Sensitivity, Throughput, and Cost Analysis of Concentration Methods for Multitarget Pathogen Wastewater Monitoring

Wastewater-based epidemiology is an efficient method for monitoring the transmission of diverse pathogens in communities. While various concentration methods are used, most were selected to detect SARS-CoV-2 and other respiratory viruses. Research is needed to guide the method selection for monitoring diverse pathogens in wastewater. In this study, a head-to-head comparison of six different concentration methods was performed, including direct extraction (with and without bead beating), electronegative (HA) filtration, solid concentration, and magnetic bead-based concentration (using Nanotrap particles; with and without bead beating). Methods were assessed for sensitivity, inhibitor removal, recovery rates, and cost, targeting 14 microorganisms including viruses, bacteria, and fungal pathogens. Results showed that the concentration method selection significantly impacts the sensitivity and economic costs of the wastewater monitoring workflow. While no single method was optimal for all targets, combining HA filtration and solid methods in parallel for the same sample is recommended to sensitively detect viruses, bacteria, and fungal pathogens. The magnetic bead-based method can be automated but costs more per sample and is less sensitive for some targets. This study provides data-driven insights to enhance the reliability and cost-effectiveness of wastewater surveillance systems that can support public health responses for a broad range of diseases.