HT7 type GPCRs from cestodes. However, little is known about another type of 5-HT GPCR, the 5-HT1 clade, which has been studied in several invertebrate phyla but not in platyhelminthes. Three putative 5-HT GPCRs from Echinococcus canadensis, Mesocestoides vogae (syn. M. corti) and Hymenolepis microstoma were cloned, sequenced and bioinformatically analyzed. Evidence grouped these new sequences within the 5-HT1 clade of GPCRs but differences in highly conserved GPCR motifs were observed. Transcriptomic analysis, heterologous expression and immunolocalization studies were performed to characterize the E. canadensis receptor, called Eca-5-HT1a. Functional heterologous expression studies showed that Eca-5-HT1a is highly specific for serotonin. 5-Methoxytryptamine and α-methylserotonin, both known 5-HT GPCR agonists, give stimulatory responses whereas methysergide, a known 5-HT GPCR ligand, give an antagonist response in Eca-5-HT1a. Mutants obtained by the substitution of key predicted residues resulted in severe impairment of receptor activity, confirming that indeed, these residues have important roles in receptor function. Immunolocalization studies on the protoscolex stage from E. canadensis, showed that Eca-5-HT1a is localized in branched fibers which correspond to the nervous system of the parasite. The patterns of immunoreactive fibers for Eca-5-HT1a and for serotonin were intimately intertwined but not identical, suggesting that they are two separate groups of fibers. These data provide the first functional, pharmacological and localization report of a serotonergic receptor that putatively belongs to the 5-HT1 type of GPCRs in cestodes. The serotonergic GPCR characterized here may represent a new target for antiparasitic intervention.

Inhibitors of protein kinases A and C impair the motility of oncospheres of the model tapeworm Hymenolepis microstoma.

The oncosphere larvae of tapeworms cyclically extend and retract their hooks during the penetration of the intestine of their intermediate hosts. The mechanisms regulating these movements are essentially unknown, in part due to the biohazardous nature of oncospheres from human pathogens. In this work, we standardized a method for the analysis of motility of hatched oncospheres (hexacanths) of the model tapeworm Hymenolepis microstoma. We used this assay to explore the relevance of protein kinases C (PKC) and A (PKA) in these processes. Pharmacological inhibition of the PKC pathway resulted in impaired larval motility. On the other hand, the PKA inhibitor H-89 potently blocked larval motility, as well as the motility of other life stages, although other inhibitors of the PKA pathway were not effective. This work represents the first study of the mechanisms that regulate the motility of oncospheres, and provides a path for further exploration.

Stage-specific transcriptomic analysis of the model cestode Hymenolepis microstoma.

Most parasitic flatworms go through different life stages with important physiological and morphological changes. In this work, we used a transcriptomic approach to analyze the main life-stages of the model tapeworm Hymenolepis microstoma (eggs, cysticercoids, and adults). Our results showed massive transcriptomic changes in this life cycle, including key gene families that contribute substantially to the expression load in each stage. In particular, different members of the cestode-specific hydrophobic ligand-binding protein (HLBP) family are among the most highly expressed genes in each life stage. We also found the transcriptomic signature of major metabolic changes during the transition from cysticercoids to adult worms. Thus, this work contributes to uncovering the gene expression changes that accompany the development of this important cestode model species, and to the best of our knowledge represents the first transcriptomic study with robust replicates spanning all of the main life stages of a tapeworm.

Extracellular vesicles secreted by model tapeworm Hymenolepis diminuta: biogenesis, ultrastructure and protein composition.

We provided the first known evidence of the presence and release of extracellular vesicles in adults of important model tapeworm Hymenolepis diminuta. Two different subtypes have been observed on the surface of the worm and among the secretory products confirmed by several microscopical methods. Proteomic analysis revealed the presence of parasite-specific proteins as well as those of the host in purified extracellular vesicles. Among the protein cargo, we identified potential drug targets, vaccine candidates and H. diminuta antigens. Finally, the protein composition further revealed proteins participating in the endosomal complex required for transport-dependent biogenesis pathway.

Complete representation of a tapeworm genome reveals chromosomes capped by centromeres, necessitating a dual role in segregation and protection.

BACKGROUND: Chromosome-level assemblies are indispensable for accurate gene prediction, synteny assessment, and understanding higher-order genome architecture. Reference and draft genomes of key helminth species have been published, but little is yet known about the biology of their chromosomes. Here, we present the complete genome of the tapeworm Hymenolepis microstoma, providing a reference quality, end-to-end assembly that represents the first fully assembled genome of a spiralian/lophotrochozoan, revealing new insights into chromosome evolution. RESULTS: Long-read sequencing and optical mapping data were added to previous short-read data enabling complete re-assembly into six chromosomes, consistent with karyology. Small genome size (169 Mb) and lack of haploid variation (1 SNP/3.2 Mb) contributed to exceptionally high contiguity with only 85 gaps remaining in regions of low complexity sequence. Resolution of repeat regions reveals novel gene expansions, micro-exon genes, and spliced leader trans-splicing, and illuminates the landscape of transposable elements, explaining observed length differences in sister chromatids. Syntenic comparison with other parasitic flatworms shows conserved ancestral linkage groups indicating that the H. microstoma karyotype evolved through fusion events. Strikingly, the assembly reveals that the chromosomes terminate in centromeric arrays, indicating that these motifs play a role not only in segregation, but also in protecting the linear integrity and full lengths of chromosomes. CONCLUSIONS: Despite strong conservation of canonical telomeres, our results show that they can be substituted by more complex, species-specific sequences, as represented by centromeres. The assembly provides a robust platform for investigations that require complete genome representation.

The tapeworm interactome: inferring confidence scored protein-protein interactions from the proteome of Hymenolepis microstoma.

BACKGROUND: Reference genome and transcriptome assemblies of helminths have reached a level of completion whereby secondary analyses that rely on accurate gene estimation or syntenic relationships can be now conducted with a high level of confidence. Recent public release of the v.3 assembly of the mouse bile-duct tapeworm, Hymenolepis microstoma, provides chromosome-level characterisation of the genome and a stabilised set of protein coding gene models underpinned by bioinformatic and empirical data. However, interactome data have not been produced. Conserved protein-protein interactions in other organisms, termed interologs, can be used to transfer interactions between species, allowing systems-level analysis in non-model organisms. RESULTS: Here, we describe a probabilistic, integrated network of interologs for the H. microstoma proteome, based on conserved protein interactions found in eukaryote model species. Almost a third of the 10,139 gene models in the v.3 assembly could be assigned interaction data and assessment of the resulting network indicates that topologically-important proteins are related to essential cellular pathways, and that the network clusters into biologically meaningful components. Moreover, network parameters are similar to those of single-species interaction networks that we constructed in the same way for S. cerevisiae, C. elegans and H. sapiens, demonstrating that information-rich, system-level analyses can be conducted even on species separated by a large phylogenetic distance from the major model organisms from which most protein interaction evidence is based. Using the interolog network, we then focused on sub-networks of interactions assigned to discrete suites of genes of interest, including signalling components and transcription factors, germline multipotency genes, and genes differentially-expressed between larval and adult worms. Results show not only an expected bias toward highly-conserved proteins, such as components of intracellular signal transduction, but in some cases predicted interactions with transcription factors that aid in identifying their target genes. CONCLUSIONS: With key helminth genomes now complete, systems-level analyses can provide an important predictive framework to guide basic and applied research on helminths and will become increasingly informative as new protein-protein interaction data accumulate.

Suppression of systemic lupus erythematosus in NZBWF1 mice infected with Hymenolepis microstoma.

Intestinal helminths induce immune suppressive responses thought to regulate inflammatory diseases including allergies and autoimmune diseases. This study was designed to evaluate whether helminthic infections suppress the natural development of systemic lupus erythematosus (SLE) in NZBWF1 mice. Infection of NZBWF1 SLE-prone mice with two nematodes failed to establish long-lasting settlement. However, the Hymenolepis microstoma (Hm) rodent tapeworm successfully established long-term parasitization of NZBWF1 mice and was used to evaluate the suppressive effects of helminth infection. Ten-month-old NZBWF1 mice developed symptoms including autoantibody generation, proteinuria, glomerular histopathology, and splenomegaly, but mice infected with Hm at 2 months of age did not show any clinical signs. Furthermore, infection with Hm reduced lymphocyte activation and increased regulatory T cells in the spleen and mesenteric lymph nodes. These results indicate that infection with Hm protects NZBWF1 mice from naturally developing SLE and suggest that pathological immunity is attenuated, presumably because of the induction of regulatory T cells.

Morphological and molecular identification of hymenolepidid cestodes in children and synanthropic rodents from rural Mexico.

Hymenolepidid cestodes of synanthropic rodents represent a risk for public health. In order to describe the occurrence of hymenolepidids in children and the role of rodents as a potential source of infection, we conducted a morphological and molecular survey on cestodes in two rural villages from Yucatan, Mexico. One hundred and thirty-five stool samples from children (64 from Paraíso and 71 from Xkalakdzonot), 233 Mus musculus (159 from Paraíso and 74 from Xkalakdzonot) and 125 Rattus rattus (7 from Paraíso and 118 from Xkalakdzonot) were analyzed for the presence of cestodes. Three hymenolepidid species were identified morphologically: Hymenolepis nana in 7.8% of children from Paraíso, Hymenolepis microstoma in 4.4% of M. musculus from Paraíso and Hymenolepis diminuta in 15.3% of R. rattus from Xkalakdzonot. The molecular characterization and phylogenetic analysis based on mitochondrial cytochrome c subunit 1 (CO1) gene and ribosomal internal transcribed spacer 1 (ITS1) region, confirmed the identity of the three cestodes isolated from Yucatan. Phylogeny of the CO1 gene identified intraspecific genetic differences within H. nana ranging from 0 to 5%, in H. microstoma from 0 to 0.4%, and in H. diminuta ranged from 0 to 6.5% which suggests, the presence of complex species within H. nana and H. diminuta infecting humans and rodents, as reported by other authors. Based on the morphological and molecular results, and the epidemiological evidence, infections with H. nana suggest a non-zoonotic transmission; however, the presence of H. microstoma and H. diminuta in synanthropic rodents serve as a possible source for human infection.

Divergent Axin and GSK-3 paralogs in the beta-catenin destruction complexes of tapeworms.

The Wnt/beta-catenin pathway has many key roles in the development of animals, including a conserved and central role in the specification of the primary (antero-posterior) body axis. The posterior expression of Wnt ligands and the anterior expression of secreted Wnt inhibitors are known to be conserved during the larval metamorphosis of tapeworms. However, their downstream signaling components for Wnt/beta-catenin signaling have not been characterized. In this work, we have studied the core components of the beta-catenin destruction complex of the human pathogen Echinococcus multilocularis, the causative agent of alveolar echinococcosis. We focused on two Axin paralogs that are conserved in tapeworms and other flatworm parasites. Despite their divergent sequences, both Axins could robustly interact with one E. multilocularis beta-catenin paralog and limited its accumulation in a heterologous mammalian expression system. Similarly to what has been described in planarians (free-living flatworms), other beta-catenin paralogs showed limited or no interaction with either Axin and are unlikely to function as effectors in Wnt signaling. Additionally, both Axins interacted with three divergent GSK-3 paralogs that are conserved in free-living and parasitic flatworms. Axin paralogs have highly segregated expression patterns along the antero-posterior axis in the tapeworms E. multilocularis and Hymenolepis microstoma, indicating that different beta-catenin destruction complexes may operate in different regions during their larval metamorphosis.

Identification and expression profiling of microRNAs in Hymenolepis.

Tapeworms (cestodes) of the genus Hymenolepis are the causative agents of hymenolepiasis, a neglected zoonotic disease. Hymenolepis nana is the most prevalent human tapeworm, especially affecting children. The genomes of Hymenolepis microstoma and H. nana have been recently sequenced and assembled. MicroRNAs (miRNAs), a class of small non-coding RNAs, are principle regulators of gene expression at the post-transcriptional level and are involved in many different biological processes. In previous work, we experimentally identified miRNA genes in the cestodes Echinococcus, Taenia and Mesocestoides. However, current knowledge about miRNAs in Hymenolepis is limited. In this work we described for the first known time the expression profile of the miRNA complement in H. microstoma, and discovered miRNAs in H. nana. We found a reduced complement of 37 evolutionarily conserved miRNAs, putatively reflecting their low morphological complexity and parasitic lifestyle. We found high expression of a few miRNAs in the larval stage of H. microstoma that are conserved in other cestodes, suggesting that these miRNAs may have important roles in development, survival and for host-parasite interplay. We performed a comparative analysis of the identified miRNAs across the Cestoda and showed that most of the miRNAs in Hymenolepis are located in intergenic regions, implying that they are independently transcribed. We found a Hymenolepis-specific cluster composed of three members of the mir-36 family. Also, we found that one of the neighboring genes of mir-10 was a Hox gene as in most bilaterial species. This study provides a valuable resource for further experimental research in cestode biology that might lead to improved detection and control of these neglected parasites. The comprehensive identification and expression analysis of Hymenolepis miRNAs can help to identify novel biomarkers for diagnosis and/or novel therapeutic targets for the control of hymenolepiasis.

Molecular data confirm the taxonomic position of Hymenolepis erinacei (Cyclophyllidea: Hymenolepididae) and host switching, with notes on cestodes of Palaearctic hedgehogs (Erinaceidae).

The cestode Hymenolepis erinacei is regarded as a widely distributed parasite in European hedgehogs of the genus Erinaceus, although the taxonomic position of this hymenolepidid has been debated for a considerable period of time. We present the first molecular data for this cestode, including partial DNA sequences of mitochondrial 16S and nuclear 28S ribosomal genes. Molecular phylogenetic analysis clusters H. erinacei in one clade together with representatives of the genus Hymenolepis from rodents. Characteristic morphological features, including the oval embryophore without filaments and shape of the embryonic hooks of H. erinacei are described. Features of these cestode eggs are proposed as a basis for non-invasive detection of parasitic infections in small mammal populations. The present study explores phylogenetic relationships within the genus Hymenolepis and the host switching related to H. erinacei. Cases of host switching in other genera of the family Hymenolepididae are reviewed. A short critical review of cestodes parasitizing hedgehogs in the Palaearctic is presented.

First survey on zoonotic helminthosis in urban brown rats (Rattus norvegicus) in Spain and associated public health considerations.

The brown rat, Rattus norvegicus, with a worldwide distribution, is the most commensal species among synanthropic rodents, since its main habitat, in urban as well as in rural areas, is always linked to humans. Therefore, people living in close proximity to rodent populations can be exposed to infection. Whereas bacteria and viruses are the best known rat-associated zoonoses in urban environments, the role of brown rats as reservoirs for helminth parasites and the associated risk for humans are less well known. Specifically, this role has not been analyzed in Spain to date. A total of 100 R. norvegicus trapped in the sewage system (n = 85), and parks (n = 15) of Barcelona was examined. The overall prevalence of helminth infection was 85%. The helminths found were Hymenolepis nana (17%), H. diminuta (33%) (Cestoda), Calodium hepaticum (17%), Eucoleus gastricus (28%), Aonchotheca annulosa (12%), Trichosomoides crassicauda (7%), Nippostrongylus brasiliensis (46%), Heterakis spumosa (62%), Gongylonema neoplasticum (20%) (Nematoda) and Moniliformis moniliformis (6%) (Acanthocephala). Five of the ten helminth species are considered zoonotic parasites, with rats acting as reservoirs for human infection, i.e. H. nana, H. diminuta, C. hepaticum, G. neoplasticum and M. moniliformis. G. neoplasticum and M. moniliformis are reported for the first time in urban rats in Europe. H. nana, H. diminuta and C. hepaticum are the most widespread species in European cities. For H. nana and C. hepaticum, rats act as effective spreaders of the human infective stage (eggs). For H. diminuta, G. neoplasticum and M. moniliformis, rats act as indirect reservoirs of the zoonoses since the eggs shed by the rats are infective for their insect intermediate hosts only. Medical practitioners need to be made aware of the range of parasites carried by rats, as there is a realistic likelihood that ill health currently caused by rat infestations may be misdiagnosed.