RNA interference dynamics in juvenile Fasciola hepatica are altered during in vitro growth and development

Paul McCusker, Wasim Hussain, Paul McVeigh, Erin McCammick, Nathan G. Clarke, Emily Robb, Fiona M. McKay, Peter M. Brophy, David J. Timson, Angela Mousley, Nikki J. Marks, Aaron G. Maule

Research output: Contribution to journalArticle

Abstract

For over a decade RNA interference (RNAi) has been an important molecular tool for functional genomics studies in parasitic flatworms. Despite this, our understanding of RNAi dynamics in many flatworm parasites, such as the temperate liver fluke (Fasciola hepatica), remains rudimentary. The ability to maintain developing juvenile fluke in vitro provides the opportunity to perform functional studies during development of the key pathogenic life stage. Here, we investigate the RNAi competence of developing juvenile liver fluke. Firstly, all life stages examined possess, and express, core candidate RNAi effectors encouraging the hypothesis that all life stages of F. hepatica are RNAi competent. RNAi effector analyses supported growing evidence that parasitic flatworms have evolved a separate clade of RNAi effectors with unknown function. Secondly, we assessed the impact of growth/development during in vitro culture on RNAi in F. hepatica juveniles and found that during the first week post-excystment liver fluke juveniles exhibit quantitatively lower RNAi mediated transcript knockdown when maintained in growth inducing media. This did not appear to occur in older in vitro juveniles, suggesting that rapidly shifting transcript dynamics over the first week following excystment alters RNAi efficacy after a single 24 h exposure to double stranded (ds)RNA. Finally, RNAi efficiency was found to be improved through use of a repeated dsRNA exposure methodology that has facilitated silencing of genes in a range of tissues, thereby increasing the utility of RNAi as a functional genomics tool in F. hepatica.

Original languageEnglish
Pages (from-to)46-55
Number of pages10
JournalInternational Journal for Parasitology: Drugs and Drug Resistance
Volume14
DOIs
Publication statusPublished - 19 Aug 2020

Bibliographical note

© 2020 The Authors. Published by Elsevier Ltd on behalf of Australian Society for Parasitology. This is an open access article under the CC BY license

Keywords

  • Argonaute
  • Calmodulin
  • Fasciola
  • Liver fluke
  • RNAi
  • σGST

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    McCusker, P., Hussain, W., McVeigh, P., McCammick, E., Clarke, N. G., Robb, E., McKay, F. M., Brophy, P. M., Timson, D. J., Mousley, A., Marks, N. J., & Maule, A. G. (2020). RNA interference dynamics in juvenile Fasciola hepatica are altered during in vitro growth and development. International Journal for Parasitology: Drugs and Drug Resistance, 14, 46-55. https://doi.org/10.1016/j.ijpddr.2020.08.004