Quantification of Viruses in Wastewater on a Centrifugal Microfluidic Disc

Rapid and onsite detection of pathogens in water is a critical first step in preventing the spread of infectious diseases from the environment to humans. However, current microbial monitoring practices are tedious, expensive, and slow. These limitations significantly impede our ability to promptly identify potential risks to public health, leading to delays in implementing the necessary interventions. In this study, we report the development of a portable centrifugal microfluidic disc (CD) that integrates sample concentration, purification, and a droplet digital reverse transcription LAMP (ddRT-LAMP) assay as a lab-on-a-chip system for rapid virus detection in the environment. Coupled with the pseudo- and nonpseudo forces generated during the CD rotation or oscillation, the assay steps for sample purification, concentration, and quantification were completed in less than 1.5 h. The results showed that the on-CD sample preparation procedures are comparable to the traditional in-tube sample preparation assay. Furthermore, the indigenous pepper mild mottle virus (PMMoV) in raw sewage at concentrations ranging from 6.0 × 10⁴ to 2.1 × 10⁷ copies/ml was successfully quantified using the complete on-CD assay, which includes on-CD sample preparation procedures and on-CD ddRT-LAMP. The concentrations of PMMoV detected by the CD assay matched well with those detected by the state-of-the-art virus nucleic acid extraction and ddRT-PCR assay for wastewater and wastewater-spiked environmental water samples, demonstrating the potential of this CD platform for environmental applications.