Protocol for Safe, Affordable, and Reproducible 1 Isolation and Quantitation 2 of SARS-CoV-2 RNA from Wastewater

Authors

Monica Trujillo, CUNY Queensborough Community CollegeFollow
Kristen Cheung, CUNY Queens College
Anna Gao, CUNY Queens College
Irene Hoxie, CUNY Graduate Center
Sherin Kannoly, CUNY Queens College
Nanami Kubota, CUNY Queens College
Kaung Myat San, CUNY Queens College
Davida S. Smyth, The New School
John J. Dennehy, CUNY Queens CollegeFollow

Document Type

Article

Publication Date

2021

Abstract

The following protocol describes our workflow for processing wastewater with the goal of detecting the genetic signal of SARS-CoV-2. The steps include pasteurization, virus concentration, RNA extraction, and quantification by RT-qPCR. We include auxiliary steps that provide new users with tools and strategies that will help troubleshoot key steps in the process. This protocol is one of the safest, cheapest, and most reproducible approaches for the detection of SARS-CoV-2 RNA in wastewater. Furthermore, the RNA obtained using this protocol, minus the pasteurization step, can be sequenced both using a targeted approach sequencing specific regions or the whole genome. The protocol was adopted by the New York City Department of Environmental Protection in August 2020 to support their efforts in monitoring SARS-CoV-2 prevalence in wastewater in all five boroughs of the city. Owing to a pasteurization step, it is safe for use in a BSL1+ facility. This step also increases the genetic signal of the virus while making the protocol safe for the personnel involved. This protocol could be used to isolate a variety of other clinically relevant viruses from wastewater and serve as a foundation of a wastewater surveillance strategy for monitoring community spread of known and emerging viral pathogens.

Comments

This submitted manuscript is also available in medRxiv. https://doi.org/10.1101/2021.02.16.21251787

This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.