Missouri's COVID-19 Sewershed Surveillance Project

Background SARS-CoV-2, the virus responsible for COVID-19, is shed in the feces of both symptomatic and asymptomatic individuals and often before the onset of symptoms. Wastewater testing for COVID-19 virus can, therefore, provide population-level information about the circulation of the virus that is not captured by clinical testing. Wastewater testing is a proactive approach to detecting the presence and prevalence of infection and developing public health strategies to mitigate spread of the virus. Missouri's Coronavirus Sewershed Surveillance Project (CSSP) is a collaborative effort to monitor community sewersheds and congregate living facilities for genetic indicators of SARS-CoV-2 in wastewater. At the start of the project in June 2020, CSSP was collecting weekly samples from 12 community wastewater treatment facilities. In subsequent months, community interest in participation in the project grew. Currently, CSSP is collecting samples from 94 community facilities and 36 state congregate facilities, representing over 70% of the state's residents. Methods In CSSP, untreated wastewater (influent) samples are collected weekly or biweekly from select sewersheds and congregate living facilities for detection and analysis of trends of 'true' prevalence of the virus. Twenty-four-hour composite samples of raw influent sewage are collected using auto-sampling devices and shipped on ice to a lab at the University of Missouri-Columbia. The wastewater samples are then filtered, concentrated using polyethylene glycol (PEG) precipitation, and RNA is extracted. Following extraction, SARS-CoV-2 levels are quantitated by qPCR using N1 and N2 primers/probes (CDC). Standard curves are included in every PCR plate so that cycle threshold (Ct) values can be converted to Copy numbers/L from wastewater. In order to account for volumetric dilution that occurs within wastewater systems (e.g. rain fall), RNA concentration (copies/L) are multiplied by facility-specific flow rates (MGD) on the day of sample collection, yielding an estimate of total copies. Samples with high and sufficient viral load are screened for variant mutations using a novel computation workflow based on amplicon sequencing of the SARS-CoV-2 spike N-terminal domain (NTD) and receptor-binding domain (RBD), allowing for detection of variants of concern (VOC) and other lineages within sewersheds. Results Using Spearman's correlation analysis, CSSP found that wastewater concentrations most strongly correlate with positive case counts 4-6 days in the future (Figure 1). In analysis of community sewershed data, the team has found that a ≥45% increase in the exponential weighted moving average (EWMA) of viral genetic fragments in wastewater is proceeded by an increase in clinical cases in the following week approximately 65% of the time. Bland-Altman plots have revealed that wastewater is indeed a reliable proxy marker of current or future clinical cases. Application of Findings CSSP publishes the community sewershed data, including viral load trends and variant mutation detections, in a storymap (https://storymaps.arcgis.com/stories/f7f5492486114da6b5d6fdc07f81aacf) posted on the Missouri Department of Health and Senior Services (MDHSS) website (Figure 3). In addition, CSSP notifies MDHSS staff, local public health agencies (LPHAs), and other stakeholders of significant increases or elevations in viral load and the presence of variant mutations in sewersheds (Figure 4). In April 2021, CSSP began using the wastewater testing results to help identify communities in need of COVID-19 testing. CSSP extends offers for MDHSS to host a free community testing event where sewershed viral load trends are significantly increasing. LPHAs have helped raise awareness of wastewater testing results and interest in testing among their community members by advertising the events, holding meetings, issuing press releases, and conducting interviews. These outreach efforts have been particularly beneficial in rural areas of the state, where vaccination rates are low, COVID-19 testing options are sometimes limited, and, in May and June 2021, mutations of the more transmissible Delta variant were first being detected in the wastewater samples. Conclusions Sewershed surveillance can be utilized to identify potentially high risk communities and assist in data-driven public health efforts/decisions. In the future, sewershed surveillance may be used to better understand the spread of other communicable diseases and possibly even opioid use.