Hemocyte siRNA uptake is increased by 5′ cholesterol-TEG addition in Biomphalaria glabrata, snail vector of schistosome

Abstract

Biomphalaria glabrata is one of the snail intermediate hosts of Schistosoma mansoni, the causative agent of intestinal schistosomiasis disease. Numerous molecular studies using comparative approaches between susceptible and resistant snails to S. mansoni infection have helped identify numerous snail key candidates supporting such susceptible/resistant status. The functional approach using RNA interference (RNAi) remains crucial to validate the function of such candidates. CRISPR-Cas systems are still under development in many laboratories, and RNA interference remains the best tool to study B. glabrata snail genetics. Herein, we describe the use of modified small interfering RNA (siRNA) molecules to enhance cell delivery, especially into hemocytes, the snail immune cells. Modification of siRNA with 5′ Cholesteryl TriEthylene Glycol (Chol-TEG) promotes cellular uptake by hemocytes, nearly eightfold over that of unmodified siRNA. FACS analysis reveals that more than 50% of hemocytes have internalized Chol-TEG siRNA conjugated to Cy3 fluorophores, 2 hours only after in vivo injection into snails. Chol-TEG siRNA targeting BgTEP1 (ThioEster-containing Protein), a parasite binding protein, reduced BgTEP1 transcript expression by 70–80% compared to control. The level of BgTEP1 protein secreted in the hemolymph was also decreased. However, despite the BgTEP1 knock-down at both RNA and protein levels, snail compatibility with its sympatric parasite is not affected suggesting functional redundancy among the BgTEP genes family in snail-schistosoma interaction.

Delivery of siRNAs against MERS-CoV in Vero and HEK-293 cells: A comparative evaluation of transfection reagents

“… Baddeley and Isalan, 2021 , Bruch et al, 2021 , Chafekar and Burtram, 2018 , Portet et al, 2021 , Sohrab et al, 2020a , Sohrab et al, 2020b , WHO, World Health Organization (WHO)., 2022 , Zhang and Jun, 2020 .…”

Accelerating snail vector genomics

“…However, it is imperative to expand this research and data generation to encompass the other snail vector taxa. The next steps are of paramount importance, these being to validate functions and interactions of shortlisted genes and confirm how they influence phenotypes in these genera, most likely through differential expression analysis, gene knockdown, or gene over-expression studies [ 24 , 25 ]. Findings from these functional studies could lead to novel approaches for controlling schistosome transmission, or directly controlling snail vectors themselves if factors concerning gastropod development, survival and environmental adaptation are uncovered.…”

Computational Design and Experimental Evaluation of MERS-CoV siRNAs in Selected Cell Lines

“…Oligonucleotide-based antiviral therapies using siRNA, shRNA, and miRNA are being evaluated. Long noncoding RNAs (lncRNAs) against cancers [9,10], bacterial infections [11], fungal infections [12,13], parasitic infections [14,15], viral infections [16]. Several potential RNA interference-based (RNAi) drugs have recently been reported [17].…”