Serologic Testing for Zika Virus: Comparison of Three Zika Virus IgM-Screening Enzyme-Linked Immunosorbent Assays and Initial Laboratory Experiences

Abstract

Serologic evaluation for Zika virus (ZIKV) infection currently includes an initial screen using an anti-ZIKV IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) followed by supplemental testing of specimens with nonnegative results by a plaque reduction neutralization test (PRNT). We compared the performance characteristics of three ELISAs for the detection of IgM class antibodies to ZIKV, including the Centers for Disease Control and Prevention (CDC) Zika MAC-ELISA, the InBios ZIKV Detect MAC-ELISA, and the Euroimmun anti-Zika Virus IgM ELISA. Additionally, we present our initial experiences with ZIKV serologic testing from a national reference laboratory perspective. Using both retrospectively and prospectively collected specimens from patients with possible ZIKV infection, we show that the CDC and InBios MAC-ELISAs perform comparably to each other, with positive agreement, negative agreement, and interrater kappa values ranging from 87.5% to 93.1%, 95.7% to 98.5%, and 0.52 to 0.83, respectively. In contrast, comparison of the Euroimmun ZIKV ELISA to either the CDC or InBios MAC-ELISAs resulted in positive agreement, negative agreement, and interrater kappa values ranging from 17.9% to 42.9%, 91.7% to 98.6%, and 0.10 to 0.39, respectively. Among the 19 prospective samples submitted for PRNT, nine were negative, eight specimens had neutralizing antibodies to a flavivirus (unable to be identified), and one sample each was confirmed for ZIKV or dengue virus infection. This study highlights the ongoing challenges associated with serologic diagnosis of ZIKV infection. Although the availability of a commercial serologic test for ZIKV has greatly expanded the national capacity for such testing, the need to further characterize and improve these assays, particularly with regard to specificity, remains. KEYWORDS Zika virus, serologyZ ika virus (ZIKV) emerged from obscurity in early 2015 following its detection in Bahia, Brazil (1). Over the next year, ZIKV spread rapidly throughout Latin America, the Caribbean, and into the southern United States, resulting in a major and still ongoing international outbreak (2). Currently, over 1 million cases of suspected or confirmed ZIKV infection have been documented in the Americas by the Pan American Health Organization (3). ZIKV, a single-stranded RNA virus and member of the Flavivirus genus, is primarily transmitted through infected Aedes species mosquitoes, which are also primary vectors for dengue (DENV) and chikungunya (CHIKV) viruses, both of which cocirculate in many regions where ZIKV is now considered endemic (4). ZIKV transmis-

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“…The assessment of ZIKV in urine or whole blood would have been a valuable addition to the study protocol and should be considered in future studies 28. The lack of detected IgM responses may be due to the limited sensitivity of the serological method used for IgM detection (Euroimmun ZIKV ELISA) 29, 30. Furthermore, IgM responses can be attenuated in infected individuals with flavivirus infections in the past31, 32, 33 and ZIKV serology is further complicated by extensive cross‐reactivity with other endemic flaviviruses 21, 34.…”

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“…43 Serologic evaluation for ZIKV infection includes an initial screening by enzyme-linked immunosorbent assay (ELISA) to detect specific class M immunoglobulins (IgM) against ZIKV followed by confirmation testing by plaque reduction neutralization test (PRNT) due to crossreactivity with other flaviviruses. 45 Indirect immunofluorescence and In the cohort from French Guiana, we proposed a definition based on early neonatal clinical, biological, and imaging features, differentiating neonates with signs potentially associated with CZS and those with complications compatible with CZS. 50 In future epidemics, other more subtle signs might be associated with CZS, and some infected infants may only develop anomalies in childhood.…”

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“…Furthermore, other viruses co-circulating with ZIKV (e.g. dengue and Chikungunya viruses) may also present similarly and serologic tests for ZIKV may cross-react with antibodies from past infections with related flaviviruses(31, 32), leading to equivocal results. The limited time window for detecting ZIKV nucleic acids also makes it difficult to detect ZIKV RNA if maternal testing did not occur at the right time.…”