Spatially Informed Wastewater Differentiation among Locations during an Ongoing Measles Outbreak in Texas, USA

Though reverse transcription-quantitative polymerase chain reaction (RT-qPCR), RT digital PCR (RT-dPCR), and RT digital droplet PCR (RT-ddPCR) are commonly used for wastewater-based epidemiology and surveillance (WBE/WBS), differences among the platforms exist. While RT-ddPCR has been suggested as an ideal approach to use globally for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) monitoring, access to RT-qPCR instrumentation is more widely available in many regions, and it is more economical. Subsequently, a larger number of studies have used RT-qPCR for SARS-CoV-2 wastewater monitoring, along with additional pathogens that can be detected with WBS. In this study, we employed RT-qPCR and RT-dPCR platforms for the comparative detection of vaccine (A genotype Edmonston) and wild-type (D8 genotype) measles RNA from wastewater in nearby cities separated by <50 km during an ongoing measles outbreak in Texas, USA, in addition to several other locations up to 30-1400 km from the outbreak location. The limit of detection (LOD) for each methodology was evaluated using a synthetic gBlock DNA gene fragment of known concentrations, with comparable LODs identified for both the RT-dPCR (∼0.5 gc/μL) and RT-qPCR (∼0.4 gc/μL) platforms. Using composite supernatant-solid wastewater samples, RNA aliquots were analyzed on each platform in parallel. The RT-qPCR platform demonstrated a higher detection rate than RT-dPCR for the vaccine strain quantified in wastewater samples, with equivalent detections for the wild-type strain in both platforms, and notable differences in the gene copies quantified in wastewater based on the platform. Our study identifies that regardless of PCR methodology employed, WBS is a particularly valuable approach for the spatially informed differentiation of measles during rapid response to an active outbreak.