Abstract
Chikungunya virus (CHIKV) has emerged as a global health concern due to its recent spread in both old and new world. So far, no CHIKV specific drug or vaccine is licensed for human use. In this study, we report production of Chikungunya virus like particles (CHIK-VLPs) using novel yeast expression system (Pichia pastoris) and its evaluation as vaccine candidate. The gene encoding structural polyprotein of CHIKV from a recent epidemic strain was cloned into yeast expression system. The multicopy integrants were processed for expression of CHIK-VLPs. The VLPs were purified and confirmed through electron microscopic analysis for their morphological identity with CHIKV. The in vitro and in vivo evaluation of CHIK-VLPs as vaccine candidate was determined in Balb/c mice. Induction of both humoral and cellular immune response was observed with different doses of CHIK-VLPs. The humoral immune response was studied through different techniques like enzyme linked immunosorbent assay, IgG Isotyping and plaque reduction neutralization test. CHIK-VLPs were found to elicit high titer of antibodies that are able to recognize native CHIKV. Higher level of IgG2a and IgG1 subtypes was identified suggestive of balanced Th1/Th2 response. Both in vitro and in vivo neutralization activity of CHIK-VLPs antibodies was observed even with low concentration, which shows its high specificity and neutralizing activity against two different CHIKV strains. Neonatal mice receiving anti-CHIK-VLPs antibodies were protected from CHIKV challenge. Induction of cellular immune response was confirmed through higher level of TNF-α, IL-10 and substantial level of IL-2, IL-4 and IFN-γ indicating a balanced response. This is the first report, where CHIK-VLPs has been expressed by Pichia pastoris and evaluated for neutralizing activity against CHIKV. These promising results indicate the utility of CHIK-VLPs as a promising vaccine candidate against emerging CHIKV.
Highlights
Chikungunya virus (CHIKV) is an emerging mosquito transmitted arbovirus of immense public health significance
The full CHIKV structural polyprotein gene, encoding 3.747 Kbp was transformed into pPIC9K yeast transfer vector (Invitrogen, USA) at AvrII and SnaBI sites resulting pPIC9K-CHIKV-C-E3-E2-6K-E1 construct
Positive transformants having full CHIKV structural polyprotein gene was confirmed by restriction digestion with AvrII and SnaBI and PCR amplification with CHIKYVLP forward and CHIKYVLP reverse primers
Summary
Chikungunya virus (CHIKV) is an emerging mosquito transmitted arbovirus of immense public health significance. It belongs to family Togaviridae and genus Alphavirus. The word Chikungunya comes from Makonde language, meaning “which bends up”, relating to the patients having contoured posture [1]. It reemerged in Kenya in 2004, from where virus spread to Indian Ocean islands, Asia, Africa and Europe. It was reported from Caribbean islands and many parts of Americas [2, 3]
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