Operationalizing a routine wastewater monitoring laboratory for SARS-CoV-2

Kantor, Rose; Greenwald, Hannah; Kennedy, Lauren C; Hinkle, Adrian; Harris-Lovett, Sasha; Metzger, Matthew; Thornton, Melissa M.; Paluba, Justin M.; Nelson, Kara L.

doi:10.1371/journal.pwat.0000007

Cited by 22 publications

(32 citation statements)

References 21 publications

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“…A possible dilution caused by inflow of rainwater can be calculated by measuring concentration of pepper mild mottle virus (PMMoV) in the sample and normalizing the SARS-CoV-2 gene copies to PMMeV gene copies. PMMeV is a fecal biomarker present in human feces through diet [25], [26]. For this pilot project, we did not take possible rainwater infiltration into account.…”

Section: Discussionmentioning

confidence: 99%

Detection Of SARS-COV-2 Variants Of Concern In Wastewater Of Leuven, Belgium

Bloemen

Rector

Raymenants

et al. 2022

Preprint

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To investigate whether wastewater surveillance can be used as an early warning system to detect a rise in SARS-CoV-2 positive cases, and to follow the circulation of specific variants of concern (VOC) in particular geographical areas, wastewater samples were collected from local neighborhood sewers and from a large regional wastewater treatment plant (WWTP) in the area of Leuven, Belgium. In two residential sampling sites, a rise in viral SARS-CoV-2 copies in wastewater preceded the peaks in positive cases. In the WWTP, peaks in the wastewater viral load were seen simultaneous with the waves op positive cases caused by the original Wuhan SARS-CoV-2 strain, the Alpha variant and the Delta variant. For the Omicron BA.1 variant associated wave, the viral load in wastewater increased to a lesser degree, and much later than the increase in positive cases, which could be attributed to a lower level of fecal excretion, as measured in hospitalized patients. Circulation of SARS-CoV-2 VOCs (Alpha, Delta and Omicron) could be detected based on the presence of specific key mutations. The shift in variants was noticeable in the wastewater, with key mutations of two different variants being present simultaneously during the transition period.We found that wastewater based surveillance is a sensitive tool to monitor SARS-CoV-2 circulation levels and VOCs in larger regions. This can prove to be highly valuable in times of reducing testing capacity. Differences in excretion levels of various SARS-CoV-2 variants should however be taken into account when using wastewater surveillance to monitor SARS-CoV-2 circulation levels in the population.

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Section: Discussionmentioning

confidence: 99%

Detection Of SARS-COV-2 Variants Of Concern In Wastewater Of Leuven, Belgium

Bloemen

Rector

Raymenants

et al. 2022

Preprint

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“…This is a significant problem because under-reporting may lead to a false sense of security among the public and hinder data-driven decisions by policymakers. Overall, this study indicates the need to continue regular surveillance and heed the warnings of viral genome concentrations in the wastewater as a representative indicator of community health as well as COVID-19 cases in the area [6,[30][31][32][33]. Smaller communities do not always have access to the same resources or information available in larger cities.…”

Section: Discussionmentioning

confidence: 92%

Wastewater surveillance in smaller college communities may aid future public health initiatives

et al. 2022

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To date, the COVID-19 pandemic has resulted in over 570 million cases and over 6 million deaths worldwide. Predominant clinical testing methods, though invaluable, may create an inaccurate depiction of COVID-19 prevalence due to inadequate access, testing, or most recently under-reporting because of at-home testing. These concerns have created a need for unbiased, community-level surveillance. Wastewater-based epidemiology has been used for previous public health threats, and more recently has been established as a complementary method of SARS-CoV-2 surveillance. Here we describe the application of wastewater surveillance for SARS-CoV-2 in two university campus communities located in rural Lincoln Parish, Louisiana. This cost-effective approach is especially well suited to rural areas where limited access to testing may worsen the spread of COVID-19 and quickly exhaust the capacity of local healthcare systems. Our work demonstrates that local universities can leverage scientific resources to advance public health equity in rural areas and enhance their community involvement.

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“…Third, we have found that results of long term-monitoring using RT-qPCR may be subject to plate-to-plate fluctuations due to differences in standard curve serial dilutions (29). While standard curve efficiency and R 2 are key components of quality control, the y-intercept should also be monitored as an indicator of standard degradation or batch differences.…”

Section: Discussionmentioning

confidence: 99%

“…Data analysis was performed using Python (v3.9.5) using modules Pandas (v1.2.5), NumPy (v1.21.0), SciPy (v1.7.0), and Plotnine (v0.8.0). RT-qPCR analysis of standard curves and unknowns was performed using custom code as previously described (29) (https://github.com/wastewaterlab/data_analysis). Pearson correlation coefficients were determined using SciPy, and linear regressions were conducted using NumPy polyfit.…”

Section: Methodsmentioning

confidence: 99%

Comparison of RT-qPCR and Digital PCR Methods for Wastewater-Based Testing of SARS-CoV-2

Hinkle

Greenwald

Metzger

et al. 2022

Preprint

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Wastewater-based epidemiology is an important tool for monitoring SARS-CoV-2 and other molecular targets in populations, using wastewater as a pooled sample. We compared the sensitivity, susceptibility to inhibition, and quantification of reverse transcription quantitative PCR (RT-qPCR), microfluidic well digital RT-PCR (RT-dPCR), and droplet digital RT-PCR (RT-ddPCR) measurements of SARS-CoV-2 (N1 gene target) and Pepper Mild Mottle Virus (PMMoV) RNA in 40 wastewater RNA extracts. All three methods were highly sensitive, but appeared less accurate at very low concentrations. Lower inhibition was observed for RT-ddPCR than RT-qPCR with both SARS-CoV-2 and PMMoV targets, but inhibition appeared to be mitigated by dilution of template RNA. The concentrations of N1 and PMMoV from all three methods were significantly correlated (Pearson's r=0.97-0.98 for N1 and r=0.89-0.93 for PMMoV), although RT-qPCR reported higher concentrations than digital methods. Taken together, this study provides support for the application of all three methods in wastewater-based epidemiology, with additional guidelines for the use of RT-qPCR.

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Operationalizing a routine wastewater monitoring laboratory for SARS-CoV-2

Cited by 22 publications

References 21 publications

Detection Of SARS-COV-2 Variants Of Concern In Wastewater Of Leuven, Belgium

Detection Of SARS-COV-2 Variants Of Concern In Wastewater Of Leuven, Belgium

Wastewater surveillance in smaller college communities may aid future public health initiatives

Comparison of RT-qPCR and Digital PCR Methods for Wastewater-Based Testing of SARS-CoV-2

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