Comparison of Seven Extraction Kits for Detection of SARS-CoV-2 in Wastewater

Wastewater-based epidemiology (WBE) has been used to trace outbreaks and monitor populations for pathogenic viruses such as poliovirus. SARS-CoV-2 has been identified as a candidate for WBE as the virus can survive for prolonged periods in wastewater. The use of commercial extraction kits alongside polyethylene glycol separation, electrostatic membrane filtration, and ultracentrifugation have been used to analyze wastewater. However, the most consistent and versatile commercial kit has not been identified. In this study, previously analyzed wastewater was extracted using seven different commercial kits and quantified using cycle threshold (CT) values from PCR. Data was analyzed using ANOVA and pairwise comparisons at 95% confidence. Results reveal the use of ThermoFisher MagMax Microbiome Kit alongside proteinase K to be the most consistent kit when analyzing soil samples. Qiagen’s EnvironWater kit was identified as the kit of choice for liquid samples. ZymoBIOMICS DNA/RNA kit was identified as a versatile kit for analyzing both solids and liquids. Liquids were found to have higher quantities of virus on average, but solids had lower CT values on average. We conclude that isolating solids should be the primary focus when concentrating virus. MagMax Microbiome Kit with proteinase K was identified as the most consistent and accurate kit for analyzing wastewater. The MagMax Microbiome Kit was found to have significantly lower CT values when compared to other kits across all viral concentrations and sample types. The method used to concentrate water is inexpensive and uses less resources compared to previous methods, which allows WBE to be used at a larger scale in more areas. This will be important for monitoring SARS-CoV-2 variants and population health as the pandemic progresses. Further studies should focus on the efficiency of the kits when using a spiked positive control as well as variance between water samples.https://digitalcommons.unmc.edu/surp2021/1031/thumbnail.jp