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

Yang Yang; Gary Wong; Baoguo Ye; Shihua Li; Shanqin Li; Haixia Zheng; Qiang Wang; Mifang Liang; George F Gao; Lei Liu; Yingxia Liu; Yuhai Bi

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

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

Authors

Yang Yang^{1

2}, Gary Wong^{1

2}, Baoguo Ye³, Shihua Li², Shanqin Li¹, Haixia Zheng¹, Qiang Wang^{1

2}, Mifang Liang⁴, George F Gao^{1

2

5

6}, Lei Liu⁷, Yingxia Liu⁸, Yuhai Bi^{9

10}

Affiliations

¹ Shenzhen Key Laboratory of Pathogen and Immunity, State Key Discipline of Infectious Disease, Shenzhen Third People's Hospital, Shenzhen, 518112, China.
² CAS Key Laboratory of Pathogenic Microbiology and Immunology, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease, Institute of Microbiology, Center for Influenza Research and Early-warning (CASCIRE), Chinese Academy of Sciences, Beijing, 100101, China.
³ China-Japan Union Hospital of Jilin University, Changchun, 130033, China.
⁴ Key Laboratory for Medical Virology, NHFPC, National Institute for Viral Disease Control and Prevention, China CDC, Beijing, 102206, China.
⁵ Office of Director-General, Chinese Center for Disease Control and Prevention, Beijing, 102206, China.
⁶ Savaid Medical School, University of Chinese Academy of Sciences, Beijing, 100049, China.
⁷ Shenzhen Key Laboratory of Pathogen and Immunity, State Key Discipline of Infectious Disease, Shenzhen Third People's Hospital, Shenzhen, 518112, China. [email protected].
⁸ Shenzhen Key Laboratory of Pathogen and Immunity, State Key Discipline of Infectious Disease, Shenzhen Third People's Hospital, Shenzhen, 518112, China. [email protected].
⁹ Shenzhen Key Laboratory of Pathogen and Immunity, State Key Discipline of Infectious Disease, Shenzhen Third People's Hospital, Shenzhen, 518112, China. [email protected].
¹⁰ CAS Key Laboratory of Pathogenic Microbiology and Immunology, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease, Institute of Microbiology, Center for Influenza Research and Early-warning (CASCIRE), Chinese Academy of Sciences, Beijing, 100101, China. [email protected].

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.

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

MeSH terms

Conserved Sequence
DNA Primers / genetics
Humans
Molecular Diagnostic Techniques / methods*
Real-Time Polymerase Chain Reaction / methods*
Reverse Transcriptase Polymerase Chain Reaction / methods*
Sensitivity and Specificity
Viral Envelope Proteins / genetics
Zika Virus / genetics
Zika Virus / isolation & purification*
Zika Virus Infection / diagnosis*
Zika Virus Infection / virology

Substances

DNA Primers
Viral Envelope Proteins