Surveillance of SARS-CoV-2 RNA in wastewater: Methods optimization and quality control are crucial for generating reliable public health information

Abstract

Monitoring for SARS-CoV-2 RNA in wastewater through the process of wastewater-based epidemiology (WBE) provides an additional surveillance tool, contributing to community-based screening and prevention efforts as these measurements have preceded disease cases in some instances. Numerous detections of SARS-CoV-2 RNA have been reported globally using various methods, demonstrating the technical feasibility of routine monitoring. However, in order to reliably interpret data produced from these efforts for informing public health interventions, additional quality control information and standardization in sampling design, sample processing, and data interpretation and reporting is needed. This review summarizes published studies of SARS-CoV-2 RNA detection in wastewater as well as available information regarding concentration, extraction, and detection methods. The review highlights areas for potential standardization including considerations related to sampling timing and frequency relative to peak fecal loading times; inclusion of appropriate information on sample volume collected; sample collection points; transport and storage conditions; sample concentration and processing; RNA extraction process and performance; effective volumes; PCR inhibition; process controls throughout sample collection and processing; PCR standard curve performance; and recovery efficiency testing. Researchers are recommended to follow the Minimum Information for Publication of Quantitative Real-Time PCR (MIQE) guidelines. Adhering to these recommendations will enable robust interpretation of wastewater monitoring results and improved inferences regarding the relationship between monitoring results and disease cases.

Concentration and Quantification of SARS-CoV-2 RNA in Wastewater Using Polyethylene Glycol-Based Concentration and qRT-PCR

“…Prior to analysis, RNA viruses in wastewater are typically concentrated, followed by the extraction and reverse transcription (RT) of RNA and PCR-based quantification, which enables the accurate and sensitive detection of viruses at genus/species/genogroup/genotype/strain level. To date, methods, including ultrafiltration, ultracentrifugation, filtration-adsorption, flocculation and polyethylene glycol (PEG) precipitation, commonly used for the detection of enteric viruses have been implemented for the detection of SARS-CoV-2 [ 7 , 8 ]. However, in most cases the efficiency of these methods for recovering and quantifying enveloped virus concentrations have not been assessed.…”

Comparison of approaches to quantify SARS-CoV-2 in wastewater using RT-qPCR: Results and implications from a collaborative inter-laboratory study in Canada

“…Standardizing analytical methods themselves is currently not practical for the diverse range of laboratories seeking to respond to the growing need for analytical services. However, doing so for QA/QC is achievable and has been shown to be effective ( Ahmed et al., 2020c ). Indeed, discrepancies between initial values reported by laboratories arose due to differences of standards and protocols used to generate standard curves for quantification from Ct values.…”

Comparing analytical methods to detect SARS-CoV-2 in wastewater

“…Additionally, WBE has the potential to be applied in high-risk settings such as nursing homes and hospital or in lowresource settings (WHO, 2020). As recently stated by WHO, WBE research should be seen as an important public health objective to advance knowledge about COVID-19, however, many technical issues still need to be addressed (Ahmed et al, 2020b;Polo et al, 2020;Rusiñol et al, 2020;WHO, 2020). In an attempt to coordinate current knowledge and data gaps, the European Commission (EC) created a Pan-European Umbrella Study to better understand the limitations and challenges of this approach including the development of a roadmap for a systemic rollout of complementing ongoing national and regional surveillances in a unique approach (EC, 2020).…”