Multiplexed detection of SARS-CoV-2 and other respiratory infections in high throughput by SARSeq
Ramesh Yelagandula, Aleksandr Bykov, Alexander Vogt, Robert Heinen, Ezgi Özkan, Marcus Strobl, Juliane Christina Baar, Kristina Uzunova, Bence Hajdusits, Darja Kordic, Erna Suljic, Amina Kurtović-Kozarić, Sebija Izetbegović, Justine Schaeffer, Peter Hufnagl, Alexander Zoufaly, Tamara Seitz, VCDI, Mariam Al-Rawi, Stefan L. Ameres, Juliane Christina Baar, Benedikt Bauer, Nikolaus Beer, Katharina Bergauer, Wolfgang H. Binder, Claudia Blaukopf, Boril Bochev, Julius Brennecke, Selina Brinnich, Aleksandra Bundalo, Meinrad Busslinger, Tim Clausen, Geert de Vries, Marcus P. S. Dekens, David Drechsel, Zuzana Džupinková, Michaela Eckmann-Mader, Michaela Fellner, Thomas Fellner, Laura Fin, Bianca Valeria Gapp, Gerlinde Grabmann, Irina Grishkovskaya, Astrid Hagelkrüys, Dominik Handler, David Haselbach, Louisa Hempel, Louisa Hill, David Hoffmann, Stefanie Horer, Harald Isemann, Robert Kalis, Max J. Kellner, Juliane Kley, Thomas Köcher, Alwin Köhler, Christian Krauditsch, Sabina Kula, Sonja Lang, Richard Latham, Marie-Christin Leitner, Thomas Leonard, Dominik Lindenhofer, Raphael A. Manzenreither, Martin Mátl, Karl Mechtler, Anton Meinhart, Stefan Mereiter, Thomas Micheler, Paul Moeseneder, Tobias Neumann, Simon Nimpf, Magnus Nordborg, Egon Ogris, Michaela Pagani, Andrea Pauli, Jan‐Michael Peters, Petra Pjevac, Clemens Plaschka, Martina Rath, Daniel Reumann, Sarah Rieser, Marianne Rocha-Hasler, Alan Rodriguez, Nathalie Ropek, James J. Ross, Harald Scheuch, Karina Schindler, Clara Schmidt, Hannes Schmidt, Jakob Schnabl, Stefan Schüchner, Tanja A. Schwickert, Andreas Sommer, Daniele Soldoroni, Johannes Stadlmann, Peter Steinlein, Marcus Strobl, Simon Strobl, Qiong Sun
Abstract
The COVID-19 pandemic has demonstrated the need for massively-parallel, cost-effective tests monitoring viral spread. Here we present SARSeq, saliva analysis by RNA sequencing, a method to detect SARS-CoV-2 and other respiratory viruses on tens of thousands of samples in parallel. SARSeq relies on next generation sequencing of multiple amplicons generated in a multiplexed RT-PCR reaction. Two-dimensional, unique dual indexing, using four indices per sample, enables unambiguous and scalable assignment of reads to individual samples. We calibrate SARSeq on SARS-CoV-2 synthetic RNA, virions, and hundreds of human samples of various types. Robustness and sensitivity were virtually identical to quantitative RT-PCR. Double-blinded benchmarking to gold standard quantitative-RT-PCR performed by human diagnostics laboratories confirms this high sensitivity. SARSeq can be used to detect Influenza A and B viruses and human rhinovirus in parallel, and can be expanded for detection of other pathogens. Thus, SARSeq is ideally suited for differential diagnostic of infections during a pandemic.