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Evaluation of Three Viral Capsid Integrity qPCR Methods for Wastewater-Based Viral Surveillance

Capsid Integrity qPCR (CI-qPCR) assays offer a promising alternative to cell culture-based infectivity assays for assessing pathogenic human virus viability in wastewater. This study compared three CI-qPCR methods: two novel (Crosslinker, TruTiter) and one established (PMAxx dye). These methods were evaluated on heat-inactivated and non-heat-inactivated 'live' viruses spiked into phosphate-buffered saline (PBS) and wastewater, as well as on viruses naturally present in wastewater samples. The viral panel included Human adenovirus 5 (HAdV), enterovirus A71 (EV), hepatitis-A virus (HAV), influenza-A H3N2 (IAV), respiratory syncytial virus A2 (RSV), norovirus GI, norovirus GII, and SARS-CoV-2. All three methods successfully differentiated between degraded, heat-inactivated, and live viruses in PBS. While all three methods were comparable for HAdV and norovirus GI, PMAxx detected significantly lower …

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