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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., (Copyright © 2023 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

Parasitic flatworms have complex neuromuscular systems that serve important functions in their life cycles. However, our understanding of neurotransmission in parasitic flatworms is limited. Pioneering studies have suggested the presence of several classical neurotransmitter systems, but their molecular components have not been characterized in most cases. Because these components are conserved in bilaterian animals, we searched the genomes of parasitic flatworms for orthologs of genes required for neurotransmitter synthesis, vesicular transport, reuptake, and reception. Our results indicate that tapeworms have lost the genes that are specifically required in other animals for synaptic signaling using the classical neurotransmitters dopamine, tyramine, octopamine, histamine and gamma-aminobutyric acid (GABA). These results imply that these signaling pathways are either absent in these parasites, or that they require completely different molecular components in comparison with other animals. The orthologs of genes related to histaminergic and GABA signaling are also missing in trematodes (although Schistosoma-specific histaminergic receptors have been previously described). In contrast, conserved genes required for glutamatergic, serotonergic and cholinergic signaling could be found in all analyzed flatworms. We analyzed the expression of selected markers of each pathway in the tapeworm Hymenolepis microstoma by whole-mount in situ hybridization. Each marker was specifically expressed in the nervous system, although with different patterns. In addition, we analyzed the expression of proprotein convertase 2 as a marker of peptidergic cells. This gene showed the widest expression in the nervous system, but was also expressed in other tissues, suggesting additional roles of peptidergic signaling in tapeworm development and reproduction., (Copyright © 2018 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

Neuropeptide mediated signalling is an ancient mechanism found in almost all animals and has been proposed as a promising target for the development of novel drugs against helminths. However, identification of neuropeptides from genomic data is challenging, and knowledge of the neuropeptide complement of parasitic flatworms is still fragmentary. In this work, we have developed an evolution-based strategy for the de novo discovery of neuropeptide precursors, based on the detection of localised sequence conservation between possible prohormone convertase cleavage sites. The method detected known neuropeptide precursors with good precision and specificity in the models Drosophila melanogaster and Caenorhabditis elegans. Furthermore, it identified novel putative neuropeptide precursors in nematodes, including the first description of allatotropin homologues in this phylum. Our search for neuropeptide precursors in the genomes of parasitic flatworms resulted in the description of 34 conserved neuropeptide precursor families, including 13 new ones, and of hundreds of new homologues of known neuropeptide precursor families. Most neuropeptide precursor families show a wide phylogenetic distribution among parasitic flatworms and show little similarity to neuropeptide precursors of other bilaterian animals. However, we could also find orthologs of some conserved bilaterian neuropeptides including pyrokinin, crustacean cardioactive peptide, myomodulin, neuropeptide-Y, neuropeptide KY and SIF-amide. Finally, we determined the expression patterns of seven putative neuropeptide precursor genes in the protoscolex of Echinococcus multilocularis. All genes were expressed in the nervous system with different patterns, indicating a hidden complexity of peptidergic signalling in cestodes., (Copyright © 2016 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

The larval stages of the cestodes Echinococcus multilocularis and Echinococcus granulosus cause the important zoonoses known as larval echinococcoses. These larvae are protected by a unique, massive, mucin-based structure known as the laminated layer. The mucin glycans of the E. granulosus laminated layer are core 1- or core 2-based O-glycans in which the core Galpβ1-3 residue can initiate a chain comprising one to three additional Galpβ1-3 residues, a motif not known in mammalian carbohydrates. This chain can be capped with a Galpα1-4 residue, and can be ramified with GlcNAcpβ1-6 residues. These, as well as the GlcNAcpβ1-6 residue in core 2, can be decorated with the Galpα1-4Galpβ1-4 disaccharide. Here we extend our analysis to the laminated layer of E. multilocularis, showing that the non-decorated cores, together with Galpβ1-3(Galpα1-4Galpβ1-4GlcNAcpβ1-6)GalNAc, comprise over 96% of the glycans in molar terms. This simple laminated layer glycome is exhibited by E. multilocularis grown either in vitro or in vivo. Interestingly, all the differences with the complex laminated layer glycome found in E. granulosus may be explained in terms of strongly reduced activity in E. multilocularis of a putative glycosyltransferase catalysing the elongation with Galpβ1-3. Comparative inter-species analysis of available genomic and transcriptomic data suggested a candidate for this enzyme, amongst more than 20 putative (non-core 1) Gal/GlcNAc β1-3 transferases present in each species as a result of a taeniid-specific gene expansion. The candidate gene was experimentally verified to be transcribed at much higher levels in the larva of E. granulosus than that of E. multilocularis., (Copyright © 2016 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.)