A high-throughput and multiplex microsphere immunoassay based on non-structural protein 1 can discriminate three flavivirus infections.

Jasmine Tyson,Duane J. Gubler,Jih-Jin Tsai,Wei-Kung Wang,Axel T. Lehrer,Susan L. Stramer,Wen-Yang Tsai,Vivek R. Nerurkar,Ludvig Mässgård

doi:10.1371/journal.pntd.0007649

Abstract

The explosive spread of Zika virus (ZIKV) and associated complications in flavivirus-endemic regions underscore the need for sensitive and specific serodiagnostic tests to distinguish ZIKV, dengue virus (DENV) and other flavivirus infections. Compared with traditional envelope protein-based assays, several nonstructural protein 1 (NS1)-based assays showed improved specificity, however, none can detect and discriminate three flaviviruses in a single assay. Moreover, secondary DENV infection and ZIKV infection with previous DENV infection, both common in endemic regions, cannot be discriminated. In this study, we developed a high-throughput and multiplex IgG microsphere immunoassay (MIA) using the NS1 proteins of DENV1-DENV4, ZIKV and West Nile virus (WNV) to test samples from reverse-transcription-polymerase-chain reaction-confirmed cases, including primary DENV1, DENV2, DENV3, WNV and ZIKV infections, secondary DENV infection, and ZIKV infection with previous DENV infection. Combination of four DENV NS1 IgG MIAs revealed a sensitivity of 94.3% and specificity of 97.2% to detect DENV infection. The ZIKV and WNV NS1 IgG MIAs had a sensitivity/specificity of 100%/87.9% and 86.1%/78.4%, respectively. A positive correlation was found between the readouts of enzyme-linked immunosorbent assay and MIA for different NS1 tested. Based on the ratio of relative median fluorescence intensity of ZIKV NS1 to DENV1 NS1, the IgG MIA can distinguish ZIKV infection with previous DENV infection and secondary DENV infection with a sensitivity of 88.9–90.0% and specificity of 91.7–100.0%. The multiplex and high-throughput assay could be applied to serodiagnosis and serosurveillance of DENV, ZIKV and WNV infections in endemic regions.

Highlights

Despite a marked decrease of Zika virus (ZIKV) infection since late 2017, the specter of congenital Zika syndrome (CZS) and its re-emergence in flavivirus-endemic regions highlight the need for sensitive and specific diagnostic tests [1,2,3,4]
There was a decrease of Zika virus (ZIKV) infection since late 2017, the specter of congenital Zika syndrome and its re-emergence in flavivirus-endemic regions emphasize the need for sensitive and specific serological tests to distinguish ZIKV, dengue virus (DENV) and other flaviviruses
Combination of four DENV nonstructural protein 1 (NS1) assays revealed a sensitivity of 94.3% and specificity of 97.2%

Summary

Introduction

Despite a marked decrease of Zika virus (ZIKV) infection since late 2017, the specter of congenital Zika syndrome (CZS) and its re-emergence in flavivirus-endemic regions highlight the need for sensitive and specific diagnostic tests [1,2,3,4]. Since many individuals test for ZIKV infection beyond the period when RNA is detectable and most (~80%) of ZIKV infections are asymptomatic, serological tests remain as a key component of ZIKV confirmation [5,6]. Given that the envelope (E) protein is the major target of antibody response after flavivirus infection, different E antigens such as recombinant E protein, inactivated virions or virus-like particles have been developed for serological tests [10,11,12,13]. PRNT can confirm ZIKV-infected individuals who acquire ZIKV as the first flavivirus infection, known as primary ZIKV (pZIKV) infection, but often can only be interpreted as unspecified flavivirus infections for those who have experienced previous DENV or other flavivirus infections, limiting its application for ZIKV serodiagnosis in flavivirus-endemic regions

Methods

Results

Discussion

Conclusion