Spatial and Temporal Measurements of SARS-CoV-2 in a Sewershed

The SARS-CoV-2 virus can be shed in the feces of persons infected with COVID-19. As a result, sewer surveillance can be used to identify the presence of COVID-19 infections in a particular sewershed. This study involved monitoring SARS-CoV-2 at strategic locations within a sewershed serving about 600,000 customers in Washington County, Oregon. Sampling was conducted at the four wastewater treatment plants (WWTPs) and at 25 manhole locations within this sewershed from spring 2020 to fall 2021. Samples were concentrated using electronegative filtration and analyzed using a ddPCR system. The 25 manholes were chosen to represent a range of characteristics in their micro-sewersheds including varying numbers of medical centers, industrial sectors, and residential demographics. The reported cases of COVID-19 from the Oregon Health Authority (by zip codes and outbreaks) were correlated with the wastewater virus concentrations. The virus concentrations in the influent to each of the four treatment plants correlated well with the reported COVID-19 cases in Washington County. All four treatment plants showed similar trends of a moderate peak of virus concentrations in summer 2020, followed by dramatic peaks over the holiday season and late summer and fall of 2021. SARS-CoV-2 concentrations at WWTPs appeared to be a leading indicator, with increases in wastewater observed two to three weeks before cases rose (Figure 1). At the manhole sites monitored during summer 2020, the frequency of high positive measurements was highest for several food-processing industrial sites where known outbreaks occurred. During the outbreaks, the virus concentrations in wastewater closely correlated with reported COVID-19 cases at the facilities. Sites with the highest virus concentrations also tended to occur in the zip codes with the highest prevalence of COVID-19. The demographics of residential areas have been shown to affect COVID-19 case prevalence, and a similar trend was also observed in virus concentrations in wastewater. Specifically, neighborhoods with high LatinX and high poverty populations also had higher SARS-CoV-2 concentrations. There was a surprising lack of detectable SARS-CoV-2 in the effluents of hospitals and health centers, even with a known presence of COVID-19 patients. The number of COVID-19 cases relative to the total number of patients in the hospital should have produced a clear signal, given previous positive signals detected from whole neighborhoods and industries with only a few documented cases. Quarternary ammonium compounds (QAC) disinfectants are commonly used to inactivate SARS-CoV-2 on surfaces in hospitals, and they are known to destroy DNA and RNA. These disinfectants may be entering the hospital wastewater via routine disinfection procedures and degrading the SARS-CoV-2 signal, leading to concentrations below the detection limit. Experiments conducted with a QAC disinfectant used in the hospitals in this study showed that there was nearly total decay in the RNA targets at 0.1% concentration, equivalent to 0.128 ounces of disinfectant per gallon of wastewater. It is feasible that disinfectants in the hospital environments are entering the wastewater at concentrations at or above this level and damaging the RNA, thus making it undetectable with the current analytical method. This surprising finding has serious implications for WBE monitoring at and downstream of hospitals and health centers. WWTP influent samples with at least 10,000 N gene copies per liter were also sequenced to determine the SARS-CoV-2 variants present in the samples. Each surge in reported cases corresponded to the dominance of a new variant, and several region-specific sub-variants were identified. The most recent surge in cases during summer and early fall 2021 was associated with the near-total dominance of the Delta variant and its sub-variants. Interestingly, wastewater concentrations during the Delta surge were much higher per reported case than during previous surges, which corresponds to the known increase in viral load in patients infected with the Delta variant. This study has demonstrated that SARS-CoV-2 can be successfully quantified at the micro-sewershed level by selection of appropriate key manholes and used as a surrogate or supplement to swab testing. Such a micro-sewershed approach can be used to identify hot spots of high infections such as industrial facilities and can correlate wastewater concentrations of the virus with community characteristics such as income levels. Such data may prove to be useful to the health care community to determine areas for increased testing or outreach and can be used to optimize data collection and reduce costs.