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. 2019 Nov 25;58(1):46–53. doi: 10.1007/s12275-020-9384-0

Development of a neutralization assay based on the pseudotyped chikungunya virus of a Korean isolate

Woo-Chang Chung 1, Kwang Yeon Hwang 2, Suk-Jo Kang 3, Jae-Ouk Kim 4, Moon Jung Song 1,✉
PMCID: PMC7091072  PMID: 31768937

Abstract

The Chikungunya virus (CHIKV) belongs to the Alphavirus genus of Togaviridae family and contains a positive-sense single stranded RNA genome. Infection by this virus mainly causes sudden high fever, rashes, headache, and severe joint pain that can last for several months or years. CHIKV, a mosquito-borne arbovirus, is considered a re-emerging pathogen that has become one of the most pressing global health concerns due to a rapid increase in epidemics. Because handling of CHIKV is restricted to Biosafety Level 3 (BSL-3) facilities, the evaluation of prophylactic vaccines or antivirals has been substantially hampered. In this study, we first iden-tified the whole structural polyprotein sequence of a CHIKV strain isolated in South Korea (KNIH/2009/77). Phylogenetic analysis showed that this sequence clustered within the East/ Central/South African CHIKV genotype. Using this sequence information, we constructed a CHIKV-pseudotyped lenti-virus expressing the structural polyprotein of the Korean CHIKV isolate (CHIKVpseudo) and dual reporter genes of green fluorescence protein and luciferase. We then developed a pseudovirus-based neutralization assay (PBNA) using CHIKVpseudo. Results from this assay compared to those from the conventional plaque reduction neutralization test showed that our PBNA was a reliable and rapid method to evaluate the efficacy of neutralizing antibodies. More importantly, the neutralizing activities of human sera from CHIKV-infected individuals were quantitated by PBNA using CHIKVpseudo. Taken together, these results suggest that our PBNA for CHIKV may serve as a useful and safe method for testing the neutralizing activity of antibodies against CHIKV in BSL-2 facilities.

Keywords: Chikungunya virus, Korean isolate, Pseudovirus, Neutralization assay, Human serum

Acknowledgments

This research was supported by a grant (16172MFDS272) from the Ministry of Food and Drug Safety in 2016-2018, the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2018R1A2B6001363), and a grant from Korea Ministry of Health and Welfare (HI15C2971). The CHIKV strain isolated in Korea (KNIH/2009/77) was obtained from the National Culture Collection for Pathogens, Republic of Korea. The Luc2P-pLVX-IRES-ZsGreenl plasmid for CHIKVpseudo was a kind gift from Dr. Nam-Hyuk Cho at Seoul National University, Republic of Korea. Human serum samples used in this paper were originally collected as part of the Dengue Vaccine Initiative (DVI) study conducted in Ouagadougou, Burkina Faso, by Centre MURAZ and AGIR.

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