Cold-chain free nucleic acid preservation using porous super-absorbent polymer (PSAP) beads to facilitate wastewater surveillance.

The instability of viral targets including SARS-CoV-2 in sewage is an important challenge in wastewater monitoring projects. The unrecognized interruptions in the 'cold-chain' transport from the sample collection to RNA quantification in the laboratory may undermine the accurate quantification of the virus. In this study, bovine serum albumin (BSA)-modified porous superabsorbent polymer (PSAP) beads were applied to absorb raw sewage samples as a simple method for viral RNA preservation. The preservation efficiency for SARS-CoV-2 and pepper mild mottle virus (PMMoV) RNA were examined during storage for 14 days at 4 °C or room temperature against the control (no beads applied). While a non-significant difference was observed at 4 °C (∼80 % retention for both control and PSAP-treated sewage), the reduction of SARS-CoV-2 RNA concentrations was significantly lower in sewage retrieved from PSAP beads (25-40 % reduction) compared to control (>60 % reduction) at room temperature. On the other hand, the recovery of PMMoV, known for its high persistence in raw sewage, from PSAP beads or controls were consistently above 85 %, regardless of the storage temperature. Our results demonstrate the applicability of PSAP beads to wastewater-based epidemiology (WBE) projects for preservation of SARS-CoV-2 RNA in sewage, especially in remote settings with no refrigeration capabilities.