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.

• Keywords
• Cestoda, life cycle, parasite, reference genome, transcriptome,
• MeSH
• Cestoda * genetics   MeSH
• Cestode Infections * parasitology   MeSH
• Fishes genetics   MeSH
• Semen MeSH
• Gene Expression Profiling MeSH
• Transcriptome MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

We provide the first ultrastructural evidence of the secretion of extracellular vesicles (EVs) across all parasitic stages of the tapeworm Schistocephalus solidus (Müller, 1776) (Cestoda: Diphyllobothriidea) using a laboratory life cycle model. We confirmed the presence of EV-like bodies in all stages examined, including the hexacanth, procercoids in the copepod, Macrocyclops albidus (Jurine, 1820), plerocercoids from the body cavity of the three-spined stickleback, Gasterosteus aculeatus Linnaeus, and adults cultivated in artificial medium. In addition, we provide description of novel tegumental structures potentially involved in EV biogenesis and the presence of unique elongated EVs similar to those previously described only in Fasciola hepatica Linnaeus, 1758 (Trematoda), Hymenolepis diminuta (Rudolphi, 1819) (Cestoda), and Trypanosoma brucei Plimmer et Bradford, 1899 (Kinetoplastida).

• Keywords
• : EVs, Cestoda, ESP, TEM, novel tegumental structures, ultrastructure,
• MeSH
• Cestoda * MeSH
• Cestode Infections * veterinary   MeSH
• Copepoda MeSH
• Extracellular Vesicles * MeSH
• Smegmamorpha MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH

The present study provides the first ultrastructural data of the vitellogenesis in a cestode species of the cyclophyllidean family Paruterinidae, aiming to expand the limited data on the vitellogenesis in cyclophyllidean cestodes and to explore the potential of ultrastructural characters associated with vitellogenesis for phylogenetic and taxonomic studies of this order. The process of vitellocyte formation in Dictyterina cholodkowskii follows the general pattern observed in other tapeworms but exhibits several specific differences in the ultrastructure of vitelline cells. The vitellarium contains vitellocytes at various stages of maturation. The periphery of the vitellarium and the space between maturing vitellocytes are occupied by interstitial cells. Differentiation into mature vitellocytes is characterized by high secretory activity, which involves the development of granular endoplasmic reticulum, Golgi complexes, mitochondria and vitelline globules of various sizes. During vitellogenesis, the progressive fusion of these globules results in the formation of two large membrane-limited vitelline vesicles that eventually fuse into a single large vesicle. Mature vitellocytes are composed of a single vitelline vesicle, a high content of cytoplasmic organelles and have no nucleus. No traces of lipid droplets and glycogen granules are detected in the cytoplasm of mature vitellocytes, which might be related to biological peculiarities of this family, i.e. the release of eggs into environment within the tissues of the paruterine organ, which may serve as a source of nutrients for embryos.

• Keywords
• Cestode, Cyclophyllidean, Dictyterina cholodkowskii, Ultrastructure, Vitelline vesicles, Vitellocytes,
• MeSH
• Cestoda physiology   ultrastructure   MeSH
• Cytoplasm ultrastructure   MeSH
• Phylogeny * MeSH
• Vitelline Membrane physiology   MeSH
• Ovarian Follicle physiology   ultrastructure   MeSH
• Vitellogenesis physiology   MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

BACKGROUND: The tapeworm Ligula intestinalis (Diphyllobothriidea) is one of the most fascinating cestode parasites because it may cause parasitic castration of its second intermediate host, teleost freshwater fishes, due to inhibition of production of fish gonadotropic hormones. Large-sized (length up to 1 m) larvae called plerocercoids develop several months in the body cavity of freshwater fish and affect host behavior to facilitate transmission to the final host, a fish-eating bird. Vitellogenesis, i.e. formation of vitellocytes, is a key process in formation and nutrition of female gametes, oocytes in many flatworms, mainly parasitic Neodermata. The present study provides the first ultrastructural evidence in flatworms (Platyhelminthes) of the process that is interpreted as cytoplasmic-like cell death, i.e. a special case of programmed cell death (paraptosis) in vitellocytes of L. intestinalis. RESULTS: As molecular markers for paraptosis are not yet available, its identification was based on morphological criteria. Electron microscopy analyses revealed evident structural changes in vitellocytes associated with progressive cytoplasmatic vacuolation, swelling of the granular endoplasmic reticulum and mitochondria. In addition, the present study has shown that vitellocytes of L. intestinalis share numerous features in common with the members of other earliest evolved eucestodes. CONCLUSIONS: The present study indicates that paraptotic-like cell death may occur in parasitic flatworms (Neodermata). The presence of GER-bodies in mature vitellocytes indicates close relationship between the Diphyllobothriidea, Caryophyllidea and Spathebothriidea, which are considered as the earliest evolved groups of the Eucestoda. Beyond the general similarities, however, a number of differences exist between the morphology, chemical composition and amount of these inclusions which could be due to the variations in their embryonic development, life cycle strategies and definitive host groups.

• Keywords
• Cestoda, Diphyllobothriidea, Ligula intestinalis, Paraptosis, Ultrastructure, Vitellogenesis,
• Publication type
• Journal Article MeSH