Interlaboratory Comparison Using Inactivated Authentic Sars-Cov-2 Variants as a Feasible Tool for Quality Control in Covid-19 Wastewater Monitoring
26 Pages Posted: 18 May 2023
Abstract
Wastewater-based SARS-CoV-2 epidemiology (WBE) has proven as an excellent tool to monitor pandemic dynamics supporting individual testing strategies. WBE can also be used as an early warning system for monitoring the emergence of novel pathogens or viral variants. However, for a timely transmission of results, sophisticated sample logistics and analytics performed in decentralized laboratories close to the sampling sites are required. Since multiple decentralized laboratories commonly use custom in-house workflows for sample purification and PCR-analysis, comparative quality control of the analytical procedures is essential to report reliable and comparable results.In this study, we performed an interlaboratory comparison at laboratories specialized for PCR and high-throughput-sequencing (HTS)-based WBE analysis. Frozen reserve samples from low COVID-19 incidence periods were spiked with different inactivated authentic SARS-CoV-2 variants in graduated concentrations and ratios. Samples were sent to the participating laboratories for analysis using laboratory specific methods and the reported viral genome copy numbers and the detection of viral variants were compared with the expected values.Despite the different procedures, a high concordance regarding the SARS-CoV-2 PCR quantification could be achieved with low variation between the workflows. PCR-based genotyping was, in dependence of the underlying PCR-assay performance, able to predict the relative amount of variant specific substitutions even in samples with low spike-in amount. The identification of variants by HTS, however, required >100 copies/mL wastewater and had limited predictive value when analyzing at a genome coverage below 60%.This interlaboratory test demonstrates that despite different extraction and analysis methods, a high agreement of the SARS-CoV-2 genome copy equivalents could be achieved. Hence, decentralized SARS-CoV-2 wastewater monitoring is feasible to generate comparable analysis results. However, since not all assays detected the correct variant, prior evaluation of PCR and sequencing workflows as well as sustained quality control such as interlaboratory comparisons are mandatory for correct variant detection.
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Funding Information: This study was supported by the German Federal Ministry of Education and Research (BMBF) funding to the projects COVIDready (grant number 02WRS1621A-D), and partly by the project Abwasser Biomarker CoV-2 (02WRS1557A). This study was in part supported in part by the Goethe-Corona-Fund of the Goethe University Frankfurt (M.W.) and by Health Holland.
Declaration of Interests: The authors report supplies provided by QIAGEN GmbH and analytical equipment provided by Analytik Jena (Endress+Hauser Group) on loan for the duration of the project. Qiagen GmbH and Endress+Hauser are associated industry partner of the COVIDready consortium. The funders had no role in the design of the study, in the collection, analyses, or interpretation of data, in the writing of the manuscript, or in the decision to publish the results. The authors (Alexander Wilhelm and Marek Widera) report supplies provided by QIAGEN GmbH and analytical equipment provided by Analytik Jena (Endress+Hauser Group) for the duration of the project. Claudia Stange, Johannes Ho, and Andreas Tiehm reports financial support was provided by Federal Ministry of Education and Research Bonn Office. Marek Widera reports a relationship with AstraZeneca GmbH that includes: speaking and lecture fees. Sandra Ciesek reports a relationship with BioNTech that includes: clinical advisory board membership. Qiagen GmbH and Endress+Hauser are associated industry partner of the COVIDready consortium. The funders had no role in the design of the study, in the collection, analyses, or interpretation of data, in the writing of the manuscript, or in the decision to publish the results.
Keywords: COVID-19 surveillance, SARS-CoV-2 monitoring, wastewater-based epidemiology (WBE), variant detection, interlaboratory comparison, genome sequencing
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