Wastewater-based epidemiology (WBE) is useful for the surveillance of severe acute
respiratory syndrome coronavirus 2 (SARS-CoV-2) in communities, complementing clinical
diagnostic testing of individuals. In this Review, we summarize recent progress and
highlight remaining challenges in monitoring SARS-CoV-2 RNA in wastewater systems for
community and environmental surveillance. Very low concentrations of viral particles and
RNA present in the complicated wastewater and sewage sample matrix require efficient
sample processing and sensitive detection. We discuss advantages and limitations of
available methods for wastewater sample processing, including collection, separation,
enrichment, RNA extraction, and purification. Efficient extraction of the viral RNA and
removal of interfering sample matrices are critical to the subsequent reverse
transcription-quantitative polymerase chain reaction (RT-qPCR) for sensitive detection
of SARS-CoV-2 in wastewater. We emphasize the importance of implementing appropriate
controls and method validation, which include the use of surrogate viruses for assessing
extraction efficiency and normalization against measurable chemical and biological
components in wastewater. Critical analysis of the published studies reveals imperative
research needs for the development, validation, and standardization of robust and
sensitive methods for quantitative detection of viral RNA and proteins in wastewater for
WBE.
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