Extraction-free RT-LAMP to detect SARS-CoV-2 is less sensitive but highly specific compared to standard RT-PCR in 101 samples

• Published in: Journal of clinical virology 2021-03, Vol.136, p.104764-104764, Article 104764
• Main Authors: Schellenberg, John J., Ormond, Margaret, Keynan, Yoav
• Format: Article
• Language: English
• Subjects: Asymptomatic Diseases; Canada; COVID-19 - diagnosis; COVID-19 Testing - methods; Extraction-free; Humans; Instrument-free; Isothermal amplification; Mass Screening - methods; Molecular Diagnostic Techniques - methods; Nasopharynx - virology; Nucleic Acid Amplification Techniques - methods; Point-of-Care Testing; Reverse Transcriptase Polymerase Chain Reaction - methods; RNA, Viral - analysis; RT-LAMP; SARS-CoV-2; SARS-CoV-2 - isolation & purification; Sensitivity and Specificity
• ISSN: 1386-6532; 1873-5967
• DOI: 10.1016/j.jcv.2021.104764

•Extraction-free RT-LAMP (isothermal amplification) offers specific advantages over other rapid testing options for SARS-CoV-2, such as Cepheid’s GeneXpert, Abbott’s ID NOW platform and Panbio Rapid Antigen cassettes.•Relative to RT-PCR in 101 samples, extract-free RT-LAMP had a specificity of 100% and sensitivity of 77%, with all strong positives (Ct < 24) detected, indicating that the most infectious individuals would be correctly identified with this assay.•Using a positive control at a known concentration, the limit of detection for RT-LAMP was determined to be 50,000 copies/mL, comparable to other studies and consistent with concentrations at which SARS-CoV-2 can be cultured, an indicator of transmissibility.•RT-LAMP could be used for faster identification of strong positives in prioritized samples at the point-of-care, or for repeated testing in workplaces, particularly if less invasive samples such as saliva are collected. The current scale of public and private testing cannot be expected to meet the emerging need for higher levels of community-level and repeated screening of asymptomatic Canadians for SARS-CoV-2. Rapid point-of-care techniques are increasingly being offered to fill the gap in screening levels required to identify undiagnosed individuals with high viral loads. However, rapid, point-of-care tests often have lower sensitivity in practice. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) for SARS-CoV-2 has proven sensitive and specific and provides visual results in minutes. Using a commercially available kit for RT-LAMP and primer set targetting nucleocapsid (N), we tested a blinded set of 101 archived nasopharyngeal (NP) swab samples with known RT-PCR results. RT-LAMP reactions were incubated at 65 °C for 30 min, using heat-inactivated nasopharyngeal swab sample in viral transport medium, diluted tenfold in water, as input. RT-LAMP agreed with all RT-PCR defined negatives (N = 51), and all positives with cycle threshold (Ct) less than 20 (N = 24), 65% of positives with Ct between 20−30 (N = 17), and no positives with Ct greater than 30 (N = 9). RT-LAMP requires fewer and different core components, so may not compete directly with the mainline testing workflow, preserving precious central laboratory resources for those with the greatest need. Careful messaging must be provided when using less-sensitive tests, so that people are not falsely reassured by negative results, but this caveat must be weighed against the