Setting-Up a Rapid SARS-CoV-2 Genome Assessment by Next-Generation Sequencing in an Academic Hospital Center (LPCE, Louis Pasteur Hospital, Nice, France).

Patrick Brest,Sylvie Leroy,Maryline Allégra ,Olivier Bordone,Virginie Lespinet‐Fabre ,Baharia Mograbi,Charles‐Hugo Marquette ,Virginie Tanga,Elisabeth Lantéri ,Myriam Salah,Simon Heeke,Paul Hofman,Marius Ilié,Charlotte Maniel,Emmanuel Chamorey,Jacques Boutros,Jonathan Benzaquen,Véronique Hofman,Renaud Schiappa,Julien Fayada

doi:10.3389/fmed.2021.730577

Abstract

Introduction: Aside from the reverse transcription-PCR tests for the diagnosis of the COVID-19 in routine clinical care and population-scale screening, there is an urgent need to increase the number and the efficiency for full viral genome sequencing to detect the variants of SARS-CoV-2. SARS-CoV-2 variants assessment should be easily, rapidly, and routinely available in any academic hospital.Materials and Methods: SARS-CoV-2 full genome sequencing was performed retrospectively in a single laboratory (LPCE, Louis Pasteur Hospital, Nice, France) in 103 SARS-CoV-2 positive individuals. An automated workflow used the Ion Ampliseq SARS-CoV-2 panel on the Genexus Sequencer. The analyses were made from nasopharyngeal swab (NSP) (n = 64) and/or saliva (n = 39) samples. All samples were collected in the metropolitan area of the Nice city (France) from September 2020 to March 2021.Results: The mean turnaround time between RNA extraction and result reports was 30 h for each run of 15 samples. A strong correlation was noted for the results obtained between NSP and saliva paired samples, regardless of low viral load and high (>28) Ct values. After repeated sequencing runs, complete failure of obtaining a valid sequencing result was observed in 4% of samples. Besides the European strain (B.1.160), various variants were identified, including one variant of concern (B.1.1.7), and different variants under monitoring.Discussion: Our data highlight the current feasibility of developing the SARS-CoV-2 next-generation sequencing approach in a single hospital center. Moreover, these data showed that using the Ion Ampliseq SARS-CoV-2 Assay, the SARS-CoV-2 genome sequencing is rapid and efficient not only in NSP but also in saliva samples with a low viral load. The advantages and limitations of this setup are discussed.

Highlights

Aside from the reverse transcription-PCR tests for the diagnosis of the COVID-19 in routine clinical care and population-scale screening, there is an urgent need to increase the number and the efficiency for full viral genome sequencing to detect the variants of SARS-CoV-2
Sequencing performed from 16 matched salivary and nasopharyngeal swabs (NSPs) samples collected at the same time, detected identical variants in 14/16 samples, whereas 2 saliva samples failed to give a valid sequencing run. These preliminary results demonstrated the feasibility of the Ion Ampliseq SARS-CoV-2 Assay to identify known SARS-CoV-2 variants in a daily practice of an academic hospital laboratory
The flow of sequencing is quite limited compared to the flow of screening using the reverse transcription (RT)-PCR method, since only a maximum of 15 samples can be run each day if a fully automated process is selected (Supplementary Table 1)

Summary

Introduction

Aside from the reverse transcription-PCR tests for the diagnosis of the COVID-19 in routine clinical care and population-scale screening, there is an urgent need to increase the number and the efficiency for full viral genome sequencing to detect the variants of SARS-CoV-2. Since March 2020, multiple SARS-CoV-2 variants emerged with a large number of mutations, notably in the S protein [3,4,5,6,7] These have been detected in different geographical regions being responsible for the continuous COVID-19 dissemination [8,9,10,11,12]. The control of the COVID-19 pandemic requires tremendous efforts to allow widespread screening for SARS-CoV-2 in the general population and to enable worldwide vaccinations These actions must include the possibility to rapidly and massively detect the known viral variants SARS-CoV-2 and quickly define the emergence of novel variants that arise throughout the pandemic [13]. Variants under monitoring (VUM) are currently not associated with virological, epidemiological, or clinical data associated with their potential risk to human health (https://www.who.int/csr/don/31december-2020-sars-cov2-variants/en/, accessed May 6, 2021)

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Results

Conclusion