Extracellular non-coding RNA signatures of the metacestode stage of Echinococcus multilocularis

• Published in: PLoS neglected tropical diseases Vol. 14; no. 11; p. e0008890
• Main Authors: Ancarola, María Eugenia, Lichtenstein, Gabriel, Herbig, Johannes, Holroyd, Nancy, Mariconti, Mara, Brunetti, Enrico, Berriman, Matthew, Albrecht, Krystyna, Marcilla, Antonio, Rosenzvit, Mara Cecilia, Kamenetzky, Laura, Brehm, Klaus, Cucher, Marcela
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 01.11.2020; Public Library of Science (PLoS)
• Subjects: Acetates; Acetic acid; Animals; Biology and life sciences; Biomarkers; Culture Media, Conditioned - analysis; Echinococcus multilocularis - genetics; Echinococcus multilocularis - metabolism; Electron microscopy; Engineering and Technology; Extracellular; Extracellular Vesicles - metabolism; Gene expression; Glutaraldehyde; High-Throughput Nucleotide Sequencing; Host-Parasite Interactions; Humans; Medical research; Medical sciences; Medicine; Medicine and Health Sciences; Mice; MicroRNAs - genetics; MicroRNAs - isolation & purification; Molecules; Nanoparticles; Nematodes; Non-coding RNA; Nucleic acids; Parasites; Phenols; Real-Time Polymerase Chain Reaction; Research and analysis methods; RNA; Secretion; Sequence Analysis, RNA; Transmission; Transmission electron microscopy; Tropical diseases; Uranyl acetate
• ISSN: 1935-2735; 1935-2727; 1935-2735
• DOI: 10.1371/journal.pntd.0008890

Extracellular RNAs (ex-RNAs) are secreted by cells through different means that may involve association with proteins, lipoproteins or extracellular vesicles (EV). In the context of parasitism, ex-RNAs represent new and exciting communication intermediaries with promising potential as novel biomarkers. In the last years, it was shown that helminth parasites secrete ex-RNAs, however, most work mainly focused on RNA secretion mediated by EV. Ex-RNA study is of special interest in those helminth infections that still lack biomarkers for early and/or follow-up diagnosis, such as echinococcosis, a neglected zoonotic disease caused by cestodes of the genus Echinococcus. In this work, we have characterised the ex-RNA profile secreted by in vitro grown metacestodes of Echinococcus multilocularis, the casuative agent of alveolar echinococcosis. We have used high throughput RNA-sequencing together with RT-qPCR to characterise the ex-RNA profile secreted towards the extra- and intra-parasite milieus in EV-enriched and EV-depleted fractions. We show that a polarized secretion of small RNAs takes place, with microRNAs mainly secreted to the extra-parasite milieu and rRNA- and tRNA-derived sequences mostly secreted to the intra-parasite milieu. In addition, we show by nanoparticle tracking analyses that viable metacestodes secrete EV mainly into the metacestode inner vesicular fluid (MVF); however, the number of nanoparticles in culture medium and MVF increases > 10-fold when metacestodes show signs of tegument impairment. Interestingly, we confirm the presence of host miRNAs in the intra-parasite milieu, implying their internalization and transport through the tegument towards the MVF. Finally, our assessment of the detection of Echinococcus miRNAs in patient samples by RT-qPCR yielded negative results suggesting the tested miRNAs may not be good biomarkers for this disease. A comprehensive study of the secretion mechanisms throughout the life cycle of these parasites will help to understand parasite interaction with the host and also, improve current diagnostic tools.