Optimization and Application of a Multiplex Digital PCR Assay for the Detection of SARS-CoV-2 Variants of Concern in Belgian Influent Wastewater
Tim Boogaerts,
Siel Van den Bogaert,
Laura A. E. Van Poelvoorde,
Diala El Masri,
Naomi De Roeck,
Nancy H. C. Roosens,
Marie Lesenfants,
Lies Lahousse,
Koenraad Van Hoorde,
Alexander L. N. van Nuijs,
Peter Delputte
Affiliations
Tim Boogaerts
Toxicological Centre, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium
Siel Van den Bogaert
Laboratory for Microbiology, Parasitology and Hygiene, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium
Laura A. E. Van Poelvoorde
Scientific Directorate of Biological Health Risks, Service Transerversal Activities in Applied Genomics, Sciensano, J. Wytsmanstraat 14, 1050 Brussels, Belgium
Diala El Masri
Laboratory for Microbiology, Parasitology and Hygiene, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium
Naomi De Roeck
Laboratory for Microbiology, Parasitology and Hygiene, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium
Nancy H. C. Roosens
Scientific Directorate of Biological Health Risks, Service Transerversal Activities in Applied Genomics, Sciensano, J. Wytsmanstraat 14, 1050 Brussels, Belgium
Marie Lesenfants
Scientific Directorate of Epidemiology and Public Health, Service Epidemiology of Infectious Diseases, Sciensano, J. Wytsmanstraat 14, 1050 Brussels, Belgium
Lies Lahousse
Department of Bioanalysis, Ghent University, Ottergemsesteenweg 460, 9000 Ghent, Belgium
Koenraad Van Hoorde
Scientific Directorate of Infectious Diseases in Humans, Service Foodborne Pathogens, Sciensano, J. Wytsmanstraat 14, 1050 Brussels, Belgium
Alexander L. N. van Nuijs
Toxicological Centre, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium
Peter Delputte
Laboratory for Microbiology, Parasitology and Hygiene, University of Antwerp, Universiteitsplein 1, 2610 Antwerp, Belgium
Since the beginning of the COVID-19 pandemic, the wastewater-based epidemiology (WBE) of SARS-CoV-2 has been used as a complementary indicator to follow up on the trends in the COVID-19 spread in Belgium and in many other countries. To further develop the use of WBE, a multiplex digital polymerase chain reaction (dPCR) assay was optimized, validated and applied for the measurement of emerging SARS-CoV-2 variants of concern (VOC) in influent wastewater (IWW) samples. Key mutations were targeted in the different VOC strains, including SΔ69/70 deletion, N501Y, SΔ241 and SΔ157. The presented bioanalytical method was able to distinguish between SARS-CoV-2 RNA originating from the wild-type and B.1.1.7, B.1.351 and B.1.617.2 variants. The dPCR assay proved to be sensitive enough to detect low concentrations of SARS-CoV-2 RNA in IWW since the limit of detection of the different targets ranged between 0.3 and 2.9 copies/µL. This developed WBE approach was applied to IWW samples originating from different Belgian locations and was able to monitor spatio-temporal changes in the presence of targeted VOC strains in the investigated communities. The present dPCR assay developments were realized to bring added-value to the current national WBE of COVID-19 by also having the spatio-temporal proportions of the VoC in presence in the wastewaters.
