Abstract
Mounting evidence suggests that solids are a reliable matrix for SARS-CoV-2 detection in wastewater, yet studies comparing solids-based methods and common concentration methods using the liquid fraction remain limited. In this study, we developed and optimized a method for SARS-CoV-2 detection in wastewater using moderate-speed centrifuged solids and evaluated it against an ultrafiltration reference method. SARS-CoV-2 was quantified in samples from 12 wastewater treatment plants from Alberta, Canada, using RT-qPCR targeting the N2 and E genes. PCR inhibition was examined by spiking salmon DNA. The effects of using different amounts of solids, adjusting the sample pH to 9.6–10, and modifying the elution volume at the final step of RNA extraction were evaluated. SARS-CoV-2 detection rate in solids from 20 mL of wastewater showed no statistically significant difference compared to the ultrafiltration method (97/139 versus 90/139, p = 0.26, McNemar’s mid-p test). The optimized wastewater solids-based method had a significantly lower rate of samples with PCR inhibition versus ultrafiltration (3% versus 9.5%, p = 0.014, Chi-square test). Our optimized moderate-speed centrifuged solids-based method had similar sensitivity when compared to the ultrafiltration reference method but had the added advantages of lower costs, fewer processing steps, and a shorter turnaround time.
Highlights
The current COVID-19 pandemic has prompted a global surge in monitoring SARSCoV-2 in wastewater as an additional and supplementary surveillance tool to inform public health authorities on disease burden
Ct values for SARS-CoV-2 did not change as solids weight increased (p = 0.57 for gene E and p = 0.94 for gene N2, linear regression slope), Ct values for spiked salmon DNA indicated a high level of PCR inhibition, we continued further testing using no more than mg of wet solids
Our ultrafiltration reference method includes a step of pH adjustment to 9.6–10 prior virus concentration to release some of the viruses adsorbed on wastewater solids into the surrounding liquid fraction
Summary
The current COVID-19 pandemic has prompted a global surge in monitoring SARSCoV-2 in wastewater as an additional and supplementary surveillance tool to inform public health authorities on disease burden. An important advantage of wastewater-based surveillance (WBS) over indicators such as clinical cases and hospitalization rates is that it provides a comprehensive snapshot of SARS-CoV-2 presence that includes symptomatic, asymptomatic, and pre-symptomatic carriers with the analysis of a single sample. SARSCoV-2 WBS has the potential to allow early detection of changes in disease burden in advance to clinical testing data and hospitalization rates [1,2]. WBS is not affected by policies and delivery of clinical diagnostics provision, which can be limited by resource allocation and availability. WBS has gained increasing importance since the start of the COVID-19 pandemic
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