The eggs of the blood fluke Schistosoma mansoni are the main cause of the clinical manifestations of chronic schistosomiasis. After laying, the egg "winners" attach to the endothelium of the mesenteric vein and, after a period of development, induce the growth of a small granuloma, which facilitates their passage to the intestinal lumen. Egg "losers" carried by the bloodstream to non-specific tissues also undergo full development and induce large granuloma formation, but their life ends there. Although these trapped eggs represent a dead end in the parasite life cycle, the vast majority of studies attempting to describe the biology of the S. mansoni eggs have studied these liver-trapped "losers" instead of migrating intestinal "winners". This raises the fundamental question of how these eggs differ. With robust comparative transcriptomic analysis performed on S. mansoni eggs isolated 7 weeks post infection, we show that gene expression is critically dependent on tissue localization, both in the early and late stages of development. While mitochondrial genes and venom allergen-like proteins are significantly upregulated in mature intestinal eggs, well-described egg immunomodulators IPSE/alpha-1 and omega-1, together with micro-exon genes, are predominantly expressed in liver eggs. In addition, several proteases and protease inhibitors previously implicated in egg-host interactions display clear tissue-specific gene expression patterns. These major differences in gene expression could be then reflected in the observed different ability of liver and intestinal soluble egg antigens to elicit host immune responses and in the shorter viability of miracidia hatched from liver eggs. Our comparative analysis provides a new perspective on the biology of parasite's eggs in the context of their development and tissue localization. These findings could contribute to a broader and more accurate understanding of parasite eggs interactions with the host, which have historically been often restricted to liver eggs and sometimes inaccurately generalized.

• MeSH
• antigeny helmintové imunologie   MeSH
• játra * parazitologie   imunologie   metabolismus   MeSH
• myši MeSH
• ovum metabolismus   imunologie   MeSH
• proteiny červů genetika   metabolismus   imunologie   MeSH
• Schistosoma mansoni * imunologie   genetika   MeSH
• schistosomiasis mansoni * imunologie   parazitologie   MeSH
• střeva parazitologie   imunologie   MeSH
• vaječné proteiny MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antigeny helmintové MeSH
• IPSE protein, Schistosoma mansoni MeSH Prohlížeč
• proteiny červů MeSH
• vaječné proteiny MeSH

Dermanyssus gallinae is a blood-feeding mite that parasitises wild birds and farmed poultry. Its remarkably swift processing of blood, together with the capacity to blood-feed during most developmental stages, makes this mite a highly debilitating pest. To identify specific adaptations to digestion of a haemoglobin-rich diet, we constructed and compared transcriptomes from starved and blood-fed stages of the parasite and identified midgut-enriched transcripts. We noted that midgut transcripts encoding cysteine proteases were upregulated with a blood meal. Mapping the full proteolytic apparatus, we noted a reduction in the suite of cysteine proteases, missing homologues for Cathepsin B and C. We have further identified and phylogenetically analysed three distinct transcripts encoding vitellogenins that facilitate the reproductive capacity of the mites. We also fully mapped transcripts for haem biosynthesis and the ferritin-based system of iron storage and inter-tissue trafficking. Additionally, we identified transcripts encoding proteins implicated in immune signalling (Toll and IMD pathways) and activity (defensins and thioester-containing proteins), RNAi, and ion channelling (with targets for commercial acaricides such as Fluralaner, Fipronil, and Ivermectin). Viral sequences were filtered from the Illumina reads and we described, in part, the RNA-virome of D. gallinae with identification of a novel virus, Red mite quaranjavirus 1.

• MeSH
• drůbež MeSH
• infestace roztoči * veterinární   parazitologie   MeSH
• kur domácí MeSH
• nemoci drůbeže * MeSH
• roztoči * genetika   MeSH
• sekvenování transkriptomu MeSH
• virom MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH

Fasciola hepatica is a global parasite of livestock which also causes a neglected zoonosis in humans. The parasite's communication with the host during its complicated lifecycle is based on an ingenious enzymatic apparatus which includes a variety of peptidases. These enzymes are implicated in parasite migration, pathogenesis of the disease, and modification of host immune response. Although the dynamics of proteolytic machinery produced by intra-mammalian F. hepatica life stages has been previously investigated in great detail, peptidases of the eggs so far received little scientific attention. In this study, we performed a comparative RNA-seq analysis aimed at identification of peptidases expressed in F. hepatica eggs, cultured at 37 °C to represent gall bladder retained eggs, for different time periods and employed mass spectrometry in order to identify and quantify peptidases translated in F. hepatica egg lysates. We demonstrated that F. hepatica eggs undergo significant molecular changes when cultured at the physiological temperature of the definitive host. Egg transcriptome is subject to numerous subtle changes while their proteome is even more variable. The peptidase profile is considerably modified on both transcriptome and proteome level. Finally, we measured and classified proteolytic activities in extracts from F. hepatica eggs using a library of fluorogenic substrates and peptidase class-selective inhibitors. Activities of threonine peptidases were detected constantly, while the cysteine peptidases prevailing in freshly laid eggs are substituted by aspartic peptidase and metallopeptidase activities in the later stages of egg development.

• MeSH
• Fasciola hepatica * enzymologie   MeSH
• ovum * enzymologie   MeSH
• proteasy * metabolismus   MeSH
• proteom * MeSH
• proteomika MeSH
• savci parazitologie   MeSH
• tělesná teplota MeSH
• transkriptom * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• proteasy * MeSH
• proteom * MeSH

Antibody trapping is a recently described strategy for immune evasion observed in the intestinal trematode Echinostoma caproni, which may aid to avoiding the host humoral response, thus facilitating parasite survival in the presence of high levels of local-specific antibodies. Parasite-derived peptidases carry out the degradation of trapped antibodies, being essential for this mechanism. Herein, we show that cathepsin-like cysteine endopeptidases are active in the excretory/secretory products (ESPs) of E. caproni and play an important role in the context of antibody trapping. Cysteine endopeptidase activity was detected in the ESPs of E. caproni adults. The affinity probe DCG-04 distinguished a cysteine peptidase band in ESPs, which was specifically recognized by an anti-cathepsin L heterologous antibody. The same antibody localized this protein in the gut and syncytial tegument of adult worms. Studies with cultured parasites showed that in vivo-bound antibodies are removed from the parasite surface in the absence of peptidase inhibitors, while addition of cathepsin L inhibitor prevented their degradation. These results indicate that cathepsin L-like peptidases are involved in the degradation of surface-trapped antibodies and suggest that cysteine peptidases are not only crucial for tissue-invading trematodes, but they can be equally relevant at the parasite-host interface in gut-dwelling flukes.

• Klíčová slova
• Antibody, Cathepsin, Cysteine peptidase, Echinostoma caproni, Immune evasion, Trematode,
• MeSH
• cysteinové endopeptidasy metabolismus   MeSH
• Echinostoma imunologie   metabolismus   MeSH
• echinostomióza imunologie   parazitologie   MeSH
• imunitní únik imunologie   MeSH
• kathepsin L antagonisté a inhibitory   MeSH
• proteolýza MeSH
• protilátky protozoální imunologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cysteinové endopeptidasy MeSH
• kathepsin L MeSH
• protilátky protozoální MeSH

Trichobilharzia species are parasitic flatworms (called schistosomes or flukes) that cause important diseases in birds and humans, but very little is known about their molecular biology. Here, using a transcriptomics-bioinformatics-based approach, we explored molecular aspects pertaining to the nutritional requirements of Trichobilharzia szidati ('visceral fluke') and T. regenti ('neurotropic fluke') in their avian host. We studied the larvae of each species before they enter (cercariae) and as they migrate (schistosomules) through distinct tissues in their avian (duck) host. Cercariae of both species were enriched for pathways or molecules associated predominantly with carbohydrate metabolism, oxidative phosphorylation and translation of proteins linked to ribosome biogenesis, exosome production and/or lipid biogenesis. Schistosomules of both species were enriched for pathways or molecules associated with processes including signal transduction, cell turnover and motility, DNA replication and repair, molecular transport and/or catabolism. Comparative informatic analyses identified molecular repertoires (within, e.g., peptidases and secretory proteins) in schistosomules that can broadly degrade macromolecules in both T. szidati and T. regenti, and others that are tailored to each species to selectively acquire nutrients from particular tissues through which it migrates. Thus, this study provides molecular evidence for distinct modes of nutrient acquisition between the visceral and neurotropic flukes of birds.

• MeSH
• cerkárie klasifikace   genetika   patogenita   MeSH
• DNA helmintů klasifikace   genetika   MeSH
• fylogeneze * MeSH
• kachny genetika   parazitologie   MeSH
• lidé MeSH
• nemoci ptáků genetika   parazitologie   MeSH
• ptáci genetika   parazitologie   MeSH
• Schistosomatidae genetika   patogenita   MeSH
• schistosomóza genetika   parazitologie   MeSH
• Trematoda klasifikace   genetika   patogenita   MeSH
• výpočetní biologie MeSH
• živiny MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA helmintů MeSH

BACKGROUND: Serpins are a superfamily of serine peptidase inhibitors that participate in the regulation of many physiological and cell peptidase-mediated processes in all organisms (e.g. in blood clotting, complement activation, fibrinolysis, inflammation, and programmed cell death). It was postulated that in the blood-feeding members of the monogenean family Diplozoidae, serpins could play an important role in the prevention of thrombus formation, activation of complement, inflammation in the host, and/or in the endogenous regulation of protein degradation. RESULTS: In silico analysis showed that the DNA and primary protein structures of serpin from Eudiplozoon nipponicum (EnSerp1) are similar to other members of the serpin superfamily. The inhibitory potential of EnSerp1 on four physiologically-relevant serine peptidases (trypsin, factor Xa, kallikrein, and plasmin) was demonstrated and its presence in the worm's excretory-secretory products (ESPs) was confirmed. CONCLUSION: EnSerp1 influences the activity of peptidases that play a role in blood coagulation, fibrinolysis, and complement activation. This inhibitory potential, together with the serpin's presence in ESPs, suggests that it is likely involved in host-parasite interactions and could be one of the molecules involved in the control of feeding and prevention of inflammatory responses.

Contexte : Les serpines sont une super-famille d’inhibiteurs de sérine peptidases qui participent, dans tous les organismes, à la régulation de nombreux processus physiologiques et à médiation par les peptidases cellulaires (par exemple la coagulation sanguine, l’activation du complément, la fibrinolyse, l’inflammation et la mort cellulaire programmée). Il a été postulé que chez les Monogènes de la famille Diplozoidae, qui sont hématophages, les serpines pourraient jouer un rôle important dans la prévention de la formation de thrombus, l’activation du complément, l’inflammation chez l’hôte et/ou la régulation endogène de la dégradation des protéines. Résultats : Une analyse in silico a montré que l’ADN et les structures primaires protéiques de la serpine d’Eudiplozoon nipponicum (EnSerp1) sont similaires aux autres membres de la superfamille des serpines. Le potentiel inhibiteur d’EnSerp1 sur quatre sérine peptidases physiologiquement pertinentes (la trypsine, le facteur Xa, la kallikréine et la plasmine) a été démontré et sa présence dans les produits excréteurs de sécrétion du ver (ESP) a été confirmée. Conclusion : EnSerp1 influence l’activité des peptidases qui jouent un rôle dans la coagulation sanguine, la fibrinolyse et l’activation du complément. Ce potentiel inhibiteur, ainsi que la présence de la serpine dans les ESP, suggèrent qu’elle est probablement impliquée dans les interactions hôte-parasite et pourrait être l’une des molécules impliquées dans le contrôle de l’alimentation et la prévention des réponses inflammatoires.

• MeSH
• DNA helmintů chemie   MeSH
• fylogeneze MeSH
• infekce červy třídy Trematoda parazitologie   veterinární   MeSH
• inhibitory serinových proteinas chemie   genetika   izolace a purifikace   metabolismus   MeSH
• kapři parazitologie   MeSH
• nemoci ryb parazitologie   MeSH
• počítačová simulace MeSH
• polymerázová řetězová reakce MeSH
• rekombinantní proteiny genetika   izolace a purifikace   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční seřazení MeSH
• serpiny chemie   genetika   izolace a purifikace   metabolismus   MeSH
• Trematoda chemie   klasifikace   enzymologie   genetika   MeSH
• žábry parazitologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA helmintů MeSH
• inhibitory serinových proteinas MeSH
• rekombinantní proteiny MeSH
• serpiny MeSH

BACKGROUND: Cysteine peptidases of clan CA, family C1 account for a major part of proteolytic activity in the haematophagous monogenean Eudiplozoon nipponicum. The full spectrum of cysteine cathepsins is, however, unknown and their particular biochemical properties, tissue localisation, and involvement in parasite-host relationships are yet to be explored. METHODS: Sequences of cathepsins L and B (EnCL and EnCB) were mined from E. nipponicum transcriptome and analysed bioinformatically. Genes encoding two EnCLs and one EnCB were cloned and recombinant proteins produced in vitro. The enzymes were purified by chromatography and their activity towards selected substrates was characterised. Antibodies and specific RNA probes were employed for localisation of the enzymes/transcripts in tissues of E. nipponicum adults. RESULTS: Transcriptomic analysis revealed a set of ten distinct transcripts that encode EnCLs. The enzymes are significantly variable in their active sites, specifically the S2 subsites responsible for interaction with substrates. Some of them display unusual structural features that resemble cathepsins B and S. Two recombinant EnCLs had different pH activity profiles against both synthetic and macromolecular substrates, and were able to hydrolyse blood proteins and collagen I. They were localised in the haematin cells of the worm's digestive tract and in gut lumen. The EnCB showed similarity with cathepsin B2 of Schistosoma mansoni. It displays molecular features typical of cathepsins B, including an occluding loop responsible for its exopeptidase activity. Although the EnCB hydrolysed haemoglobin in vitro, it was localised in the vitelline cells of the parasite and not the digestive tract. CONCLUSIONS: To our knowledge, this study represents the first complex bioinformatic and biochemical characterisation of cysteine peptidases in a monogenean. Eudiplozoon nipponicum adults express a variety of CLs, which are the most abundant peptidases in the worms. The properties and localisation of the two heterologously expressed EnCLs indicate a central role in the (partially extracellular?) digestion of host blood proteins. High variability of substrate-binding sites in the set of EnCLs suggests specific adaptation to a range of biological processes that require proteolysis. Surprisingly, a single cathepsin B is expressed by the parasite and it is not involved in digestion, but probably in vitellogenesis.

• Klíčová slova
• Blood digestion, Cathepsin, Cysteine peptidase, Diplozoidae, Eudiplozoon nipponicum, Fish parasite, Haematophagy, Monogenea, Protease, S2 subsite,
• MeSH
• gastrointestinální trakt parazitologie   MeSH
• hydrolýza MeSH
• interakce hostitele a parazita MeSH
• kapři parazitologie   MeSH
• kathepsin B chemie   genetika   izolace a purifikace   metabolismus   MeSH
• kathepsin L chemie   genetika   izolace a purifikace   metabolismus   MeSH
• proteolýza MeSH
• rekombinantní proteiny analýza   genetika   izolace a purifikace   MeSH
• stanovení celkové genové exprese MeSH
• Trematoda enzymologie   genetika   MeSH
• zavlečené druhy MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kathepsin B MeSH
• kathepsin L MeSH
• rekombinantní proteiny MeSH

Schistosomiasis is one of a number of chronic helminth diseases of poverty that severely impact personal and societal well-being and productivity. Peptidases (proteases) are vital to successful parasitism, and can modulate host physiology and immunology. Interference of peptidase action by specific drugs or vaccines can be therapeutically beneficial. To date, research on peptidases in the schistosome parasite has focused on either the functional characterization of individual peptidases or their annotation as part of global genome or transcriptome studies. We were interested in functionally characterizing the complexity of peptidase activity operating at the host-parasite interface, therefore the excretory-secretory products of key developmental stages of Schistosoma mansoni that parasitize the human were examined. Using class specific peptidase inhibitors in combination with a multiplex substrate profiling assay, a number of unique activities derived from endo- and exo-peptidases were revealed in the excretory-secretory products of schistosomula (larval migratory worms), adults and eggs. The data highlight the complexity of the functional degradome for each developmental stage of this parasite and facilitate further enquiry to establish peptidase identity, physiological and immunological function, and utility as drug or vaccine candidates.

• Klíčová slova
• Excretion, Fluke, Inhibitor, Parasite, Protease, Secretion,
• MeSH
• interakce hostitele a patogenu MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• ovum metabolismus   MeSH
• proteasy metabolismus   MeSH
• proteiny červů metabolismus   MeSH
• proteolýza MeSH
• Schistosoma mansoni růst a vývoj   metabolismus   fyziologie   MeSH
• schistosomiasis mansoni parazitologie   MeSH
• sekvence aminokyselin MeSH
• stadia vývoje MeSH
• substrátová specifita MeSH
• vazebná místa MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• proteasy MeSH
• proteiny červů MeSH

To date, most molecular investigations of schistosomatids have focused principally on blood flukes (schistosomes) of humans. Despite the clinical importance of cercarial dermatitis in humans caused by Trichobilharzia regenti and the serious neuropathologic disease that this parasite causes in its permissive avian hosts and accidental mammalian hosts, almost nothing is known about the molecular aspects of how this fluke invades its hosts, migrates in host tissues and how it interacts with its hosts' immune system. Here, we explored selected aspects using a transcriptomic-bioinformatic approach. To do this, we sequenced, assembled and annotated the transcriptome representing two consecutive life stages (cercariae and schistosomula) of T. regenti involved in the first phases of infection of the avian host. We identified key biological and metabolic pathways specific to each of these two developmental stages and also undertook comparative analyses using data available for taxonomically related blood flukes of the genus Schistosoma. Detailed comparative analyses revealed the unique involvement of carbohydrate metabolism, translation and amino acid metabolism, and calcium in T. regenti cercariae during their invasion and in growth and development, as well as the roles of cell adhesion molecules, microaerobic metabolism (citrate cycle and oxidative phosphorylation), peptidases (cathepsins) and other histolytic and lysozomal proteins in schistosomula during their particular migration in neural tissues of the avian host. In conclusion, the present transcriptomic exploration provides new and significant insights into the molecular biology of T. regenti, which should underpin future genomic and proteomic investigations of T. regenti and, importantly, provides a useful starting point for a range of comparative studies of schistosomatids and other trematodes.

• MeSH
• biologická adaptace * MeSH
• interakce hostitele a patogenu * MeSH
• kachny parazitologie   MeSH
• metabolické sítě a dráhy genetika   MeSH
• molekulární sekvence - údaje MeSH
• Schistosomatidae genetika   růst a vývoj   MeSH
• sekvenční analýza DNA MeSH
• stadia vývoje MeSH
• stanovení celkové genové exprese * MeSH
• výpočetní biologie * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Blood flukes (Schistosoma spp.) are parasites that can survive for years or decades in the vasculature of permissive mammalian hosts, including humans. Proteolytic enzymes (proteases) are crucial for successful parasitism, including aspects of invasion, maturation and reproduction. Most attention has focused on the 'cercarial elastase' serine proteases that facilitate skin invasion by infective schistosome larvae, and the cysteine and aspartic proteases that worms use to digest the blood meal. Apart from the cercarial elastases, information regarding other S. mansoni serine proteases (SmSPs) is limited. To address this, we investigated SmSPs using genomic, transcriptomic, phylogenetic and functional proteomic approaches. METHODOLOGY/PRINCIPAL FINDINGS: Genes encoding five distinct SmSPs, termed SmSP1 - SmSP5, some of which comprise disparate protein domains, were retrieved from the S. mansoni genome database and annotated. Reverse transcription quantitative PCR (RT- qPCR) in various schistosome developmental stages indicated complex expression patterns for SmSPs, including their constituent protein domains. SmSP2 stood apart as being massively expressed in schistosomula and adult stages. Phylogenetic analysis segregated SmSPs into diverse clusters of family S1 proteases. SmSP1 to SmSP4 are trypsin-like proteases, whereas SmSP5 is chymotrypsin-like. In agreement, trypsin-like activities were shown to predominate in eggs, schistosomula and adults using peptidyl fluorogenic substrates. SmSP5 is particularly novel in the phylogenetics of family S1 schistosome proteases, as it is part of a cluster of sequences that fill a gap between the highly divergent cercarial elastases and other family S1 proteases. CONCLUSIONS/SIGNIFICANCE: Our series of post-genomics analyses clarifies the complexity of schistosome family S1 serine proteases and highlights their interrelationships, including the cercarial elastases and, not least, the identification of a 'missing-link' protease cluster, represented by SmSP5. A framework is now in place to guide the characterization of individual proteases, their stage-specific expression and their contributions to parasitism, in particular, their possible modulation of host physiology.

• MeSH
• biochemie MeSH
• fylogeneze MeSH
• genomika MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• molekulární sekvence - údaje MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• proteomika MeSH
• Schistosoma mansoni enzymologie   genetika   MeSH
• sekvenční analýza DNA MeSH
• sekvenční homologie MeSH
• serinové proteasy genetika   metabolismus   MeSH
• shluková analýza MeSH
• transkriptom MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• serinové proteasy MeSH