West Nile Virus ( Orthoflavivirus nilense ) RNA concentrations in wastewater solids at five wastewater treatment plants in the United States
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Background
Orthoflavivirus nilense , formerly known as West Nile Virus (WNV), has become endemic to the United States since its introduction in 1999. It has caused an estimated 4.2 million infections since then, and nearly 3,000 deaths. While 80% of infections remain asymptomatic, approximately half of confirmed cases develop into neuroinvasive disease, which is responsible for significant morbidity. Current surveillance methods rely primarily on mosquito pool testing, which is both costly and time-intensive, at best providing delayed epidemiological information. Wastewater-based epidemiology (WBE) has proven an effective method for the surveillance of various pathogens, including other orthoflaviviruses such as Dengue. Given current knowledge of WNV shedding, with over 50% of patients with neurological symptoms shedding it in urine, WBE represents a potentially valuable surveillance approach that has so far been underexplored.
Methods
This study evaluates the viability of using wastewater to assess community prevalence of WNV. We used a targeted droplet digital RT-PCR approach (ddRT-PCR) to measure WNV concentrations in wastewater retrospectively from five locations and in over 600 samples. Three of these locations were in communities with multiple confirmed WNV infections, while two were not. Samples were collected during periods corresponding to typical WNV seasonality (spring to fall). SARS-CoV-2 RNA was measured simultaneously to assess nucleic acid degradation during sample storage. Publicly available confirmed WNV case data were compiled from the California and Nebraska departments of public health and their weekly arboviral reports.
Results
We demonstrated that WNV RNA can be detected in wastewater samples during periods of known viral circulation within a community. We show that the adapted ddRT-PCR assay is highly specific and sensitive, and that detections in wastewater solids correspond to the occurrence of cases in the season and location of sampling. WNV was detected in 9 samples in 3 locations with known WNV clinical cases – wastewater positivity rates in these locations ranged from 3.3% to 13%. The results suggest that wastewater monitoring could serve as an effective complement to traditional surveillance methods, particularly for sentinel surveillance in locations which do not have extensive mosquito and clinical testing systems.
