Causes and Clinical Relevance of Inconclusive SARS-CoV-2 Real-Time Reverse Transcription-PCR Test Results

Aram Kim1   Heerah Lee1   Kyu Wha Hur1   Heungsup Sung1   Mi-Na Kim1*   

1 Department of Laboratory Medicine, Asan Medical Center, University of Ulsan College of Medicine, Seoul,

* Corresponding author: Tel: +82-2-3010-4511, Fax: +82-2-478-0884, E-mail: mnkim@amc.seoul.kr

Abstract

Background: Inconclusive SARS-CoV-2 real-time reverse transcription-PCR (rRT-PCR) test results, which are positive for one or more target genes but not all, are problematic in clinical laboratories. In this study, we aimed to investigate the cause and clinical relevance of such inconclusive results.

Methods: rRT-PCR was performed using the Allplex 2019-nCoV assay kit (Seegene Inc., Korea) targeting the following three genes: E, RdRp, and N. For all inconclusive test results reported from March to June 2020, the frequency per kit, lot number, specimen type, cycle threshold (Ct) and peak values of the amplification curves, positive target genes, and results of repeated or consecutive tests were analyzed.

Results: A total of 43,268 tests were conducted, of which 93 (0.21%) were inconclusive—49 from 11 coronavirus disease (COVID-19) patients and 44 from non-COVID-19 patients. In COVID-19 patients, the results were inconclusive 11.9 ± 4.7 days after diagnosis and were negative 8.8 ± 5.5 days after the inconclusive results were reported. However, in non-COVID-19 patients, they were all negative upon retest and 81.8% of them were identified to have yielded in 2 out of 8 lots. The most frequently positive target genes were N (55.4%) in COVID-19 and RdRp (61.2%) in non-COVID-19 patients, respectively. No difference was observed in the Ct or peak values of the amplification curves for inconclusive samples between COVID-19 and non-COVID-19 cases.

Conclusion: Inconclusive test results should be reported neither positive nor negative. Such results can be reported as inconclusive without retesting in COVID-19 patients; however, they should certainly be confirmed by a retest in non-COVID-19 patients or newly diagnosed cases.

Figures & Tables

Fig. 1. Classification of the inconclusive results according to COVID-19 diagnosis and retest or consecutive test results.