BACKGROUND: Parasitism as a life strategy has independently evolved multiple times within the eukaryotic tree of life. Each lineage has developed mechanisms to invade hosts, exploit resources, and ensure replication, but our knowledge of survival mechanisms in many parasitic taxa remain extremely limited. One such group is the Myxozoa, which are obligate, dixenous cnidarians. Evidence suggests that myxozoans evolved from free-living ancestors to endoparasites around 600 million years ago and are likely one of the first metazoan parasites on Earth. Some myxozoans pose significant threats to farmed and wild fish populations, negatively impacting aquaculture and fish stocks; one such example is Sphaerospora molnari, which forms spores in the gills of common carp (Cyprinus carpio), disrupting gill epithelia and causing somatic and respiratory failure. Sphaerospora molnari undergoes sequential development in different organs of its host, with large numbers of morphologically distinct stages occurring in the blood, liver, and gills of carp. We hypothesize that these parasite life-stages differ in regards to their host exploitation, pathogenicity, and host immune evasion strategies and mechanisms. We performed stage-specific transcriptomic profiling to identify differentially expressed key functional gene groups that relate to these functions and provide a fundamental understanding of the mechanisms S. molnari uses to optimize its parasitic lifestyle. We aimed to identify genes that are likely related to parasite pathogenicity and host cell exploitation mechanisms, and we hypothesize that genes unique to S. molnari might be indicative of evolutionary innovations and specific adaptations to host environments. RESULTS: We used parasite isolation protocols and comparative transcriptomics to study early proliferative and spore-forming stages of S. molnari, unveiling variation in gene expression between each stage. We discovered several apparent innovations in the S. molnari transcriptome, including proteins that are likely to function in the uptake of previously unknown key nutrients, immune evasion factors that may contribute to long-term survival in hosts, and proteins that likely improve adhesion to host cells that may have arisen from horizontal gene transfer. Notably, we identified genes that are similar to known virulence factors in other parasitic organisms, particularly blood and intestinal parasites like Plasmodium, Trypanosoma, and Giardia. Many of these genes are absent in published cnidarian and myxozoan datasets and appear to be specific to S. molnari; they may therefore represent potential innovations enabling Sphaerospora to exploit the host's blood system. CONCLUSIONS: In order to address the threat posed by myxozoans to both cultured fish species and wild stocks, it is imperative to deepen our understanding of their genetics. Sphaerospora molnari offers an appealing model for stage-specific transcriptomic profiling and for identifying differentially expressed key functional gene groups related to parasite development. We identified genes that are thus far unique to S. molnari, which reveal their evolutionary novelty and likely role as adaptations to specific host niches. In addition, we describe the pathogenicity-associated genetic toolbox of S. molnari and discuss the implications of our discoveries for disease control by shedding light on specific targets for potential intervention strategies.

• Klíčová slova
• Sphaerospora molnari, Differential expression, Myxozoans, Pathogenicity related, Species specific genes,
• MeSH
• fyziologická adaptace genetika   MeSH
• interakce hostitele a parazita genetika   MeSH
• kapři parazitologie   MeSH
• Myxozoa * genetika   patogenita   fyziologie   MeSH
• nemoci ryb parazitologie   MeSH
• stadia vývoje genetika   MeSH
• stanovení celkové genové exprese MeSH
• transkriptom * MeSH
• žábry parazitologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

BACKGROUND: Helminth extracellular vesicles (EVs) are known to have a three-way communication function among parasitic helminths, their host and the host-associated microbiota. They are considered biological containers that may carry virulence factors, being therefore appealing as therapeutic and prophylactic target candidates. This study aims to describe and characterise EVs secreted by Sparicotyle chrysophrii (Polyopisthocotyla: Microcotylidae), a blood-feeding gill parasite of gilthead seabream (Sparus aurata), causing significant economic losses in Mediterranean aquaculture. METHODS: To identify proteins involved in extracellular vesicle biogenesis, genomic datasets from S. chrysophrii were mined in silico using known protein sequences from Clonorchis spp., Echinococcus spp., Fasciola spp., Fasciolopsis spp., Opisthorchis spp., Paragonimus spp. and Schistosoma spp. The location and ultrastructure of EVs were visualised by transmission electron microscopy after fixing adult S. chrysophrii specimens by high-pressure freezing and freeze substitution. EVs were isolated and purified from adult S. chrysophrii (n = 200) using a newly developed ultracentrifugation-size-exclusion chromatography protocol for Polyopisthocotyla, and EVs were characterised via nanoparticle tracking analysis and tandem mass spectrometry. RESULTS: Fifty-nine proteins involved in EV biogenesis were identified in S. chrysophrii, and EVs compatible with ectosomes were observed in the syncytial layer of the haptoral region lining the clamps. The isolated and purified nanoparticles had a mean size of 251.8 nm and yielded 1.71 × 108 particles · mL-1. The protein composition analysis identified proteins related to peptide hydrolases, GTPases, EF-hand domain proteins, aerobic energy metabolism, anticoagulant/lipid-binding, haem detoxification, iron transport, EV biogenesis-related, vesicle-trafficking and other cytoskeletal-related proteins. Several identified proteins, such as leucyl and alanyl aminopeptidases, calpain, ferritin, dynein light chain, 14-3-3, heat shock protein 70, annexin, tubulin, glutathione S-transferase, superoxide dismutase, enolase and fructose-bisphosphate aldolase, have already been proposed as target candidates for therapeutic or prophylactic purposes. CONCLUSIONS: We have unambiguously demonstrated for the first time to our knowledge the secretion of EVs by an ectoparasitic flatworm, inferring their biogenesis machinery at a genomic and transcriptomic level, and by identifying their location and protein composition. The identification of multiple therapeutic targets among EVs' protein repertoire provides opportunities for target-based drug discovery and vaccine development for the first time in Polyopisthocotyla (sensu Monogenea), and in a fish-ectoparasite model.

• Klíčová slova
• Drug target candidates, Ectosomes, Electron microscopy, Exosomes, Monogenea, Peptidases, Polyopisthocotyla, Prophylactic target candidates,
• MeSH
• extracelulární vezikuly * MeSH
• mořan zlatý * parazitologie   MeSH
• ploštěnci * MeSH
• proteomika MeSH
• Trematoda * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Hybrid generations usually face either a heterosis advantage or a breakdown, that can be expressed by the level of parasite infection in hybrid hosts. Hybrids are less infected by parasites than parental species (especially F1 generations) or more infected than parental species (especially post-F1 generations). We performed the experiment with blood-feeding gill parasite Paradiplozoon homoion (Monogenea) infecting leuciscid species, Abramis brama and Rutilus rutilus, their F1 generation and two backcross generations. Backcross generations tended to be more parasitized than parental lines and the F1 generation. The number of differentially expressed genes (DEGs) was lower in F1 hybrids and higher in backcross hybrids when compared to each of the parental lines. The main groups of DEGs were shared among lines; however, A. brama and R. rutilus differed in some of the top gene ontology (GO) terms. DEG analyses revealed the role of heme binding and erythrocyte differentiation after infection by blood-feeding P. homoion. Two backcross generations shared some of the top GO terms, representing mostly downregulated genes associated with P. homoion infection. KEGG analysis revealed the importance of disease-associated pathways; the majority of them were shared by two backcross generations. Our study revealed the most pronounced DEGs associated with blood-feeding monogeneans in backcross hybrids, potentially (but not exclusively) explainable by hybrid breakdown. The lower DEGs reported in F1 hybrids being less parasitized than backcross hybrids is in line with the hybrid advantage.

• Klíčová slova
• Monogenea, Paradiplozoon homoion, RNA seq, differential gene expression, freshwater fish, hybrid breakdown, hybrid heterosis, hybridization,
• Publikační typ
• časopisecké články MeSH

Host blood protein digestion plays a pivotal role in the ontogeny and reproduction of hematophagous vectors. The gut of hematophagous arthropods stores and slowly digests host blood and represents the primary gateway for transmitted pathogens. The initial step in blood degradation is induced lysis of host red blood cells (hemolysis), which releases hemoglobin for subsequent processing by digestive proteolytic enzymes. The activity cycles and characteristics of hemolysis in vectors are poorly understood. Hence, we investigated hemolysis in two evolutionarily distant blood-feeding arthropods: The mosquito Culex pipiens and the soft tick Argas persicus, both of which are important human and veterinary disease vectors. Hemolysis in both species was cyclical after blood meal ingestion. Maximum digestion occurs under slightly alkaline conditions in females. Hemolytic activity appears to be of lipoid origin in C. pipiens and enzymatic activity (proteolytic) in A. persicus. We have assessed the effect of pH, incubation time, and temperature on hemolytic activity and the hemolysin. The susceptibility of red blood cells from different hosts to the hemolysin and the effect of metabolic inhibition of hemolytic activity were assessed. We conclude that in C. pipiens and A. persicus midgut hemolysins control the amplitude of blood lysis step to guarantee an efficient blood digestion.

• MeSH
• členovci - vektory fyziologie   MeSH
• členovci MeSH
• Culex MeSH
• Culicidae MeSH
• erytrocyty MeSH
• hematologické testy MeSH
• hemolýza * MeSH
• hemolyziny MeSH
• komáří přenašeči fyziologie   MeSH
• lidé MeSH
• stravovací zvyklosti fyziologie   MeSH
• trávicí systém MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hemolyziny MeSH

The human parasite Trypanosoma brucei does not synthesize heme de novo and instead relies entirely on heme supplied by its vertebrate host or its insect vector, the tsetse fly. In the host bloodstream T. brucei scavenges heme via haptoglobin-hemoglobin (HpHb) receptor-mediated endocytosis occurring in the flagellar pocket. However, in the procyclic developmental stage, in which T. brucei is confined to the tsetse fly midgut, this receptor is apparently not expressed, suggesting that T. brucei takes up heme by a different, unknown route. To define this alternative route, we functionally characterized heme transporter TbHrg in the procyclic stage. RNAi-induced down-regulation of TbHrg in heme-limited culture conditions resulted in slower proliferation, decreased cellular heme, and marked changes in cellular morphology so that the cells resemble mesocyclic trypomastigotes. Nevertheless, the TbHrg KO developed normally in the tsetse flies at rates comparable with wild-type cells. T. brucei cells overexpressing TbHrg displayed up-regulation of the early procyclin GPEET and down-regulation of the late procyclin EP1, two proteins coating the T. brucei surface in the procyclic stage. Light microscopy of immunostained TbHrg indicated localization to the flagellar membrane, and scanning electron microscopy revealed more intense TbHrg accumulation toward the flagellar pocket. Based on these findings, we postulate that T. brucei senses heme levels via the flagellar TbHrg protein. Heme deprivation in the tsetse fly anterior midgut might represent an environmental stimulus involved in the transformation of this important human parasite, possibly through metabolic remodeling.

• Klíčová slova
• differentiation, flagellum, heme, import, parasite, procyclin, transporter, trypanosome,
• MeSH
• biologický transport MeSH
• down regulace MeSH
• flagella metabolismus   MeSH
• hem metabolismus   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• membránové transportní proteiny metabolismus   MeSH
• mikroskopie elektronová rastrovací MeSH
• moucha tse-tse parazitologie   MeSH
• proliferace buněk MeSH
• protozoální proteiny metabolismus   MeSH
• receptory buněčného povrchu metabolismus   MeSH
• RNA interference MeSH
• sekvence aminokyselin MeSH
• stadia vývoje MeSH
• transgeny MeSH
• Trypanosoma brucei brucei metabolismus   MeSH
• vývojová regulace genové exprese MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hem MeSH
• hemoglobin-haptoglobin receptor MeSH Prohlížeč
• membránové transportní proteiny MeSH
• protozoální proteiny MeSH
• receptory buněčného povrchu MeSH

Haem and iron homeostasis in most eukaryotic cells is based on a balanced flux between haem biosynthesis and haem oxygenase-mediated degradation. Unlike most eukaryotes, ticks possess an incomplete haem biosynthetic pathway and, together with other (non-haematophagous) mites, lack a gene encoding haem oxygenase. We demonstrated, by membrane feeding, that ticks do not acquire bioavailable iron from haemoglobin-derived haem. However, ticks require dietary haemoglobin as an exogenous source of haem since, feeding with haemoglobin-depleted serum led to aborted embryogenesis. Supplementation of serum with haemoglobin fully restored egg fertility. Surprisingly, haemoglobin could be completely substituted by serum proteins for the provision of amino-acids in vitellogenesis. Acquired haem is distributed by haemolymph carrier protein(s) and sequestered by vitellins in the developing oocytes. This work extends, substantially, current knowledge of haem auxotrophy in ticks and underscores the importance of haem and iron metabolism as rational targets for anti-tick interventions.

• Klíčová slova
• biochemistry, haem auxotrophy, haem oxygenase, haematophagy, infectious disease, iron metabolism, microbiology, ticks,
• MeSH
• fertilita MeSH
• hem metabolismus   MeSH
• klíšťata metabolismus   fyziologie   MeSH
• rozmnožování MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hem MeSH

• MeSH
• biologická evoluce MeSH
• energetický metabolismus MeSH
• hem metabolismus   MeSH
• interakce hostitele a parazita MeSH
• parazitární nemoci metabolismus   terapie   MeSH
• paraziti metabolismus   MeSH
• Plasmodium malariae metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hem MeSH