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Comparative Study
. 2020 Jun;127:104374.
doi: 10.1016/j.jcv.2020.104374. Epub 2020 Apr 20.

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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Free PMC article
Comparative Study

Interpret With Caution: An Evaluation of the Commercial AusDiagnostics Versus In-House Developed Assays for the Detection of SARS-CoV-2 Virus

H Rahman et al. J Clin Virol. .
Free PMC article

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.

Keywords: Covid-19; NAT; SARS-CoV-2.

Figures

Fig. 1
Fig. 1
Example of a false positive AusDiagnostics assay result showing a flat, non-sigmoidal amplification curve.
Fig. 2
Fig. 2
Comparison of cycle threshold values of the E gene versus RdRp gene for the detection of SARS-CoV-2.. E gene mean Ct value = 21.75, RdRp mean Ct value = 28.1, p = 0.0031.

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