RNA interference in < i > Fasciola gigantica < /i > : Establishing and optimization of experimental RNAi in the newly excysted juveniles of the fluke

by Arun Anandanarayanan, Opinder Krishen Raina, Hniang Lalrinkima, Ajayta Rialch, Muthu Sankar, Anju Varghese Fasciolosis caused byFasciola gigantica is a neglected tropical disease but a constraint on the growth and productivity of cattle, buffaloes and sheep in the tropical countries of Asia and Africa. Resistance to commonly used anthelmintics inFasciola has increased the need to search for alternative therapeutic targets. RNA interference is the current tool of choice in the search for such targets inFasciola. The susceptibility of juvenileFasciola hepatica to double stranded (ds) RNA induced RNAi has been established but inF.gigantica a single preliminary report on RNAi induced mRNA transcript knockdown is available. Here we optimized conditions for RNAi in the liver flukeF.gigantica targeting six genes including superoxide dismutase (SOD), σ class of glutathione-s-transferase (GST), cathepsin (Cat) L1-D, Cat B1, Cat B2 and Cat B3 that showed robust transcriptional silencing of the targets following exposure of the newly excysted juveniles (NEJs) to long (170–223 nt) dsRNA. Knockdown was shown to be concentration dependent with sign ificant mRNA transcript suppression occurring at 5 ng / μl that showed further suppression with the increase in the dsRNA concentration. The dsRNA induced persistent silencing of the mRNA transcript of SOD and σGST up to 15 days of observation. Delivery of the long dsRNA and siRNA to the newly exc ysted juveniles by soaking method was f...
Source: PLoS Neglected Tropical Diseases - Category: Tropical Medicine Authors: Source Type: research