Polyethylenimine mediated recovery of SARS-CoV-2 and total viral RNA: Impact of aqueous conditions on behaviour and recovery.

ABSTRACT

Wastewater-based epidemiology (WBE) is an emerging, practical surveillance tool for monitoring community levels of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, SC2). However, a paucity of data exists regarding SARS-CoV-2 and viral biomarker behaviour in aqueous and wastewater environments. Therefore, there is a pressing need to develop efficient and robust methods that both improve method sensitivity and reduce time and cost. We present a novel method for SARS-CoV-2, Human Coronavirus 229E (229E), and Pepper Mild Mottle Virus (PMMoV) recovery utilizing surface charge-based attraction via the branched cationic polymer, polyethylenimine (PEI). Initially, dose-optimization experiments demonstrated that low concentrations of PEI (0.001% w/v) proved most effective at flocculating suspended viruses and viral material, including additional unbound SC2 viral fragments and/or RNA from raw wastewater. A design-of-experiments (DOE) approach was used to optimize virus and/or viral material aggregation behaviour and recovery across varying aqueous conditions, revealing pH as a major influence on recoverability in this system, combinatorially due to both a reduction in viral material surface charge and increased protonation of PEI-bound amine groups. Overall, this method has shown great promise in significantly improving quantitative viral recovery, providing a straightforward and effective augmentation to standard centrifugation techniques.