Abstract
BackgroundChikungunya virus (CHIKV) is a re-emerging alphavirus that causes chikungunya fever and persistent arthralgia in humans. Currently, there is no effective vaccine or antiviral against CHIKV infection. Therefore, this study evaluates whether RNA interference which targets at viral genomic level may be a novel antiviral strategy to inhibit the medically important CHIKV infection.MethodsPlasmid-based small hairpin RNA (shRNA) was investigated for its efficacy in inhibiting CHIKV replication. Three shRNAs designed against CHIKV Capsid, E1 and nsP1 genes were transfected to establish stable shRNA-expressing cell clones. Following infection of stable shRNA cells clones with CHIKV at M.O.I. 1, viral plaque assay, Western blotting and transmission electron microscopy were performed. The in vivo efficacy of shRNA against CHIKV replication was also evaluated in a suckling murine model of CHIKV infection.ResultsCell clones expressing shRNAs against CHIKV E1 and nsP1 genes displayed significant inhibition of infectious CHIKV production, while shRNA Capsid demonstrated a modest inhibitory effect as compared to scrambled shRNA cell clones and non-transfected cell controls. Western blot analysis of CHIKV E2 protein expression and transmission electron microscopy of shRNA E1 and nsP1 cell clones collectively demonstrated similar inhibitory trends against CHIKV replication. shRNA E1 showed non cell-type specific anti-CHIKV effects and broad-spectrum silencing against different geographical strains of CHIKV. Furthermore, shRNA E1 clones did not exert any inhibition against Dengue virus and Sindbis virus replication, thus indicating the high specificity of shRNA against CHIKV replication. Moreover, no shRNA-resistant CHIKV mutant was generated after 50 passages of CHIKV in the stable cell clones. More importantly, strong and sustained anti-CHIKV protection was conferred in suckling mice pre-treated with shRNA E1.ConclusionTaken together, these data suggest the promising efficacy of anti-CHIKV shRNAs, in particular, plasmid-shRNA E1, as a novel antiviral strategy against CHIKV infection.
Highlights
Chikungunya virus (CHIKV) is an alphavirus member from the family Togaviridae
We investigated the efficacy of plasmidbased small hairpin RNA against CHIKV replication in three CHIKV-permissive cell lines, namely, HeLa, RD and BHK cells
Establishment of CHIKV Growth Kinetics in HeLa Cell Line In order to determine a suitable time point for investigating the anti-CHIKV efficacy of small hairpin RNA (shRNA) expressed in the stable cell clones, CHIKV growth kinetic in non-transfected HeLa cells was established at the start of the study
Summary
Chikungunya virus (CHIKV) is an alphavirus member from the family Togaviridae. CHIKV causes acute infection in humans with clinical symptoms characterized by sudden-onset chills and fever, headache, maculopapular rash and persistent arthralgia [1,2]. In recent years from 2004–2007, major outbreaks of CHIKV infection in Kenya, India and islands in Indian Ocean have involved a second vector, Aedes albopictus [5,6]. Chikungunya virus (CHIKV) is a re-emerging alphavirus that causes chikungunya fever and persistent arthralgia in humans. There is no effective vaccine or antiviral against CHIKV infection. This study evaluates whether RNA interference which targets at viral genomic level may be a novel antiviral strategy to inhibit the medically important CHIKV infection
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