Capsid integrity RT-qPCR for the selective detection of intact SARS-CoV-2 in wastewater

被引：27

作者：

Canh, Vu Duc ^{[1
]}

Torii, Shotaro ^{[1
]}

Yasui, Midori ^{[1
]}

Kyuwa, Shigeru ^{[2
]}

Katayama, Hiroyuki ^{[1
]}

机构：

[1] Univ Tokyo, Grad Sch Engn, Bunkyo Ku, 7-3-1 Hongo, Tokyo 1138656, Japan

[2] Univ Tokyo, Grad Sch Agr & Life Sci, Bunkyo Ku, 7-3-1 Hongo, Tokyo 1138657, Japan

来源：

SCIENCE OF THE TOTAL ENVIRONMENT | 2021年 / 791卷

关键词：

Capsid integrity RT-qPCR; SARS-CoV-2; intact virus; concentration method; wastewater; PROPIDIUM MONOAZIDE; ETHIDIUM MONOAZIDE; INACTIVATION; NOROVIRUS; PRETREATMENT; VIRUSES; HEAT;

D O I：

10.1016/j.scitotenv.2021.148342

中图分类号：

X [环境科学、安全科学];

学科分类号：

08 ; 0830 ;

摘要：

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes have been detected in wastewater worldwide. However, the assessment of SARS-CoV-2 infectivity in wastewater has been limited due to the stringent requirements of biosafety level 3. The main objective of this study is to investigate the applicability of capsid integrity RT-qPCR for the selective detection of intact SARS-CoV-2 in wastewater. Three capsid integrity reagents, namely ethidium monoazide (EMA, 0.1-100 mu M), propidium monoazide (PMA, 0.1-100 mu M), and cisdichlorodiammineplatinum (CDDP, 0.1-1000 mu M), were tested for their effects on different forms (including free genomes, intact and heat-inactivated) of murine hepatitis virus (MHV), which was used as a surrogate for SARS-CoV-2. CDDP at a concentration of 100 mu M was identified as the most efficient reagent for the selective detection of infectious MHV by RT-qPCR (CDDP-RT-qPCR). Next, two common virus concentration methods including ultrafiltration (UF) and polyethylene glycol (PEG) precipitation were investigated for their compatibility with capsid integrity RT-qPCR. The UF method was more suitable than the PEG method since it recovered intact MHV (mean +/- SD, 38% +/- 29%) in wastewater much better than the PEG method did (0.013% +/- 0.015%). Finally, CDDPRT-qPCR was compared with RT-qPCR alone for the detection of SARS-CoV-2 in 16 raw wastewater samples collected in the Greater Tokyo Area. Five samples were positive for SARS-CoV-2 when evaluated by RT-qPCR alone. However, intact SARS-CoV-2 was detected in only three positive samples when determined by CDDP-RT-qPCR. Although CDDP-RT-qPCR was unable to determine the infectivity of SARS-CoV-2 in wastewater, this method could improve the interpretation of positive results of SARS-CoV-2 obtained by RT-qPCR. (c) 2021 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http:// creativecommons.org/licenses/by/4.0/).

引用

页数：10

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