Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan

Wastewater-based epidemiology is expected to be able to identify SARS-CoV-2 variants at an early stage via next-generation sequencing. In the present study, we developed a highly sensitive amplicon sequencing method targeting the spike gene of SARS-CoV-2, which allows for sequencing viral genomes fr...

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Autores principales: Iwamoto, Ryo, Yamaguchi, Kiyoshi, Katayama, Kotoe, Ando, Hiroki, Setsukinai, Ken-ichi, Kobayashi, Hiroyuki, Okabe, Satoshi, Imoto, Seiya, Kitajima, Masaaki
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Authors. Published by Elsevier B.V. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10129341/
https://www.ncbi.nlm.nih.gov/pubmed/37105480
http://dx.doi.org/10.1016/j.scitotenv.2023.163706
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author Iwamoto, Ryo
Yamaguchi, Kiyoshi
Katayama, Kotoe
Ando, Hiroki
Setsukinai, Ken-ichi
Kobayashi, Hiroyuki
Okabe, Satoshi
Imoto, Seiya
Kitajima, Masaaki
author_facet Iwamoto, Ryo
Yamaguchi, Kiyoshi
Katayama, Kotoe
Ando, Hiroki
Setsukinai, Ken-ichi
Kobayashi, Hiroyuki
Okabe, Satoshi
Imoto, Seiya
Kitajima, Masaaki
author_sort Iwamoto, Ryo
collection PubMed
description Wastewater-based epidemiology is expected to be able to identify SARS-CoV-2 variants at an early stage via next-generation sequencing. In the present study, we developed a highly sensitive amplicon sequencing method targeting the spike gene of SARS-CoV-2, which allows for sequencing viral genomes from wastewater containing a low amount of virus. Primers were designed to amplify a relatively long region (599 bp) around the receptor-binding domain in the SARS-CoV-2 spike gene, which could distinguish initial major variants of concern. To validate the methodology, we retrospectively analyzed wastewater samples collected from a septic tank installed in a COVID-19 quarantine facility between October and December 2020. The relative abundance of D614G mutant in SARS-CoV-2 genomes in the facility wastewater increased from 47.5 % to 83.1 % during the study period. The N501Y mutant, which is the characteristic mutation of the Alpha-like strain, was detected from wastewater collected on December 24, 2020, which agreed with the fact that a patient infected with the Alpha-like strain was quarantined in the facility on this date. We then analyzed archived municipal wastewater samples collected between November 2020 and January 2021 that contained low SARS-CoV-2 concentrations ranging from 0.23 to 0.43 copies/qPCR reaction (corresponding to 3.30 to 4.15 log(10) copies/L). The targeted amplicon sequencing revealed that the Alpha-like variant with D614G and N501Y mutations was present in municipal wastewater collected on December 4, 2020 and later, suggesting that the variant had already spread in the community before its first clinical confirmation in Japan on December 25, 2020. These results demonstrate that targeted amplicon sequencing of wastewater samples is a powerful surveillance tool applicable to low COVID-19 prevalence periods and may contribute to the early detection of emerging variants.
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spelling pubmed-101293412023-04-26 Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan Iwamoto, Ryo Yamaguchi, Kiyoshi Katayama, Kotoe Ando, Hiroki Setsukinai, Ken-ichi Kobayashi, Hiroyuki Okabe, Satoshi Imoto, Seiya Kitajima, Masaaki Sci Total Environ Article Wastewater-based epidemiology is expected to be able to identify SARS-CoV-2 variants at an early stage via next-generation sequencing. In the present study, we developed a highly sensitive amplicon sequencing method targeting the spike gene of SARS-CoV-2, which allows for sequencing viral genomes from wastewater containing a low amount of virus. Primers were designed to amplify a relatively long region (599 bp) around the receptor-binding domain in the SARS-CoV-2 spike gene, which could distinguish initial major variants of concern. To validate the methodology, we retrospectively analyzed wastewater samples collected from a septic tank installed in a COVID-19 quarantine facility between October and December 2020. The relative abundance of D614G mutant in SARS-CoV-2 genomes in the facility wastewater increased from 47.5 % to 83.1 % during the study period. The N501Y mutant, which is the characteristic mutation of the Alpha-like strain, was detected from wastewater collected on December 24, 2020, which agreed with the fact that a patient infected with the Alpha-like strain was quarantined in the facility on this date. We then analyzed archived municipal wastewater samples collected between November 2020 and January 2021 that contained low SARS-CoV-2 concentrations ranging from 0.23 to 0.43 copies/qPCR reaction (corresponding to 3.30 to 4.15 log(10) copies/L). The targeted amplicon sequencing revealed that the Alpha-like variant with D614G and N501Y mutations was present in municipal wastewater collected on December 4, 2020 and later, suggesting that the variant had already spread in the community before its first clinical confirmation in Japan on December 25, 2020. These results demonstrate that targeted amplicon sequencing of wastewater samples is a powerful surveillance tool applicable to low COVID-19 prevalence periods and may contribute to the early detection of emerging variants. The Authors. Published by Elsevier B.V. 2023-08-20 2023-04-25 /pmc/articles/PMC10129341/ /pubmed/37105480 http://dx.doi.org/10.1016/j.scitotenv.2023.163706 Text en © 2023 The Authors Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.
spellingShingle Article
Iwamoto, Ryo
Yamaguchi, Kiyoshi
Katayama, Kotoe
Ando, Hiroki
Setsukinai, Ken-ichi
Kobayashi, Hiroyuki
Okabe, Satoshi
Imoto, Seiya
Kitajima, Masaaki
Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan
title Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan
title_full Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan
title_fullStr Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan
title_full_unstemmed Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan
title_short Identification of SARS-CoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19 prevalence period in Japan
title_sort identification of sars-cov-2 variants in wastewater using targeted amplicon sequencing during a low covid-19 prevalence period in japan
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10129341/
https://www.ncbi.nlm.nih.gov/pubmed/37105480
http://dx.doi.org/10.1016/j.scitotenv.2023.163706
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