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Updated: Aug 10, 2026

Quantification and Whole Genome Characterization of SARS-CoV-2 RNA in Wastewater and Air Samples
Published on: June 30, 2023
Accounting for methodological variabilities in disease outbreak surveillance using wastewater-based epidemiology: a
James Boxall-Clasby1,2,3, Nicola Ceolotto4,5,6, Marjorie J Gibbon2,7
1Department of Chemistry, University of Bath, Bath, BA2 7AY, UK.
Wastewater surveillance of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) using an analytical framework improved data sensitivity. Refrigeration enhanced epidemiological insights compared to freezing, and CrAssphage showed high variability for normalization.
Area of Science:
- Environmental microbiology
- Public health surveillance
- Molecular diagnostics
Background:
- Wastewater-based surveillance (WBS) is crucial for community health monitoring.
- Existing SARS-CoV-2 RNA quantification protocols lack comprehensive variability characterization.
Purpose of the Study:
- To develop an analytical framework to quantify variability in SARS-CoV-2 RNA measurement in wastewater.
- To assess the impact of storage, processing, and PCR quantification on epidemiological interpretation.
Main Methods:
- Developed a framework for SARS-CoV-2 RNA quantification in wastewater.
- Analyzed 20,124 RT-qPCR data points for N1, E-Sarbeco, and CrAssphage across four WRCs over 24 months.
Main Results:
- Sample refrigeration provided richer epidemiological data than freezing.
- CrAssphage exhibited high variability, limiting its utility as a normalization biomarker.
- A composite metric (Total Estimated SARS-CoV-2) enhanced sensitivity and reduced non-detects.
Conclusions:
- The developed framework improves sensitivity and interpretation of WBS data.
- Characterizing analytical variability is key for robust pathogen monitoring in wastewater.
- This approach has broad applicability for future WBS methodologies.
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