Correlation between SARS-CoV-2 concentration in wastewater and COVID-19 case incidence in California National Wastewater Surveillance System (NWSS) sewersheds

Background Monitoring of the SARS-CoV-2 virus, causative agent of coronavirus disease 2019 (COVID-19), in wastewater has been proposed as an adjunctive public health surveillance mechanism that can provide a more comprehensive look at real-time community transmission of viral activity independent of health care seeking or testing behavior. With funding from the Centers for Disease Control and Prevention (CDC) National Wastewater Surveillance System (NWSS) program, the California Department of Public Health (CDPH) partnered with the State Water Resources Control Board (SWRCB) and five sanitation agencies in January 2021 to monitor wastewater at six wastewater treatment plants (WWTPs) servicing large (population > 200,000) metropolitan regions of the state, representing over 30% of California's population; these sites make up CA NWSS. Goals included assessing if wastewater SARS-CoV-2 measurements reflected community COVID-19 case incidence and whether changes in variant predominance and vaccination rates impacted the correlation between wastewater concentration and case incidence. Thus, we evaluated the correlation between wastewater SARS-CoV-2 concentration with COVID-19 case incidence within each sewershed and explored these relationships during the 'winter' 2021 surge and the 'summer' 2021 surge in California. Methods Wastewater influent (24-hour composite, flow or time weighted) were collected three times a week at the main headworks of six California WWTPs (two in Los Angeles County, two in San Francisco County, and one each in San Diego and Orange counties) from January 1 -- August 15, 2021. Analysis occurred at four different laboratories using distinct methods of concentration, extraction, and quantification (qPCR and ddPCR). Measurements were performed for the N1 gene at all laboratories, and N2 and pepper mild mottled virus (PMMoV), a fecal control, at a subset of laboratories at their discretion. Case counts for each sewershed were estimated using shapefiles of treatment plant service area and geocoded line lists of all PCR-confirmed COVID-19 cases in California based on residential address and episode date. To estimate the correlation between the relevant RNA-based metric (e.g., N1, Averaged N1, N1/PMMoV, Averaged N1/PMMoV) and sewershed case counts, a simple linear regression model was applied on the log10-transformed data for each plant. The 'winter' 2021 surge was defined as January 1 -- April 15, 2021, with April 15 chosen as an end date given case counts from the winter surge had plateaued at a low level, vaccination rates in California were around 25-40% and increasing, and regional circulation of Delta was estimated to be <10% of all circulating variants. The 'summer' 2021 surge was defined as June 15-August 15, 2021, with June 15 chosen as a start date as case counts were still low but starting to increase, statewide vaccination rates had reached approximately 50-75%, and prevalence of Delta was > 50% of isolates sequenced. Results Strong, positive, and linear correlations (r2 > 0.75) between sewershed case counts and 10-day average wastewater SARS-CoV-2 concentrations (either N1 or N2 gene copies/l wastewater) were observed for all six sewersheds (Table 1, Figure 1, Figure 2). Correlations were stronger when using the N2 gene compared to N1, with small and variable changes after normalization with PMMoV at sites where those measurements were available. Linear trends and correlations between 'winter' and 'summer' surge time periods appeared to be similar (Figure 3). Conclusion Strong correlations were observed between the weekly averaged wastewater viral RNA concentrations and incidence rates for all six sewersheds. Although Delta variant predominated and vaccination rates increased during the summer surge compared to winter surge, correlations between wastewater concentrations and case counts remained similarly positive during these two periods. These results support the use of wastewater surveillance as an adjunctive estimate of community COVID-19 incidence. Additional analyses could explore the reasons for lower correlation at some sites compared to others, including with normalization to PMMoV.