Prediction of SARS-CoV-2 Variant Lineages Using the S1-Encoding Region Sequence Obtained by PacBio Single-Molecule Real-Time Sequencing.

Sébastien Lhomme,Gérald Salin,Pauline Trémeaux,Nicolas Jeanne,Justine Latour,Noémie Ranger,Jacques Izopet,Marion Migueres,Cécile Donnadieu

doi:10.3390/v13122544

Abstract

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is the causal agent of the COVID-19 pandemic that emerged in late 2019. The outbreak of variants with mutations in the region encoding the spike protein S1 sub-unit that can make them more resistant to neutralizing or monoclonal antibodies is the main point of the current monitoring. This study examines the feasibility of predicting the variant lineage and monitoring the appearance of reported mutations by sequencing only the region encoding the S1 domain by Pacific Bioscience Single Molecule Real-Time sequencing (PacBio SMRT). Using the PacBio SMRT system, we successfully sequenced 186 of the 200 samples previously sequenced with the Illumina COVIDSeq (whole genome) system. PacBio SMRT detected mutations in the S1 domain that were missed by the COVIDseq system in 27/186 samples (14.5%), due to amplification failure. These missing positions included mutations that are decisive for lineage assignation, such as G142D (n = 11), N501Y (n = 6), or E484K (n = 2). The lineage of 172/186 (92.5%) samples was accurately determined by analyzing the region encoding the S1 domain with a pipeline that uses key positions in S1. Thus, the PacBio SMRT protocol is appropriate for determining virus lineages and detecting key mutations.

Highlights

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a member of the Coronaviridae family, that caused the COVID-19 pandemic is a ~30 kb single-stranded, positive-sense RNA virus [1,2]
Clades were identified by NextClade and lineages were determined using the PANGOLIN classification based on the analysis of full-length genome sequences (Table 2)
In January 2021, three strains were more frequently detected: the Alpha variant that belonged to the clade 20I and the lineage B.1.1.7 (48/146, 32.9%), followed by 37 (25.3%) strains belonging to the clade 20A and the lineage B.1.160 and 37 variants belonging to the clade 20E(EU1) and the lineage B.1.177

Summary

Introduction

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a member of the Coronaviridae family, that caused the COVID-19 pandemic is a ~30 kb single-stranded, positive-sense RNA virus [1,2]. The European lineage B.1.177 (clade 20E(EU1)) differs from ancestral sequences at 6 or more positions, including a A222V mutation in the spike protein This variant arose in Spain early in the summer of 2020 and subsequently spread across Europe, perhaps carried by infected holiday travelers [10]. Another variant belonging to lineage B.1.160 (clade 20A, previously identified 20A.EU2) that has a S477N substitution in the spike protein was common in Autumn 2020 and early 2021 in some European countries, including France [10]. Many B.1.617.2 sublineages were described ( called AY lineages, for which AY. corresponds to B.1.617.2.1) and their number is currently increasing

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