Abstract
Strategies for monitoring the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are crucial for combating the pandemic. Detection and mutation surveillance of SARS-CoV-2 and other respiratory viruses require separate and complex workflows that rely on highly specialized facilities, personnel, and reagents. To date, no method can rapidly diagnose multiple viral infections and determine variants in a high-throughput manner.
We describe a method for multiplex isothermal amplification-based sequencing and real-time analysis of multiple viral genomes, termed nanopore sequencing of isothermal rapid viral amplification for near real-time analysis (NIRVANA). It can simultaneously detect SARS-CoV-2, influenza A, human adenovirus, and human coronavirus and monitor mutations for up to 96 samples in real time.
NIRVANA showed high sensitivity and specificity for SARS-CoV-2 in 70 clinical samples with a detection limit of 20 viral RNA copies per μL of extracted nucleic acid. It also detected the influenza A co-infection in two samples. The variant analysis results of SARS-CoV-2-positive samples mirror the epidemiology of coronavirus disease 2019 (COVID-19). Additionally, NIRVANA could simultaneously detect SARS-CoV-2 and pepper mild mottle virus (PMMoV) (an omnipresent virus and water-quality indicator) in municipal wastewater samples.
NIRVANA provides high-confidence detection of both SARS-CoV-2 and other respiratory viruses and mutation surveillance of SARS-CoV-2 on the fly. We expect it to offer a promising solution for rapid field-deployable detection and mutational surveillance of pandemic viruses.
M.L. is supported by KAUST Office of Sponsored Research (BAS/1/1080-01). This work is supported by KAUST Competitive Research Grant (URF/1/3412-01-01; M.L. and J.C.I.B.) and Universidad Catolica San Antonio de Murcia (J.C.I.B.). A.M.H. is supported by Saudi Ministry of Education (project 436).
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Multiplex isothermal amplification of viral genomes followed by Nanopore sequencingSimultaneous detection of SARS-CoV-2 and co-infecting viruses in up to 96 patientsSimultaneous detection of SARS-CoV-2 and pepper mild mottle virus in wastewaterReal-time and field-deployable surveillance of SARS-CoV-2 variants
It is challenging to simultaneously identify severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and detect viral mutations. Bi et al. develop a convenient sequencing-based method that can simultaneously detect SARS-CoV-2, influenza A (FluA), human adenovirus, and human coronavirus and monitor viral mutations for up to 96 samples in real time. The method not only showed high sensitivity and specificity for SARS-CoV-2 in 70 clinical samples but also detected cases of FluA co-infection. Additionally, it could simultaneously detect SARS-CoV-2 and other viruses in municipal wastewater samples. This rapid and field-deployable method promises to help contain the spread of coronavirus disease 2019 (COVID-19) and ensure the effectiveness of current vaccines.
Bi et al. develop a convenient sequencing-based method that can simultaneously detect SARS-CoV-2, influenza A, human adenovirus, and human coronavirus and monitor viral mutations for up to 96 samples in real time. This rapid and field-deployable method promises to help contain the spread of COVID-19 and ensure the effectiveness of vaccines.