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Interpret with caution: An evaluation of the commercial AusDiagnostics versus in-house developed assays for the detection of SARS-CoV-2 virus.

ABSTRACT: INTRODUCTION:There is limited data on the analytical performance of commercial nucleic acid tests (NATs) for laboratory confirmation of COVID-19 infection. METHODS:Nasopharyngeal, combined nose and throat swabs, nasopharyngeal aspirates and sputum was collected from persons with suspected SARS-CoV-2 infection, serial dilutions of SARS-CoV-2 viral cultures and synthetic positive controls (gBlocks, Integrated DNA Technologies) were tested using i) AusDiagnostics assay (AusDiagnostics Pty Ltd); ii) in-house developed assays targeting the E and RdRp genes; iii) multiplex PCR assay targeting endemic respiratory viruses. Discrepant SARS-CoV-2 results were resolved by testing the N, ORF1b, ORF1ab and M genes. RESULTS:Of 52 clinical samples collected from 50 persons tested, respiratory viruses were detected in 22 samples (42 %), including SARS CoV-2 (n?=?5), rhinovirus (n?=?7), enterovirus (n?=?5), influenza B (n?=?4), hMPV (n?=?5), influenza A (n?=?2), PIV-2 (n?=?1), RSV (n?=?2), CoV-NL63 (n?=?1) and CoV-229E (n?=?1). SARS-CoV-2 was detected in four additional samples by the AusDiagnostics assay. Using the in-house assays as the "gold standard", the sensitivity, specificity, positive and negative predictive values of the AusDiagnostics assay was 100 %, 92.16 %, 55.56 % and 100 % respectively. The Ct values of the real-time in-house-developed PCR assay targeting the E gene was significantly lower than the corresponding RdRp gene assay when applied to clinical samples, viral culture and positive controls (mean 21.75 vs 28.1, p?=?0.0031). CONCLUSIONS:The AusDiagnostics assay is not specific for the detection SARS-CoV-2. Any positive results should be confirmed using another NAT or sequencing. The case definition used to investigate persons with suspected COVID-19 infection is not specific.

SUBMITTER: Rahman H

PROVIDER: S-EPMC7195305 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Interpret with caution: An evaluation of the commercial AusDiagnostics versus in-house developed assays for the detection of SARS-CoV-2 virus.

Rahman H H Carter I I Basile K K Donovan L L Kumar S S Tran T T Ko D D Alderson S S Sivaruban T T Eden J-S JS Rockett R R O'Sullivan M V MV Sintchenko V V Chen S C-A SC Maddocks S S Dwyer D E DE Kok J J

Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology 20200420

<h4>Introduction</h4>There is limited data on the analytical performance of commercial nucleic acid tests (NATs) for laboratory confirmation of COVID-19 infection.<h4>Methods</h4>Nasopharyngeal, combined nose and throat swabs, nasopharyngeal aspirates and sputum was collected from persons with suspected SARS-CoV-2 infection, serial dilutions of SARS-CoV-2 viral cultures and synthetic positive controls (gBlocks, Integrated DNA Technologies) were tested using i) AusDiagnostics assay (AusDiagnostic ...[more]

PMID: 32361322

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Project description:IntroductionAccurate and sensitive measurement of antibodies is critical to assess the prevalence of infection, especially asymptomatic infection, and to analyze the immune response to vaccination during outbreaks and pandemics. A broad variety of commercial and in-house serological assays are available to cater to different laboratory requirements; however direct comparison is necessary to understand utility.Materials and methodsWe investigate the performance of six serological methods against SARS-CoV-2 to determine the antibody profile of 250 serum samples, including 234 RT-PCR-confirmed SARS-CoV-2 cases, the majority with asymptomatic presentation (87.2%) at 1-51 days post laboratory diagnosis. First, we compare to the performance of two in-house antibody assays: (i) an in-house IgG ELISA, utilizing UV-inactivated virus, and (ii) a live-virus neutralization assay (PRNT) using the same Cambodian isolate as the ELISA. In-house assays are then compared to standardized commercial anti-SARS-CoV-2 electrochemiluminescence immunoassays (Elecsys ECLIAs, Roche Diagnostics; targeting anti-N and anti-S antibodies) along with a flow cytometry based assay (FACS) that measures IgM and IgG against spike (S) protein and a multiplex microsphere-based immunoassay (MIA) determining the antibodies against various spike and nucleoprotein (N) antigens of SARS-CoV-2 and other coronaviruses (SARS-CoV-1, MERS-CoV, hCoVs 229E, NL63, HKU1).ResultsOverall, specificity of assays was 100%, except for the anti-S IgM flow cytometry based assay (96.2%), and the in-house IgG ELISA (94.2%). Sensitivity ranged from 97.3% for the anti-S ECLIA down to 76.3% for the anti-S IgG flow cytometry based assay. PRNT and in-house IgG ELISA performed similarly well when compared to the commercial ECLIA: sensitivity of ELISA and PRNT was 94.7 and 91.1%, respectively, compared to S- and N-targeting ECLIA with 97.3 and 96.8%, respectively. The MIA revealed cross-reactivity of antibodies from SARS-CoV-2-infected patients to the nucleocapsid of SARS-CoV-1, and the spike S1 domain of HKU1.ConclusionIn-house serological assays, especially ELISA and PRNT, perform similarly to commercial assays, a critical factor in pandemic response. Selection of suitable immunoassays should be made based on available resources and diagnostic needs.

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Interpret with caution: An evaluation of the commercial AusDiagnostics versus in-house developed assays for the detection of SARS-CoV-2 virus.

Publications

Interpret with caution: An evaluation of the commercial AusDiagnostics versus in-house developed assays for the detection of SARS-CoV-2 virus.

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