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. 2017 May 19;32(3):199–206. doi: 10.1007/s12250-017-3958-y

Development of a reverse transcription quantitative polymerase chain reaction-based assay for broad coverage detection of African and Asian Zika virus lineages

Yang Yang 1,2,#, Gary Wong 1,2,#, Baoguo Ye 3,#, Shihua Li 2, Shanqin Li 1, Haixia Zheng 1, Qiang Wang 1,2, Mifang Liang 4, George F Gao 1,2,5,6, Lei Liu 1,✉, Yingxia Liu 1,✉, Yuhai Bi 1,2,✉
PMCID: PMC6598927  PMID: 28530022

Abstract

The Zika virus (ZIKV) is an arbovirus that has spread rapidly worldwide within recent times. There is accumulating evidence that associates ZIKV infections with Guillain-Barré Syndrome (GBS) and microcephaly in humans. The ZIKV is genetically diverse and can be separated into Asian and African lineages. A rapid, sensitive, and specific assay is needed for the detection of ZIKV across various pandemic regions. So far, the available primers and probes do not cover the genetic diversity and geographic distribution of all ZIKV strains. To this end, we have developed a one-step quantitative reverse transcription polymerase chain reaction (qRT-PCR) assay based on conserved sequences in the ZIKV envelope (E) gene. The detection limit of the assay was determined to be five RNA transcript copies and 2.94 × 10–3 50% tissue culture infectious doses (TCID50) of live ZIKV per reaction. The assay was highly specific and able to detect five different ZIKV strains covering the Asian and African lineages without nonspecific amplification, when tested against other flaviviruses. The assay was also successful in testing for ZIKV in clinical samples. Our assay represents an improvement over the current methods available for the detection ZIKV and would be valuable as a diagnostic tool in various pandemic regions.

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Keywords: Flavivirus, Zika virus(ZIKV), molecular diagnostics, qRT-PCR

Acknowledgments

This work was supported by the National Science and Technology Major Project (2016ZX10004222), the Sanming Project of Medicine in Shenzhen (ZDSYS2015 04301534057), the Key specialized fund for infectious diseases in Shenzhen City (No. 201161), the intramural special grant for influenza virus research from the Chinese Academy of Sciences (KJZD-EW-L09 and KJZD-EWL15), and the Shenzhen Science and Technology Research and Development Project (JCYJ20160427151920801 and JCYJ20160427153238750). G.F.G. is a leading principal investigator of the National Natural Science Foundation of China (NSFC) Innovative Research Group (81621091). Y.B. is supported by the Youth Innovation Promotion Association of Chinese Academy of Sciences (CAS) (2017122). G.W. is the recipient of a Banting Postdoctoral Fellowship from the Canadian Institutes of Health Research (CIHR) and the President’s International Fellowship Initiative from the CAS. We thank Dr. Chengfeng Qin and Gary Kobinger for supplying strains of DENVs and ZIKVs.

Footnotes

These authors contributed equally to this work.

Contributor Information

Lei Liu, Email: liulei3322@aliyun.com.

Yingxia Liu, Email: yingxialiu@hotmail.com.

Yuhai Bi, Email: beeyh@im.ac.cn.

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