Minimizing errors in RT-PCR detection and quantification of SARS-CoV-2 RNA for wastewater surveillance.
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Ahmed Warish, Simpson Stuart L, Bertsch Paul M, Bibby Kyle, Bivins Aaron, Blackall Linda L, Bofill-Mas Sílvia, Bosch Albert, Brandão João, Choi Phil M, Ciesielski Mark, Donner Erica, D'Souza Nishita, Farnleitner Andreas H, Gerrity Daniel, Gonzalez Raul, Griffith John F, Gyawali Pradip, Haas Charles N, Hamilton Kerry A, Hapuarachchi Hapuarachchige Chanditha, Harwood Valerie J, Haque Rehnuma, Jackson Greg, Khan Stuart J, Khan Wesaal, Kitajima Masaaki, Korajkic Asja, La Rosa Giuseppina, Layton Blythe A, Lipp Erin, McLellan Sandra L, McMinn Brian, Medema Gertjan, Metcalfe Suzanne, Meijer Wim G, Mueller Jochen F, Murphy Heather, Naughton Coleen C, Noble Rachel T, Payyappat Sudhi, Petterson Susan, Pitkänen Tarja, Rajal Veronica B, Reyneke Brandon, Roman Fernando A, Rose Joan B, Rusiñol Marta, Sadowsky Michael J, Sala-Comorera Laura, Setoh Yin Xiang, Sherchan Samendra P, Sirikanchana Kwanrawee, Smith Wendy, Steele Joshua A, Sabburg Rosalie, Symonds Erin M, Thai Phong, Thomas Kevin V, Tynan Josh, Toze Simon, Thompson Janelle, Whiteley Andy S, Wong Judith Chui Ching, Sano Daisuke, Wuertz Stefan, Xagoraraki Irene, Zhang Qian, Zimmer-Faust Amity G, Shanks Orin C
Аннотация
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) in wastewater can potentially provide an early warning 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 in wastewater, 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, particularly when the incidence of SARS-CoV-2 in wastewater is low. Corrective and confirmatory actions must be in place for inconclusive results or results diverging from current 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.