ABSTRACT
Here we describe a retrospective clinical evaluation of the QIAGEN artus® SARS-CoV-2 Prep&Amp UM RT-PCR assay that detects SARS-CoV-2 RNA without the need for a nucleic acid eluate extraction procedure. Using Roche SARS-CoV-2 RT-PCR on the cobas® 8800 platform as a reference standard, a total of 225 confirmed SARS-CoV-2 positive and 320 negative nasopharyngeal swabs in viral transport media, were used to evaluate the artus® assay. Using the RT-PCR cycle threshold as a semi-quantitative marker of viral load, an assessment of over 370,000 SARS-CoV-2 RT-PCR positive results was used in the design of the reference positive specimen cohort. The viral load of all reference positive specimens used in the evaluation was a unique and accurate representation of the range and levels of SARS-CoV-2 positivity observed over a 13-month period of the COVID-19 pandemic. The artus® RT-PCR detects the presence of SARS-CoV-2 RNA, an internal control, and the human RNase P gene to ensure specimen quality. The diagnostic sensitivity of artus® was 92.89% with a specificity of 100%. To assess the analytical sensitivity, a limit of detection was performed using the 1st WHO NIBSC SARS-CoV-2 international standard, recording a 95% LOD of 1.1 × 103 IU/ml. The total invalid rate of specimens was 7.34% due to a lack of detectable RNase P (Ct >35). The artus® SARS-CoV-2 Prep&Amp UM RT-PCR assay is a new rapid RT-PCR assay, which may be considered to produce acceptable levels of diagnostic sensitivity and specificity whilst potentially halving the laboratory processing time.
ABSTRACT
The COVID-19 pandemic has led to the rapid development of a plethora of molecular diagnostic assays with real-time polymerase chain reaction (RT-PCR) at the forefront. In this review, we will discuss the history and utility of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) molecular diagnostics and the associated current and future regulatory process in Europe. We will assess the performance characteristics of a range of the most common SARS-CoV-2 molecular tests currently used in Europe with a focus on as rapid molecular platforms, stand-alone RT-PCR kits, the role of low-throughput and high-throughput end-to-end testing platforms, and the rapidly evolving field of SARS-CoV-2 variant of concern identification.
Subject(s)
COVID-19 , SARS-CoV-2 , COVID-19/diagnosis , Europe/epidemiology , Humans , Pandemics , Pathology, Molecular , SARS-CoV-2/geneticsABSTRACT
Large scale screening of health care workers and the general population for asymptomatic COVID-19 infection requires modalities that are amenable to testing at scale while retaining acceptable levels of sensitivity and specificity. This study evaluated a novel COVID-19 Direct-RT LAMP assay using saliva samples in asymptomatic individuals by comparison to RT-PCR. Additional studies were performed using VTM collected from routine diagnostic testing. Analytical sensitivity was determined for Direct RT-LAMP assay using the WHO International Standard. Finally, quantified results from RT-PCR testing of 9177 nose and throat swabs obtained from routine diagnostic testing were used to estimate the sensitivity of Direct RT-LAMP using the limit of detection curve obtained from the analytical sensitivity data. Results from saliva testing demonstrated a sensitivity of 40.91% and a specificity of 100% for Direct RT-LAMP. The sensitivity and specificity for nose and throat swabs were 44.85% and 100% respectively. The 95% limit of detection (LOD) for Direct RT-LAMP was log 7.13 IU/ml (95% 6.9-7.5). The estimated sensitivity for Direct-RT LAMP based on the results of 9117 nose and throat swabs was 34% and 45% for saliva and VTM respectively. The overall diagnostic sensitivity of Direct RT-LAMP was low compared to RT-PCR. Testing of nose and throat swabs and estimating the sensitivity based on a large cohort of clinical samples demonstrated similar results. This study highlights the importance of utilising the prospective collection of samples from the intended target population in the assessment of diagnostic sensitivity.