Application of a high-throughput quantitative PCR system for simultaneous monitoring of SARS-CoV-2 variants and other pathogenic viruses in wastewater

Malla, Bikash, Thakali, Ocean, Shrestha, Sadhana, Segawa, Takahiro, Kitajima, Masaaki and Haramoto, Eiji, (2022). Application of a high-throughput quantitative PCR system for simultaneous monitoring of SARS-CoV-2 variants and other pathogenic viruses in wastewater. Science of the Total Environment, 853 158659-158667

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    Author Malla, Bikash
    Thakali, Ocean
    Shrestha, Sadhana
    Segawa, Takahiro
    Kitajima, Masaaki
    Haramoto, Eiji
    Title Application of a high-throughput quantitative PCR system for simultaneous monitoring of SARS-CoV-2 variants and other pathogenic viruses in wastewater
    Appearing in Science of the Total Environment
    Volume 853
    Publication Date 2022-09-09
    Place of Publication Amsterdam
    Publisher Elsevier B.V.
    Start page 158659
    End page 158667
    Language eng
    Abstract Variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are continuously emerging, highlighting the importance of regular surveillance of SARS-CoV-2 and other epidemiologically significant pathogenic viruses in the current context. Reverse transcription-quantitative PCR (RT-qPCR) is expensive, time-consuming, labor-intensive, requires a large reagent volume, and only tests a few targets in a single run. High-throughput qPCR (HT-qPCR) utilizing the Biomark HD system (Fluidigm) can be used as an alternative. This study applied an HT-qPCR to simultaneously detect SARS-CoV-2, SARS-CoV-2 nucleotide substituted RNA, and other pathogenic viruses in wastewater. Wastewater samples were collected from the coronavirus disease 2019 (COVID-19) quarantine facility between October 2020 and February 2021 (n = 4) and from the combined and separated sewer lines of a wastewater treatment plant (WWTP) in Yokkaichi, Mie Prefecture, Japan, between March and August 2021 (n = 23 each). The samples were analyzed by HT-qPCR using five SARS-CoV-2, nine SARS-CoV-2 spike gene nucleotide substitution-specific, five pathogenic viruses, and three process control assays. All samples from the quarantine facility tested positive for SARS-CoV-2 and the nucleotide substitutions N501Y and S69-70 del (Alpha variant) were detected in the December 2020 sample, coinciding with the first clinical case in Japan. Only three WWTP samples were positive when tested with a single SARS-CoV-2 assay, whereas more than eight samples were positive when tested with all assays, indicating that using multiple assays increases the likelihood of detection. The nucleotide substitution L452R (Delta variant) was detected in the WWTP samples of Mie Prefecture in April 2021, but the detection of Delta variant from patients had not been reported until May 2021. Aichi virus 1 and norovirus GII were prevalent in WWTP samples. This study demonstrated that HT-qPCR may be the most time- and cost-efficient method for tracking COVID-19 and broadly monitoring community health.
    Keyword High-throughput
    qPCR
    Pathogenic viruses
    SARS-CoV-2
    Wastewater
    Copyright Holder The Authors
    Copyright Year 2022
    Copyright type Creative commons
    DOI https://doi.org/10.1016/j.scitotenv.2022.158659
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    Created: Fri, 23 Sep 2022, 15:16:59 JST by Hanna Takemoto on behalf of UNU IAS