Comparison of five polyethylene glycol precipitation procedures for the RT-qPCR based recovery of murine hepatitis virus, bacteriophage phi6, and pepper mild mottle virus as a surrogate for SARS-CoV-2 from wastewater

Shotaro Torii, Wakana Oishi, Yifan Zhu, Ocean Thakali, Bikash Malla, Zaizhi Yu, Bo Zhao, Chisato Arakawa, Masaaki Kitajima, Akihiko Hata, Masaru Ihara, Shigeru Kyuwa, Daisuke Sano, Eiji Haramoto, Hiroyuki Katayama

Research output: Contribution to journalArticlepeer-review

Abstract

Polyethylene glycol (PEG) precipitation is one of the conventional methods for virus concentration. This technique has been used to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in wastewater. The procedures and seeded surrogate viruses were different among implementers; thus, the reported whole process recovery efficiencies considerably varied among studies. The present study compared five PEG precipitation procedures, with different operational parameters, for the RT-qPCR-based whole process recovery efficiency of murine hepatitis virus (MHV), bacteriophage phi6, and pepper mild mottle virus (PMMoV), and molecular process recovery efficiency of murine norovirus using 34 raw wastewater samples collected in Japan. The five procedures yielded significantly different whole process recovery efficiency of MHV (0.070%–2.6%) and phi6 (0.071%–0.51%). The observed concentration of indigenous PMMoV ranged from 8.9 to 9.7 log (8.2 × 108 to 5.6 × 109) copies/L. Interestingly, PEG precipitation with 2-h incubation outperformed that with overnight incubation partially due to the difference in molecular process recovery efficiency. The recovery load of MHV exhibited a positive correlation (r = 0.70) with that of PMMoV, suggesting that PMMoV is the potential indicator of the recovery efficiency of SARS-CoV-2. In addition, we reviewed 13 published studies and found considerable variability between different studies in the whole process recovery efficiency of enveloped viruses by PEG precipitation. This was due to the differences in operational parameters and surrogate viruses as well as the differences in wastewater quality and bias in the measurement of the seeded load of surrogate viruses, resulting from the use of different analytes and RNA extraction methods. Overall, the operational parameters (e.g., incubation time and pretreatment) should be optimized for PEG precipitation. Co-quantification of PMMoV may allow for the normalization of SARS-CoV-2 RNA concentration by correcting for the differences in whole process recovery efficiency and fecal load among samples.

Original languageEnglish
Article number150722
JournalScience of the Total Environment
Volume807
DOIs
Publication statusPublished - 2022 Feb 10

Keywords

  • Polyethylene glycol precipitation
  • SARS-CoV-2
  • Surrogates
  • Virus concentration
  • Wastewater-based epidemiology

ASJC Scopus subject areas

  • Environmental Engineering
  • Environmental Chemistry
  • Waste Management and Disposal
  • Pollution

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