The Advisory Committee on Immunization Practices (ACIP) recommended that dengue pre-vaccination screening tests for Dengvaxia administration have at least 98% specificity and 75% sensitivity. This study evaluates the performance of commercial anti-DENV IgG tests to identify tests that could be used for pre-vaccination screening. First, for seven tests, we evaluated sensitivity and specificity in early convalescent dengue virus (DENV) infection, using 44 samples collected 7-30 days after symptom onset and confirmed by RT-PCR. Next, for the five best-performing tests and two additional tests (with and without an external test reader) that became available later, we evaluated performance to detect past dengue infection among a panel of 44 specimens collected in 2018-2019 from healthy 9- to 16-year-old children from Puerto Rico. Finally, a full-scale evaluation was done with the four best-performing tests using 400 specimens from the same population. We used virus focus reduction neutralization test and an in-house DENV IgG ELISA as reference standards. Of seven tests, five showed ≥75% sensitivity in detecting anti-DENV IgG in early convalescent specimens with low cross-reactivity to the Zika virus. For the detection of previous DENV infections, the tests with the highest performance were the Euroimmun NS1 IgG ELISA (sensitivity 84.5%, specificity 97.1%) and CTK Dengue IgG rapid test R0065C with the test reader (sensitivity 76.2% specificity 98.1%). There are IgG tests available that can be used to accurately classify individuals with previous DENV infection as eligible for dengue vaccination to support safe vaccine implementation. IMPORTANCE: The Advisory Committee on Immunization Practices (ACIP) has set forth recommendations that dengue pre-vaccination screening tests must exhibit at least 98% specificity and 75% sensitivity. Our research rigorously assesses the performance of various commercial tests against these benchmarks using well-characterized specimens from Puerto Rico. The findings from our study are particularly relevant given FDA approval and ACIP recommendation of Sanofi Pasteur's Dengvaxia vaccine, highlighting the need for accurate pre-vaccination screening tools.

BACKGROUND: Anti-dengue virus (DENV) immunoglobulin M (IgM) seroconversion has been the reference standard for dengue diagnosis. However, paired specimens are rarely obtained, and the interval for this testing negates its usefulness in guiding clinical case management. The presence of DENV viremia and appearance of IgM during the febrile phase of dengue provides the framework for dengue laboratory diagnosis by using a single serum specimen. METHODS: Archived paired serum specimens (n = 1234) from patients with laboratory-confirmed dengue from 2005 through 2011 were used to determine the diagnostic performance of real-time reverse transcription polymerase chain reaction (RT-PCR), for detection of DENV serotypes 1-4, and enzyme-linked immunosorbent assays (ELISAs), for detection of DENV nonstructural protein 1 (NS1) antigen and anti-DENV IgM. RESULTS: During 1-3 days after illness onset, real-time RT-PCR and NS1 antigen testing detected 82%-69% and 90%-84% of cases, respectively, as viremia levels declined, while anti-DENV IgM ELISA detected 5%-41% of cases as antibody appeared. Over the 10-day period of the febrile phase of dengue, the cumulative effect of using these 3 types of tests in a diagnostic algorithm confirmed ≥90% of dengue cases. CONCLUSIONS: The use of molecular or NS1 antigen tests to detect DENV and one to detect anti-DENV IgM in a single serum specimen collected during the first 10 days of illness accurately identified ≥90% of dengue primary and secondary cases.

A laboratory testing algorithm was evaluated to confirm West Nile virus (WNV) infection in human serum following its introduction in Puerto Rico in 2007. This testing algorithm used two standard diagnostic assays, the IgM antibody capture enzyme linked immunosorbent assay (MAC ELISA) and real time reverse transcriptase polymerase chain reaction (RT-PCR), along with two non-conventional assays, the non-structural protein 1 (NS1) ELISA and plaque reduction neutralization test at 90% (PRNT(90)) with IgG depletion for dengue virus (DENV) and WNV. A total of 2321 serum samples from suspected WNV human cases were submitted for testing. Approximately one third (867, 37%) were cross reactive for DENV and WNV by MAC-ELISA and had negative RT-PCR results for both viruses. Of a subset of 43 samples tested, 31 (72%) of these cases were identified as dengue in the PRNT(90) IgG depletion assay and 8 (19%) were positive in the dengue NS1 antigen ELISA. These two assays combined differentiated 36 (84%) of the samples that could not be diagnosed using the standard diagnostic testing methods.

Annually, over 2.5 billion people are at risk for infection by dengue virus (DENV), while between 50 and 100 million people contract the disease. There is an urgent need for alternative diagnostic tools that can detect DENV during acute infection. Recent studies have shown that dengue nonstructural protein 1 (NS1) is detectable in the blood as early as the onset of symptoms and persist well into the convalescent phase of the infection. We evaluated the utility of the BioRad Platelia Dengue NS1 Antigen Capture(R) kit in combination with real time reverse transcriptase polymerase chain reaction (RT-PCR) and IgM antibody capture enzyme linked immunosorbent assay (MAC ELISA) for refining a new diagnostic algorithm for the diagnosis of an acute or convalescent DENV from a single clinical sample. We tested the BioRad kit in three panels of sera. These panels were designed to evaluate the sensitivity of the NS1 kit in (1) early convalescent samples, (2) PCR false negative acute samples, and (3) confirmed secondary dengue infections with IgM negative convalescent samples. Results show that the NS1 can be detected in 22% serum collected after 10 days post onset of illness and in 22% of samples that did not elicit an IgM response. Additionally, NS1 was detected in 37% of the PCR false negative acute samples tested suggesting that NS1 detection may be valuable in increasing the sensitivity of current acute phase diagnostics. These results will improve diagnosis in a single acute or early convalescent sample for disease surveillance and clinical diagnosis.