The Complex Interpretation and Management of Zika Virus Test Results

Kenneth W. Lin, MD, MPH; John D. Kraemer, JD, MPH; Rachael Piltch-Loeb, MSPH; Michael A. Stoto, PhD

Disclosures

J Am Board Fam Med. 2018;31(6):924-930. 

In This Article

Zika Virus Test Types and Performance

Interpreting Zika virus test results is complicated by the absence of a single testing approach with superior validity across contexts and populations. Rather, there is substantial variation across 3 dimensions: (1) what the test seeks to detect; (2) the test's sensitivity and specificity under idealized conditions; and (3) moderators that affect test validity under real world conditions, such as pregnancy status, the timing of a test, what fluids are tested, and cross-reactivity with other, similar viruses.

The US Food and Drug Administration (FDA) has authorized 2 classes of Zika virus tests for human use: molecular and serologic (Table 1). Molecular tests identify individuals with a current infection by identifying Zika virus RNA in blood serum or urine. Most molecular tests use polymerase chain reaction (PCR), although 1 uses transcription-mediated amplification.

Exact estimates of sensitivity and specificity are not available because there is no gold standard comparison. However, currently available molecular tests have high specificity—they correctly identify patients without Zika virus—under idealized conditions.[4] Their sensitivity varies substantially, however. The tests currently authorized for use in the United States have labeled limits of detection ranging across 3 orders of magnitude.[5] As a result, while some tests are extremely sensitive,[6] the least sensitive tests would have failed to detect over half of the cases in a large Nicaraguan cohort.[7]

Several factors influence the sensitivity of molecular tests, of which the timing of the test is most important. These tests generally cease returning a positive test once a person is no longer viremic, usually around 2 weeks after symptom onset[8] to a maximum of a few weeks after symptoms resolve.[5,9] Furthermore, only a small fraction of infected people develop symptoms that are likely to be identified as Zika virus,[10,11] complicating the correct timing of tests. Several studies suggest that whole-blood samples have higher virus levels and that detectable virus may persist longer than in serum or urine samples.[12] Coinfection with related viruses, which tends to reduce virus levels, may reduce sensitivity;[13] however, cross-reactivity, which would reduce test specificity, is rarely reported.[5] Finally, limited evidence suggests that pregnant women may have higher and longer-lasting viremia, which would increase the test's sensitivity.[14]

Serologic tests, on the other hand, use the enzyme-linked immunosorbent assay (ELISA) to identify antibodies against Zika virus,[15] which indicates past infection. All US tests identify immunoglobulin M (IgM), which develop shortly (usually a few days) after symptom onset and decline after about 3 months (although IgM can sometimes be detected for over a year).[10] A few tests identify IgG, which develops later, although as early as 10 days after symptoms in some cases, but persists for years.[16] Under ideal conditions, serologic tests have sensitivities ranging from 37% to 100% and specificities between 66% and 100%.[15]

The performance of serologic tests depends substantially on the timing of testing relative to infection. Immediately after infection, antibodies against Zika virus will not have developed yet. Too long after infection, the IgM antibody against Zika virus will no longer be present. IgG will persist but cannot distinguish recent from previous infection.[17] Serologic tests are more likely to cross-react with viruses that are related to Zika virus, including dengue, West Nile virus, and yellow fever virus. The risk of cross-reactivity depends on where a person has lived and traveled. While limited data exist, 1 commercially available test found cross-reactivity 19% of the time for IgG and 8% for IgM.[4] Another test cross-reacted in 40% of cases where dengue but not Zika virus IgM was present.[17]

Because there is a significant risk of false positives and cross-reaction with ELISA tests, the CDC recommends that negative tests be considered definitive but positive or indeterminate tests be followed up with a plaque reduction neutralization test for both Zika virus and dengue. The plaque reduction neutralization test is highly accurate and usually able to differentiate between antibodies against Zika virus and other related viruses, but it can only be run by CDC or other specialized labs. It takes up to 4 weeks for results and, like all antibody tests, it cannot definitely determine when infection occurred.[10]

Because of the differences in the sensitivity and specificity of available tests, clinicians must know which tests were done and when they were done relative to the approximate time of infection (estimated by the time of symptom onset and/or known exposure) to properly interpret the results. Any test's positive and negative predictive value, however, also depends on the pretest probability of infection (see Sidebar below). Therefore, it is also important to understand patient characteristics, including why they were tested.

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