Solid–liquid partitioning of dengue, West Nile, Zika, hepatitis A, influenza A, and SARS-CoV-2 viruses in wastewater from across the USA

Limited information is available on the fate of respiratory and arthropod-borne viruses in wastewater. Enteric viruses have been extensively studied in wastewater treatment plants, however partition coefficients have not been well documented. This information is essential for interpreting wastewater-based surveillance (WBS) data and optimizing sample collection and processing methods. In this study, we examined the solid–liquid partitioning behavior of dengue, West Nile, Zika, hepatitis A, influenza A, and SARS-CoV-2 viruses in wastewater. Samples were collected from the primary sludge line of eleven wastewater treatment plants across the United States and spiked with varying concentrations of each virus. Solid and liquid fractions were separated via centrifugation. Viral nucleic acids were extracted and quantified using reverse-transcription digital droplet PCR (RT-ddPCR). Partition coefficients (K_F), determined using the Freundlich adsorption model, ranged from 4.0 × 10² mL g⁻¹ to 3.9 × 10⁶ mL g⁻¹ (median = 1.1 × 10⁴ mL g⁻¹). We applied a multiple linear regression model to evaluate the effects of factors like viruses and wastewater treatment plants on virus partitioning. We found that the individual effects of those variables were not significant, however, their combined effect was significant. Specifically, significant differences were observed between K_F for Zika and West Nile virus between wastewater treatment plants. Further research is needed to understand how wastewater characteristics might impact the partition of viral markers. The results from this experiment underscore the importance of considering wastewater solids for the early detection and monitoring of viral infectious diseases, particularly in regions with low prevalence of infections.