RESUMO
BACKGROUND: Tapeworms are parasitic flatworms that independently evolved a segmented body plan, historically confounding comparisons with other animals. Anteroposterior (AP) patterning in free-living flatworms and in tapeworm larvae is associated with canonical Wnt signaling and positional control genes (PCGs) are expressed by their musculature in regionalized domains along the AP axis. Here, we extend investigations of PCG expression to the adult of the mouse bile-duct tapeworm Hymenolepis microstoma, focusing on the growth zone of the neck region and the initial establishment of segmental patterning. RESULTS: We show that the adult musculature includes new, segmental elements that first appear in the neck and that the spatial patterns of Wnt factors are consistent with expression by muscle cells. Wnt factor expression is highly regionalized and becomes AP-polarized in segments, marking them with axes in agreement with the polarity of the main body axis, while the transition between the neck and strobila is specifically demarcated by the expression domain of a Wnt11 paralog. CONCLUSION: We suggest that segmentation could originate in the muscular system and participate in patterning the AP axis through regional and polarized expression of PCGs, akin to the gene regulatory networks employed by free-living flatworms and other animals.
RESUMO
Ligula intestinalis (Cestoda: Diphyllobothriidae) is an emerging model organism for studies on parasite population biology and host-parasite interactions. However, a well-resolved genome and catalogue of its gene content has not been previously developed. Here, we present the first genome assembly of L. intestinalis, based on Oxford Nanopore Technologies, Illumina and Omni-C sequencing methodologies. We use transcriptome profiling to compare plerocercoid larvae and adult worms and identify differentially expressed genes (DEGs) associated with these life stages. The genome assembly is 775.3 mega (M)bp in size, with scaffold N50 value of 118 Mbp and encodes 27 256 predicted protein-coding sequences. Over 60% of the genome consists of repetitive sequences. Synteny analyses showed that the 10 largest scaffolds representing 75% of the genome display high correspondence to full chromosomes of cyclophyllidean tapeworms. Mapping RNA-seq data to the new reference genome, we identified 3922 differentially expressed genes in adults compared with plerocercoids. Gene ontology analyses revealed over-represented genes involved in reproductive development of the adult stage (e.g. sperm production) and significantly enriched DEGs associated with immune evasion of plerocercoids in their fish host. This study provides the first insights into the molecular biology of L. intestinalis and provides the most highly contiguous assembly to date of a diphyllobothriid tapeworm useful for population and comparative genomic investigations of parasitic flatworms.
Assuntos
Cestoides , Infecções por Cestoides , Animais , Masculino , Sêmen , Cestoides/genética , Infecções por Cestoides/parasitologia , Peixes/genética , Perfilação da Expressão Gênica , TranscriptomaRESUMO
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.
Assuntos
Cromossomos/metabolismo , Elementos de DNA Transponíveis/genética , Genoma Helmíntico , Hymenolepis/genética , Sintenia , Animais , Centrômero/metabolismo , Segregação de CromossomosRESUMO
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.
Assuntos
Hymenolepis/genética , Mapeamento de Interação de Proteínas , Proteoma/genética , Animais , Bases de Dados Genéticas , Eucariotos/classificação , Eucariotos/genética , Eucariotos/metabolismo , Redes Reguladoras de Genes , Genoma Helmíntico/genética , Proteínas de Helminto/genética , Proteínas de Helminto/metabolismo , Hymenolepis/classificação , Hymenolepis/metabolismo , ProbabilidadeRESUMO
Chimaeras, or ratfishes, are the only extant group of holocephalan fishes and are the sole host group of gyrocotylidean cestodes, which represent a sister group of the true tapeworms (Eucestoda). These unique, non-segmented cestodes have been known since the 1850s and multiple species and genera have been erected despite a general agreement that the delineation of species on the basis of morphology is effectively impossible. Thus, in the absence of molecular studies, the validity of gyrocotylid taxa and their specific host associations has remained highly speculative. Here we report the presence of Gyrocotyle spp. from rarely-caught deep-sea chimaeras collected in the North-East Atlantic, and describe two new species: G. haffii n. sp. from the bent-nose chimaera, Harriota raleighana Goode & Bean, and G. discoveryi n. sp. from the large-eyed rabbit fish, Hydrolagus mirabilis (Collett). Nuclear ribosomal sequence data were generated for individual parasites taken from different host species collected on different dates and from different localities and were combined with previously published sequences. Phylogenetic analyses supported the recognition of independent lineages and clusters, indicative of species, but were indecisive in recovering the root of the tree in analyses that included non-gyrocotylid outgroup taxa. The molecular data reveal variation not reflected in morphology and point to a complex picture of genetic divergence shaped by both isolation and migration in the deep-sea environment.
Assuntos
Cestoides/classificação , Cestoides/genética , Peixes/parasitologia , Filogenia , Animais , Oceano Atlântico , DNA de Helmintos/genética , Variação Genética , Especificidade da EspécieRESUMO
Tapeworms (Cestoda) cause neglected diseases that can be fatal and are difficult to treat, owing to inefficient drugs. Here we present an analysis of tapeworm genome sequences using the human-infective species Echinococcus multilocularis, E. granulosus, Taenia solium and the laboratory model Hymenolepis microstoma as examples. The 115- to 141-megabase genomes offer insights into the evolution of parasitism. Synteny is maintained with distantly related blood flukes but we find extreme losses of genes and pathways that are ubiquitous in other animals, including 34 homeobox families and several determinants of stem cell fate. Tapeworms have specialized detoxification pathways, metabolism that is finely tuned to rely on nutrients scavenged from their hosts, and species-specific expansions of non-canonical heat shock proteins and families of known antigens. We identify new potential drug targets, including some on which existing pharmaceuticals may act. The genomes provide a rich resource to underpin the development of urgently needed treatments and control.
Assuntos
Adaptação Fisiológica/genética , Cestoides/genética , Genoma Helmíntico/genética , Parasitos/genética , Animais , Evolução Biológica , Cestoides/efeitos dos fármacos , Cestoides/fisiologia , Infecções por Cestoides/tratamento farmacológico , Infecções por Cestoides/metabolismo , Sequência Conservada/genética , Echinococcus granulosus/genética , Echinococcus multilocularis/efeitos dos fármacos , Echinococcus multilocularis/genética , Echinococcus multilocularis/metabolismo , Genes de Helmintos/genética , Genes Homeobox/genética , Proteínas de Choque Térmico HSP70/genética , Humanos , Hymenolepis/genética , Redes e Vias Metabólicas/genética , Terapia de Alvo Molecular , Parasitos/efeitos dos fármacos , Parasitos/fisiologia , Proteoma/genética , Células-Tronco/citologia , Células-Tronco/metabolismo , Taenia solium/genéticaRESUMO
Neoplasms occur naturally in invertebrates but are not known to develop in tapeworms. We observed nests of monomorphic, undifferentiated cells in samples from lymph-node and lung biopsies in a man infected with the human immunodeficiency virus (HIV). The morphologic features and invasive behavior of the cells were characteristic of cancer, but their small size suggested a nonhuman origin. A polymerase-chain-reaction (PCR) assay targeting eukaryotes identified Hymenolepis nana DNA. Although the cells were unrecognizable as tapeworm tissue, immunohistochemical staining and probe hybridization labeled the cells in situ. Comparative deep sequencing identified H. nana structural genomic variants that are compatible with mutations described in cancer. Invasion of human tissue by abnormal, proliferating, genetically altered tapeworm cells is a novel disease mechanism that links infection and cancer.
Assuntos
Transformação Celular Neoplásica , Himenolepíase/patologia , Hymenolepis nana/genética , Mutação , Adulto , Animais , Análise Mutacional de DNA , DNA de Helmintos/isolamento & purificação , Humanos , Hymenolepis nana/citologia , Masculino , Microscopia Eletrônica de Transmissão , Filogenia , Reação em Cadeia da PolimeraseRESUMO
BACKGROUND: Early developmental patterns of flatworms are extremely diverse and difficult to compare between distant groups. In parasitic flatworms, such as tapeworms, this is confounded by highly derived life cycles involving indirect development, and even the true orientation of the tapeworm antero-posterior (AP) axis has been a matter of controversy. In planarians, and metazoans generally, the AP axis is specified by the canonical Wnt pathway, and we hypothesized that it could also underpin axial formation during larval metamorphosis in tapeworms. RESULTS: By comparative gene expression analysis of Wnt components and conserved AP markers in the tapeworms Echinococcus multilocularis and Hymenolepis microstoma, we found remarkable similarities between the early stages of larval metamorphosis in tapeworms and late embryonic and adult development in planarians. We demonstrate posterior expression of specific Wnt factors during larval metamorphosis and show that scolex formation is preceded by localized expression of Wnt inhibitors. In the highly derived larval form of E. multilocularis, which proliferates asexually within the mammalian host, we found ubiquitous expression of posterior Wnt factors combined with localized expression of Wnt inhibitors that correlates with the asexual budding of scoleces. As in planarians, muscle cells are shown to be a source of secreted Wnt ligands, providing an explanation for the retention of a muscle layer in the immotile E. multilocularis larva. CONCLUSIONS: The strong conservation of gene expression between larval metamorphosis in tapeworms and late embryonic development in planarians suggests, for the first time, a homologous developmental period across this diverse phylum. We postulate these to represent the phylotypic stages of these flatworm groups. Our results support the classical notion that the scolex is the true anterior end of tapeworms. Furthermore, the up-regulation of Wnt inhibitors during the specification of multiple anterior poles suggests a mechanism for the unique asexual reproduction of E. multilocularis larvae.
Assuntos
Echinococcus multilocularis/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Helminto/genética , Hymenolepis/crescimento & desenvolvimento , Proteínas Wnt/genética , Animais , Echinococcus multilocularis/genética , Echinococcus multilocularis/ultraestrutura , Hymenolepis/genética , Hymenolepis/ultraestrutura , Metamorfose BiológicaRESUMO
Australian spiny mountain crayfish (Euastacus, Parastacidae) and their ecotosymbiotic temnocephalan flatworms (Temnocephalida, Platyhelminthes) may have co-occurred and interacted through deep time, during a period of major environmental change. Therefore, reconstructing the history of their association is of evolutionary, ecological, and conservation significance. Here, time-calibrated Bayesian phylogenies of Euastacus species and their temnocephalans (Temnohaswellia and Temnosewellia) indicate near-synchronous diversifications from the Cretaceous. Statistically significant cophylogeny correlations between associated clades suggest linked evolutionary histories. However, there is a stronger signal of codivergence and greater host specificity in Temnosewellia, which co-occurs with Euastacus across its range. Phylogeography and analyses of evolutionary distinctiveness (ED) suggest that regional differences in the impact of climate warming and drying had major effects both on crayfish and associated temnocephalans. In particular, Euastacus and Temnosewellia show strong latitudinal gradients in ED and, conversely, in geographical range size, with the most distinctive, northern lineages facing the greatest risk of extinction. Therefore, environmental change has, in some cases, strengthened ecological and evolutionary associations, leaving host-specific temnocephalans vulnerable to coextinction with endangered hosts. Consequently, the extinction of all Euastacus species currently endangered (75%) predicts coextinction of approximately 60% of the studied temnocephalans, with greatest loss of the most evolutionarily distinctive lineages.
Assuntos
Astacoidea/parasitologia , Evolução Biológica , Turbelários/fisiologia , Animais , Proteínas de Artrópodes/genética , Astacoidea/genética , Austrália , Teorema de Bayes , DNA/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Filogenia , Filogeografia , Análise de Sequência de DNA , Turbelários/genéticaRESUMO
Tapeworms of the order Spathebothriidea Wardle et McLeod, 1952 (Cestoda) are reviewed. Molecular data made it possible to assess, for the first time, the phylogenetic relationships of all genera and to confirm the validity of Bothrimonus Duvernoy, 1842, Diplocotyle Krabbe, 1874 and Didymobothrium Nybelin, 1922. A survey of all species considered to be valid is provided together with new data on egg and scolex morphology and surface ultrastructure (i.e. microtriches). The peculiar morphology of the members of this group, which is today represented by five effectively monotypic genera whose host associations and geographical distribution show little commonality, indicate that it is a relictual group that was once diverse and widespread. The order potentially represents the earliest branch of true tapeworms (i.e. Eucestoda) among extant forms.
Assuntos
Cestoides/genética , Cestoides/fisiologia , Óvulo/classificação , Animais , Cestoides/ultraestrutura , Óvulo/ultraestrutura , Filogenia , Especificidade da EspécieRESUMO
Members of several genera in the digenean superfamily Microphalloidea, namely Renschetrema, Rohdetrema, Cryptotropa, Cephalouterina and Pseudocryptotropa, are characterized by an unusual dorsal position of the genital atrium. In the absence of phylogenetic data, their systematic position has been unstable. In the most recent taxonomic revision of the Microphalloidea, they were housed in three different families. We studied the morphology and obtained DNA sequences of several microphallolideans with a dorsal genital pore, collected from vertebrates in Southeast Asia and South America. We used sequences of the nuclear 28S gene to infer a superfamily-level phylogeny and sequences of the mitochondrial cox1 gene for family-level phylogeny and species-level comparisons. Based on the combination of molecular phylogenetic data and morphological features, we restore the Cryptotropidae and synonymize the Renschetrematidae with the Cryptotropidae. We erect new genera Paracryptotropagen. nov. and Armadoatriumgen. nov., provide amended diagnoses of the Cryptotropidae and Renschetrema and provide keys to the identification of genera within the family. We resurrect the genus Novetrema, previously synonymized with Pseudocryptotropa, and remove Renschetrema indicum from Renschetrema. We describe four new species from lizards in Vietnam and birds in the Philippines and Peru. The unique morphological features and unusually broad host associations and geographical distributions of cryptotropids are discussed.
RESUMO
The 'pygmaeus' microphallids (MPG) are a closely related group of 6 digenean (Platyhelminthes: Trematoda) Microphallus species that share a derived 2-host life cycle in which metacercariae develop inside daughter sporocysts in the intermediate host (intertidal and subtidal gastropods, mostly of the genus Littorina) and are infective to marine birds (ducks, gulls and waders). Here we investigate MPG transmission patterns in coastal ecosystems and their diversification with respect to historical events, host switching and host-parasite co-evolution. Species phylogenies and phylogeographical reconstructions are estimated on the basis of 28S, ITS1 and ITS2 rDNA data and we use a combination of analyses to test the robustness and stability of the results, and the likelihood of alternative biogeographical scenarios. Results demonstrate that speciation within the MPG was not associated with co-speciation with either the first intermediate or final hosts, but rather by host-switching events coincident with glacial cycles in the Northern Hemisphere during the late Pliocene/Pleistocene. These resulted in the expansion of Pacific biota into the Arctic-North Atlantic and periodic isolation of Atlantic and Pacific populations. Thus we hypothesize that contemporary species of MPG and their host associations resulted from fragmentation of populations in regional refugia during stadials, and their subsequent range expansion from refugial centres during interstadials.
Assuntos
Interações Hospedeiro-Parasita/fisiologia , Estágios do Ciclo de Vida , Filogenia , Trematódeos/classificação , Trematódeos/genética , Infecções por Trematódeos/parasitologia , Animais , Organismos Aquáticos/parasitologia , Regiões Árticas , Doenças das Aves/parasitologia , Doenças das Aves/transmissão , Aves , DNA Ribossômico/genética , Dados de Sequência Molecular , Infecções por Trematódeos/transmissãoRESUMO
Wnt genes encode secreted glycoproteins that act in cell-cell signalling to regulate a wide array of developmental processes, ranging from cellular differentiation to axial patterning. Discovery that canonical Wnt/ß-catenin signalling is responsible for regulating head/tail specification in planarian regeneration has recently highlighted their importance in flatworm (phylum Platyhelminthes) development, but examination of their roles in the complex development of the diverse parasitic groups has yet to be conducted. Here, we characterise Wnt genes in the model tapeworm Hymenolepis microstoma and mine genomic resources of free-living and parasitic species for the presence of Wnts and downstream signalling components. We identify orthologs through a combination of BLAST and phylogenetic analyses, showing that flatworms have a highly reduced and dispersed complement that includes orthologs of only five subfamilies (Wnt1, Wnt2, Wnt4, Wnt5 and Wnt11) and fewer paralogs in parasitic flatworms (5-6) than in planarians (9). All major signalling components are identified, including antagonists and receptors, and key binding domains are intact, indicating that the canonical (Wnt/ß-catenin) and non-canonical (planar cell polarity and Wnt/Ca(2+)) pathways are functional. RNA-Seq data show expression of all Hymenolepis Wnts and most downstream components in adults and larvae with the notable exceptions of wnt1, expressed only in adults, and wnt2 expressed only in larvae. The distribution of Wnt subfamilies in animals corroborates the idea that the last common ancestor of the Cnidaria and Bilateria possessed all contemporary Wnts and highlights the extent of gene loss in flatworms.
Assuntos
Hymenolepis/genética , Hymenolepis/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Animais , Evolução Biológica , Padronização Corporal , Expressão Gênica , Genoma , Hymenolepis/embriologia , Filogenia , Via de Sinalização WntRESUMO
Here we examine 3157 foxes from 6 districts of the Slovak Republic in order to determine for the first time the distribution, prevalence and identity of Mesocestodes spp. endemic to this part of central Europe. During the period 2001-2006, an average of 41.9% of foxes were found to harbour Mesocestoides infections. Among the samples we confirmed the widespread and common occurrence of M. litteratus (Batsch, 1786), and report the presence, for the first time, of M. lineatus (Goeze, 1782) in the Slovak Republic, where it has a more restricted geographical range and low prevalence (7%). Using a combination of 12S rDNA, CO1 and ND1 mitochondrial gene sequences together with analysis of 13 morphometric characters, we show that the two species are genetically distinct and can be differentiated by discrete breaks in the ranges of the male and female reproductive characters, but not by the more commonly examined characters of the scolex and strobila. Estimates of interspecific divergence within Mesocestoides ranged from 9 to 18%, whereas intraspecific variation was less than 2%, and phylogenetic analyses of the data showed that despite overlapping geographical ranges, the two commonly reported European species are not closely related, with M. litteratus more closely allied to North American isolates of Mesocestoides than to M. lineatus. We confirm that morphological analysis of reproductive organs can be used to reliably discriminate between these often sympatric species obtained from red foxes.
Assuntos
Infecções por Cestoides/veterinária , Raposas/parasitologia , Mesocestoides/classificação , Animais , Sequência de Bases , Infecções por Cestoides/epidemiologia , Infecções por Cestoides/parasitologia , DNA de Helmintos/química , DNA de Helmintos/genética , DNA Mitocondrial/química , DNA Mitocondrial/genética , Feminino , Genes Mitocondriais/genética , Masculino , Mesocestoides/anatomia & histologia , Mesocestoides/genética , Dados de Sequência Molecular , Fenótipo , Filogenia , Prevalência , RNA Ribossômico/genética , Alinhamento de Sequência , Análise de Sequência de DNA , Eslováquia/epidemiologiaRESUMO
Complete ssrDNA and partial lsrDNA (D1-D3) of 31 species, mainly from the Indo-Pacific region, were sequenced and added to 66 species of the marine cestode order Trypanorhyncha; thus 35% of the 277 known species were sampled. The resulting phylogenetic tree resolved two major clades that represent trypanorhynchs originally parasitizing rajiform (skate and ray) or galeoform hosts. The tree topology supports an earlier classification based on morphology that splits the order into the superfamily Eutetrarhynchoidea together with the Tentacularioidea, and the Gymnorhynchoidea together with the Lacistorhynchoidea and Otobothrioidea. Three of the five recognized superfamilies are monophyletic (Tentacularioidea, Gymnorhynchoidea, Otobothrioidea). Nodal support for the Eutetrarhynchidae and Lacistorhynchidae was poor and resulted in paraphyletic clades. Mapping of morphological characters showed the tentacular armature of the scolex to be highly variable within clades, demonstrating that armature patterns used traditionally in classification, are homoplasious. Similarly, the tetrabothriate scolex, currently utilized as a family-distinguishing character in traditional classifications, has developed independently in multiple groups. Synapomorphies for the higher taxa are detailed. Sequence data from duplicate taxa confirmed interoceanic distribution patterns and low intraspecific genetic divergence and host specificity for nine trypanorhynch species belonging to five families and four superfamilies. Four distinct lineages of trypanorhynchs can be recognized with one mainly infecting rajiform hosts whereas the others infecting both, sharks and rays. Eutetrarhynchoids and tentacularioids have secondarily invaded shark hosts whilst utilising the marine food web involving teleost fishes as intermediate hosts. Three cases of host switching from sharks to rays can be inferred within the lacistorhynchoids, in one case caused by a switch from perciform to gadiform intermediate hosts. This likely enabled a radiation into the deep sea environment. Implications of the molecular phylogeny for the classification and evolutionary developments within the order are discussed.
Assuntos
Evolução Molecular , Especiação Genética , Filogenia , Platelmintos/genética , Animais , Teorema de Bayes , DNA de Helmintos/genética , DNA Ribossômico/genética , Interações Hospedeiro-Parasita/genética , Funções Verossimilhança , Modelos Genéticos , Platelmintos/classificação , Alinhamento de Sequência , Análise de Sequência de DNA , Tubarões/parasitologia , Rajidae/parasitologiaRESUMO
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.
Assuntos
Perfilação da Expressão Gênica , Hymenolepis/genética , MicroRNAs/análise , MicroRNAs/genética , Análise de Sequência de RNA , AnimaisRESUMO
BACKGROUND: Tapeworms are agents of neglected tropical diseases responsible for significant health problems and economic loss. They also exhibit adaptations to a parasitic lifestyle that confound comparisons of their development with other animals. Identifying the genetic factors regulating their complex ontogeny is essential to understanding unique aspects of their biology and for advancing novel therapeutics. Here we use RNA sequencing to identify up-regulated signalling components, transcription factors and post-transcriptional/translational regulators (genes of interest, GOI) in the transcriptomes of Larvae and different regions of segmented worms in the tapeworm Hymenolepis microstoma and combine this with spatial gene expression analyses of a selection of genes. RESULTS: RNA-seq reads collectively mapped to 90% of the > 12,000 gene models in the H. microstoma v.2 genome assembly, demonstrating that the transcriptome profiles captured a high percentage of predicted genes. Contrasts made between the transcriptomes of Larvae and whole, adult worms, and between the Scolex-Neck, mature strobila and gravid strobila, resulted in 4.5-30% of the genes determined to be differentially expressed. Among these, we identified 190 unique GOI up-regulated in one or more contrasts, including a large range of zinc finger, homeobox and other transcription factors, components of Wnt, Notch, Hedgehog and TGF-ß/BMP signalling, and post-transcriptional regulators (e.g. Boule, Pumilio). Heatmap clusterings based on overall expression and on select groups of genes representing 'signals' and 'switches' showed that expression in the Scolex-Neck region is more similar to that of Larvae than to the mature or gravid regions of the adult worm, which was further reflected in large overlap of up-regulated GOI. CONCLUSIONS: Spatial expression analyses in Larvae and adult worms corroborated inferences made from quantitative RNA-seq data and in most cases indicated consistency with canonical roles of the genes in other animals, including free-living flatworms. Recapitulation of developmental factors up-regulated during larval metamorphosis suggests that strobilar growth involves many of the same underlying gene regulatory networks despite the significant disparity in developmental outcomes. The majority of genes identified were investigated in tapeworms for the first time, setting the stage for advancing our understanding of developmental genetics in an important group of flatworm parasites.
RESUMO
Dicrocoelium dendriticum (Rudolphi, 1819) and Dicrocoelium hospes (Looss, 1907) are recognised to affect the liver of domestic and wild ruminants. A third species, Dicrocoelium orientalis which was described from musk deer in the Baikal region of the former Soviet Union and re-named to Dicrocoelium chinensis (Sudarikov and Ryjikov, 1951) Tang and Tang, 1978 was isolated from other species of deer in Asian countries and from mouflon and roe deer in Europe. Scant information is available for D. chinensis, including the range of species that act as definitive and intermediate hosts. To provide morphological and molecular evidences differentiating D. chinensis versus D. dendriticum, 239 Dicrocoelium spp. specimens were collected from sheep, cattle and sika deer from different localities in Austria, Germany and Italy. Specimens were morphologically identified based on the testes orientation, overall size, and level of maximum body width and other morphometric measurements. From this sample, 10 specimens of D. chinensis and 25 of D. dendriticum from different hosts and geographical localities were characterized molecularly through sequencing of partial 18S rDNA (approximately 1400 bp) and ITS-2 (including the 5.8S and 28S flanking regions; approximately 600 bp). Interspecific differences between D. dendriticum and D. chinensis of 0.14% and 3.8% were recorded in 18S rRNA and ITS-2 sequences, respectively. Phylogenetic analyses via Bayesian inference were conducted using sequences of ITS-2 (276 bp) and partial 28S (221 bp) of the above species of Dicrocoelium together with 20 species belonging to the Xiphidiata within the Plagiorchiida available in GenBank. Both gene regions were strongly concordant in differentiating the Dicrocoeliidae, Gorgoderidae and Plagiorchiidae and were in agreement with their current classification. Morphological and molecular characterization clearly differentiate D. dendriticum and D. chinensis as two distinct digeneans infecting ruminants. The implications on the separate status of D. chinensis on the etiology, biology and diagnosis of dicrocoeliosis are discussed.
Assuntos
DNA de Helmintos/genética , Dicrocoelium/genética , Genoma Helmíntico , Animais , Áustria , Bovinos , DNA de Helmintos/química , Cervos , Dicrocoelium/classificação , Dicrocoelium/citologia , Alemanha , Itália , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico/genética , RNA Ribossômico 18S/genética , RNA Ribossômico 28S/genética , Análise de Sequência de DNA , Ovinos , Especificidade da EspécieRESUMO
The success of any population translocation programme relies heavily on the measures implemented to control and monitor the spread of disease. Without these measures, programmes run the risk of releasing immunologically naïve species or, more dangerously, introducing novel infectious agents to native populations. As a precaution, a reintroduction programme for the common or hazel dormouse, Muscardinus avellanarius, in England screens dormice before release following captive breeding. Using PCR sequencing of a range of genes, we tested whether the same species of tapeworm(s) were present in captive and free-living dormice. Whilst only Rodentolepis straminea were identified in free-living dormice, cestode ova found in a captive individual produced a molecular match closely related to Hymenolepis microstoma and a previously unrecorded Rodentolepis species. To prevent putting at risk the free-living population, we recommended the continued treatment of dormice showing tapeworm infection before release. Our work demonstrates how molecular techniques can be used to inform reintroduction programmes, reduce risk from disease and increase chances of reintroduction success.