Abstract
Wastewater-based epidemiology (WBE) has enabled us to describe Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infections in populations. However, implementation of wastewater monitoring of SARS-CoV-2 is limited due to the need for expert staff, expensive equipment, and prolonged processing times. As WBE increases in scope (beyond SARS-CoV-2) and scale (beyond developed regions), there is a need to make WBE processes simpler, cheaper, and faster. We developed an automated workflow based on a simplified method termed exclusion-based sample preparation (ESP). Our automated workflow takes 40 min from raw wastewater to purified RNA, which is several times faster than conventional WBE methods. The total assay cost per sample/replicate is $6.50 which includes consumables and reagents for concentration, extraction, and RT-qPCR quantification. The assay complexity is reduced significantly, as extraction and concentration steps are integrated and automated. The high recovery efficiency of the automated assay (84.5 ± 25.4%) yielded an improved Limit of Detection (LoDAutomated=40 copies/mL) compared to the manual process (LoDManual=206 copies/mL), increasing analytical sensitivity. We validated the performance of the automated workflow by comparing it with the manual method using wastewater samples from several locations. The results from the two methods correlated strongly (r = 0.953), while the automated method was shown to be more precise. In 83% of the samples, the automated method showed lower variation between replicates, which is likely due to higher technical errors in the manual process e.g., pipetting. Our automated wastewater workflow can support the expansion of WBE in the fight against Coronavirus Disease of 2019 (COVID-19) and other epidemics.
Original language | English |
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Article number | 109595 |
Journal | Journal of Environmental Chemical Engineering |
Volume | 11 |
Issue number | 2 |
DOIs | |
State | Published - Apr 2023 |
Bibliographical note
Publisher Copyright:© 2023 Elsevier Ltd
Funding
The work was funded by National Institutes of Health (NIH) grants U01DA053903-01 and P30 ES026529 , Centers for Disease Control and Prevention (CDC) contract BAA 75D301-20-R-68024 . The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, NIH: Quantitative PCR (qPCR) Extraction Control from Heat-Inactivated SARS-Related Coronavirus 2, Isolate USA-WA1/2020, NR-52350. The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, NIH: Genomic RNA from SARS-Related Coronavirus 2, Isolate USA-CA4/2020, NR-52508. The work was funded by National Institutes of Health (NIH) grants U01DA053903-01 and P30 ES026529, Centers for Disease Control and Prevention (CDC) contract BAA 75D301-20-R-68024. The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, NIH: Quantitative PCR (qPCR) Extraction Control from Heat-Inactivated SARS-Related Coronavirus 2, Isolate USA-WA1/2020, NR-52350. The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, NIH: Genomic RNA from SARS-Related Coronavirus 2, Isolate USA-CA4/2020, NR-52508. We thank Blazan Mijatovic, Cullen Hunter, and Savannah Tucker, field technicians at the University of Kentucky, for their assistance in collection of wastewater samples.
Funders | Funder number |
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BEI Resources | |
National Institutes of Health (NIH) | U01DA053903-01, P30 ES026529 |
Centers for Disease Control and Prevention | BAA 75D301-20-R-68024 |
National Institute of Allergy and Infectious Diseases | NR-52508, NR-52350 |
Keywords
- Automation
- COVID-19
- SARS-CoV-2
- Wastewater
- Wastewater-based epidemiology
ASJC Scopus subject areas
- Chemical Engineering (miscellaneous)
- Waste Management and Disposal
- Pollution
- Process Chemistry and Technology