Use of a Blockade-of-Binding ELISA and Microneutralization Assay to Evaluate Zika Virus Serostatus in Dengue-Endemic Areas

Zika virus (ZIKV) serological diagnostics are compromised in areas where dengue viruses (DENV) co-circulate because of their high levels of protein sequence homology. Here, we describe the characterization of a Zika blockade-of-binding ELISA (Zika BOB) and a Zika microneutralization assay (Zika MN) for the detection of ZIKV nonstructural protein 1 (NS1)–specific antibodies and ZIKV neutralizing antibodies, respectively. Zika BOB and Zika MN cutoffs were established as 10 and 100 endpoint titers, respectively, using samples collected pre- and post-virologically confirmed ZIKV infection from subjects living in DENV-endemic areas. Specificity of the assays was equally high, whereas sensitivity of Zika BOB was lower than that of Zika MN, especially in samples collected > 6 months post-infection. Immunosurveillance analysis, using combined results from both Zika BOB and Zika MN, carried out also in DENV-endemic regions in Colombia, Honduras, Mexico, and Puerto Rico before (2013–2014) and after (2017–2018) ZIKV introduction in the Americas suggests unapparent ZIKV seroprevalence rates ranged from 25% to 80% over the specified period of time in the regions investigated.