Minimizing errors in RT-PCR detection and quantification of SARS-CoV-2 RNA for wastewater surveillance

  • Home
  • Minimizing errors in RT-PCR detection and quantification of SARS-CoV-2 RNA for wastewater surveillance

Minimizing errors in RT-PCR detection and quantification of SARS-CoV-2 RNA for wastewater surveillance

25, November 2021 |

Authors:

Ahmed W Simpson SL Bertsch PM Bibby K Bivins A Blackall LL Bofill-Mas S Bosch A Brandão J Choi PM Ciesielski M Donner E D'Souza N Farnleitner AH Gerrity D Gonzalez R Griffith JF Gyawali P Haas CN Hamilton KA Hapuarachchi HC Harwood VJ Haque R Jackson G Khan SJ Khan W Kitajima M Korajkic A La Rosa G Layton BA Lipp E McLellan SL McMinn B Medema G Metcalfe S Meijer WG Mueller JF Murphy H Naughton CC Noble RT Payyappat S Petterson S Pitkänen T Rajal VB Reyneke B Roman FA Jr Rose JB Rusiñol M Sadowsky MJ Sala-Comorera L Setoh YX Sherchan SP Sirikanchana K Smith W Steele JA Sabburg R Symonds EM Thai P Thomas KV Tynan J Toze S Thompson J Whiteley AS Wong JCC Sano D Wuertz S Xagoraraki I Zhang Q Zimmer-Faust AG Shanks OC.

Abstract


Wastewater surveillance for pathogens using reverse transcription-polymerase chain reaction (RT-PCR) is an effective and resource-efficient tool for gathering community-level public health information, including the incidence of coronavirus disease-19 (COVID-19). Surveillance of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) inwastewater can potentially provide an earlywarning signal of COVID-19 infections in a community. The capacity of the world's environmental microbiology and virology laboratories for SARS-CoV-2 RNA characterization in wastewater is increasing rapidly. However, there are no standardized protocols or harmonized quality assurance and quality control (QA/QC) procedures for SARS-CoV-2 wastewater surveillance. This paper is a technical review of factors that can cause false-positive and false-negative errors in the surveillance of SARS-CoV-2 RNA inwastewater, culminating in recommended strategies that can be implemented to identify and mitigate some of these errors. Recommendations include stringent QA/QC measures, representative sampling approaches, effective virus concentration and efficient RNA extraction, PCR inhibition assessment, inclusion of sample processing controls, and considerations for RT-PCR assay selection and data interpretation. Clear data interpretation guidelines (e.g., determination of positive and negative samples) are critical, particularlywhen the incidence of SARS-CoV-2 inwastewater is low. Corrective and confirmatory actionsmust be in place for inconclusive results or results diverging fromcurrent trends (e.g., initial onset or reemergence of COVID-19 in a community). It is also prudent to perform interlaboratory comparisons to ensure results' reliability and interpretability for prospective and retrospective analyses. The strategies that are recommended in this review aim to improve SARS-CoV-2 characterization and detection for wastewater surveillance applications. A silver lining of the COVID-19 pandemic is that the efficacy of wastewater surveillance continues to be demonstrated during this global crisis. In the future, wastewater should also play an important role in the surveillance of a range of other communicable diseases.