Assessing the impact of fluctuating viral loads on virus recovery from electronegative membrane filters used for wastewater surveillance of biologicals

• Electronegative membrane filters (EMFs) do not reliably capture a cumulative or average viral load under fluctuating concentration scenarios. • Results determined EMFs exhibit inconsistent virus retention, as well as desorption of MS2 over time, reducing recovery and potentially underestimating viral loads. • While EMFs remain a cost-effective and easy approach, they may be unsuitable for accurately quantifying viral loads in fluctuating wastewater systems. The emergence of SARS-CoV-2 hastened the adoption of wastewater-based epidemiology (WBE) for public health surveillance. While passive sampling results for SARS-CoV-2 appeared to agree with clinical sampling and infection numbers within the community, at the time there was limited investigation into factors that may affect the accuracy of results obtained through passive sampling. This study investigated the impact of fluctuating viral loads on virus recovery from a passive sampling material (electronegative membrane filters [EMFs]) using F-RNA bacteriophage MS2 as a model virus. Three models of fluctuating viral loads were tested: steady-state, substantial increase/decrease, and intermittent exposure. This resulted in eight concentration fluctuation scenarios. Experiments assessed whether concentration fluctuations impact virus recovery from EMFs, and whether there is an average or cumulative uptake of the exposed viral load. Findings showed that virus recovery reflected neither scenario, indicating that concentration fluctuations in wastewater impact the predictability of recovery from EMFs. Two predicted recovery models were calculated based on each recovery hypothesis, whether EMFs have an average or cumulative recovery. These models consistently under-predicted virus recovery during scenarios of increasing concentrations and over-predicted recovery during stable or decreasing concentrations. The measured recovery sometimes reflected the most recent concentration the samplers were exposed to rather than providing a reliable cumulative or average measurement, highlighting their limitations in accurately capturing dynamic or steady viral load conditions. This research enhances understanding of the complexities associated with using passive sampling for WBE, informing public health surveillance and policy development.