Evaluation of Loop-mediated isothermal amplification (LAMP) method as an effective molecular point-of-care technique for the rapid diagnosis of SARS-CoV-2 infection

ABSTRACT

Background: The novel Coronavirus disease-2019 (Covid-19) pandemic emergency is a concrete example of the existing gap between availability of advanced diagnostics and need for cost-effective methodology. The current standard method for Coronavirus detection is the reverse transcription-PCR (RT-PCR), but the recent validation of a new rapid SARS-CoV-2 RT-LAMP assay offers an alternative diagnostic pathway. Unlike PCR tests, LAMP (loop-mediated isothermal amplification) do not require sequential changes of temperature and so can turnaround test results more rapidly. We explored the diagnostic effectiveness of LAMP compared with RTqPCR traditional assay.Methods: A sample of 1652 UTM NP swabs (collected from suspected or non suspected patient of ED) were tested for SARS Cov2 infection, using IC GENE SARS-CoV-2 POC (Enbiotech) that detect two specific viral targets S gene and N gene. After a rapid termal RNA extraction protocol, a Real Time amplifier and fluorescence reader allowed us to achieve the result (simultaneously, and up to 12 samples per run) in a time between 30 and 60 minutes. The same samples were further analyzed with a RT-qPCR traditional assay (Cephied Xpert Xpress® SARS cov-2 and Altona RealStar® SARS-cov-2 RT-PCR).Results: The technical performance of assay demonstrated a sensitivity of 72.2 % (95%CI 61.4/80.8) and specificity of 86.8 % (95%CI 85.0/88.4), VPP 21.5%, VPN 98.4%, in comparison to current standard of care RT-qPCR testing after RNA extraction, across all samples tested (CT <45 by RTqPCR), increasing to a sensitivity of 96.6 % for those samples with a higher viral load (CT <25 by RTqPCR). Our results shows that main limitation of LAMP is the high number of false positive samples (80% of all positive test). Conclusions: in our experience the ICGene RNA RT-LAMP kit only reliably detects very strong positives patients (Ct<25), however, statistically, these are the most infectious cases and so the most urgent to find quickly, particularly in vulnerable settings. Whereby RNA RT-LAMP could replace rRT-PCR where there is need to rapidly identify highly contagious individuals within emergency departments, ensuring results still get laboratory confirmation with highly sensitive nucleic acid amplification testing (NAAT).