Improved Scalability, Sensitivity, and Interpretability of Pathogen Detection, Including SARS-CoV-2, in Wastewater using High-Throughput, Highly Multiplexed Digital Array PCR Technology

Presently, the application of molecular technology such as RT-qPCR and digital PCR (dPCR) to quantify SARS-CoV-2 and related targets in wastewater is cumbersome, time-consuming, and costly. While progress has been made on the development of methods and the interpretation of data, much remains to be improved for the technology to be used as a public health management tool. A major drawback in the current approaches are: 1) the lack of streamlined and consistent pre-analytical processing steps, 2) coverage across the relevant targets requires a high number of reactions (>20) from any single sample to provide quantitative information, and 3) a lack of vision of the development of a pathogen/marker panel, much like those used in clinical arenas, for interpretation of the data across different states, regions and nations. The goal of this project was to successfully navigate these three limitations toward development of an advanced digital array PCR system as part of the development of public health warning system that is not dependent on clinical testing and has the ability to rapidly address novel pathogen thrust in the future.