Chip-Based Digital PCR Approach for Improved SARS-CoV-2 Quantification for Endemic Wastewater Surveillance

Area of Science:

• Environmental microbiology
• Molecular epidemiology
• Public health surveillance

Background:

• Wastewater-based epidemiology (WBE) is crucial for public health surveillance.
• Low SARS-CoV-2 prevalence challenges traditional WBE methods.
• Improved detection sensitivity is needed for endemic settings.

Purpose of the Study:

• To enhance SARS-CoV-2 detection sensitivity in wastewater during low prevalence.
• To evaluate chip-based digital PCR (dPCR) with magnetic bead concentration.
• To compare N1+N2 and E-Sarbeco gene assays for SARS-CoV-2 detection.

Main Methods:

• Applied chip-based digital PCR (dPCR) targeting CDC N1 and N2 genes.
• Utilized magnetic bead concentration for wastewater sample processing.
• Compared dPCR results with electronegative filtration and analyzed N1+N2 vs. E gene assays.

Main Results:

• dPCR with magnetic bead concentration showed higher viral loads and lower variability than electronegative filtration.
• The N1+N2 gene assay yielded significantly higher SARS-CoV-2 concentrations (4.73 log10 GC/L) compared to the E gene assay (4.27 log10 GC/L).
• PMMoV normalization showed a non-significant increase in correlation between wastewater SARS-CoV-2 and clinical data.

Conclusions:

• Chip-based dPCR combined with magnetic bead concentration improves SARS-CoV-2 detection sensitivity in wastewater.
• The N1+N2 gene target is more sensitive for SARS-CoV-2 detection in endemic wastewater.
• These methods enhance SARS-CoV-2 wastewater surveillance capabilities in low-prevalence settings.