Most days each week, a courier transports a soda can-sized container of wastewater collected from a large sewer main in the middle of the Stanford campus to a nearby independent lab for analysis.

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These daily samples from the Codiga sewershed, an area covering more than 150 buildings and 10,000 people on campus, provide clues about the rise and fall of the virus’ prevalence on campus, often several days ahead of the trends identified by the university’s testing program.

Within 24-48 hours of drop-off at the lab, Alexandria Boehm, a professor of civil and environmental engineering at Stanford, receives the results online. Boehm, known to colleagues as Ali, reviews the data and uses it to create reports about the prevalence of COVID-19 on campus. In April, Stanford’s COVID Dashboards expanded to include these measurements and trends.

The streamlined process began as a pilot project involving a team of researchers overseen by Boehm and Marlene Wolfe, a Stanford postdoctoral scholar at the time and now an assistant professor at Emory University, and led by civil and environmental engineering graduate student Winnie Zambrana. The Stanford Provost’s office funded the pilot because of its potential for anticipating COVID prevalence on campus and informing decisions on health protocols.

Separately, Boehm and Wolfe also lead the Sewer Coronavirus Alert Network (SCAN), a project that samples and analyzes wastewater solids for SARS-CoV-2 in California communities.

The SCAN collaboration involves the California Department of Public Health, local public health officers, and 11 wastewater treatment plants in the greater Bay Area and Central California, as well as the life science commercial laboratory Verily. Early on, Stanford began working locally with Santa Clara County.

“The COVID-19 response has been predicated on the ability to make rapid, data-informed decisions, which depend on access to timely and accurate data,” said Michael Balliet, deputy director of the county’s Public Health Department. “Santa Clara County has been the only jurisdiction in California to consistently monitor over 98% of its population using this groundbreaking surveillance method. We were the first to start posting it for public consumption on our website, and have successfully been using it in our daily information briefings for almost two years. This is all thanks to the robust public/academic partnership with Stanford.”

Here, Boehm discusses her team’s work and the great promise that wastewater monitoring holds for anticipating the spread of other diseases. The interview has been edited for clarity.

Why does this wastewater monitoring work for detecting COVID?

Although mostly a respiratory disease, SARS-CoV-2 is excreted in the feces of people who have COVID-19, as well as in respiratory secretions. When people who are infected use the bathroom, shower, etc. the virus will enter the sewer system. We can take a sample from the wastewater in the sewers or at a treatment plant to estimate the disease occurrence in the area that the wastewater came from.

Walk us through the steps from obtaining a wastewater sample to estimating COVID rates in a community.

A sample of wastewater is liquid with a bunch of suspended solids in it. We use the wastewater solids for our project because the virus is in much higher concentrations in the wastewater solids compared to the liquid – it essentially sticks to the solids. We take those solids and extract viral genetic information found in RNA. We then use a lab technique called droplet digital PCR to measure the concentrations of the genes in SARS-CoV-2. We are looking both for genes that are present in all SARS-CoV-2 genomes (and thus representing all cases of COVID-19), and genes that are characteristic of different variants of concern. These PCR tests are a lot like the PCR tests that are used for clinical testing, in that they target a section of the genome of SARS-CoV-2 and amplify, or copy, it until it can be detected with a fluorescent, or glowing, molecular probe or tag. The concentrations we measure are strongly associated with COVID-19 cases in the surrounding communities.

How are the results different from COVID tests?

Clinical COVID tests give you results on whether one individual is infected or not, and then you need to aggregate the data from everyone who has taken a test to tell what the incidence rates of COVID are.

Using wastewater, you can take one sample, and the concentration of SARS-CoV-2 RNA in the wastewater is associated with the number of cases in the area. So it’s just one test instead of many, and it also captures people who may not be testing for whatever reason (asymptomatic, no access to testing, etc.).

Also, it’s important to note that results from rapid tests are not compiled by local public health officials, so information on disease incidence cannot be gleaned from rapid test results. So wastewater will increase in importance in tracking COVID incidence as more and more people use rapid tests.

How did this project start?

When we heard about the novel coronavirus circulating in Wuhan, China, in January 2020, I and, at the time, visiting Professor Krista Wigginton of the University of Michigan applied for a RAPID grant from National Science Foundation (NSF) to monitor wastewater on campus for coronaviruses; we also had other research aims in the proposal. The NSF program officer at the time questioned whether the proposed work was relevant for the nation, an important criterion for selecting grants to fund, but after the very first cases of COVID-19 were detected locally in Santa Clara County, they agreed to fund our proposal.

The university shut down the next day. Starting our work with locked doors and a shelter in place order was one of the hardest things I have ever experienced as a researcher. But with the support of the Civil and Environmental Engineering Department manager and other staff, as well as Ellyn Segal and Ryan McAllister from the university’s Biosafety & Biosecurity group, we were able to start our work under these very challenging conditions. When we started this project, we did not know if detection of SARS-CoV-2 in wastewater was even possible. Now, we know its concentrations correlate very strongly with COVID-19 cases and that wastewater surveillance is a valuable tool for pandemic response.

Wastewater monitoring has been used at universities around the country since early in the pandemic. We did studies on different areas of campus to demonstrate the feasibility and use of this system from January to March 2021. That research, which was just published, led us to focus on sampling at the William and Cloy Codiga Resource Recovery Center to give an indicator for the whole campus.

How did you collaborate within Stanford, the broader research community, and public health officials to develop and deploy this technology?

Early in the pandemic, we connected with great collaborators at the CDC, the California Department of Public Health, and the Santa Clara County Public Health Department. Santa Clara County has been supportive since early in the pandemic and engaged early on with us through weekly, and often more frequent, meetings over Zoom.

At Stanford, the Provost’s office provided seed funding in late 2020 for a pilot project using wastewater to gain insights about COVID-19 on campus. That project involved School of Medicine collaborators Ami Bhatt, Ben Pinsky, and Bonnie Maldonado, as well as Julia Nussbaum of the Stanford Water Planning & Stewardship team. A number of students and postdocs were involved in designing the whole program, including the methods we’re now using on campus, and Winnie Zambrana, a PhD student in the Department of Civil and Environmental Engineering, led the study.

We meet regularly and discuss our newest data, plans for testing new variants and pathogens, and how to use the data for pandemic response. We also work closely with Russell Furr’s group at Environmental Health & Safety and communicate with them weekly. All these groups are essential partners for our work.

What are some other potential ways this technology might be used in the future that you’re excited about?

We have shown that wastewater can be used to track changing trends in respiratory syncytial virus (RSV) and influenza A incidence in communities, and we are working to investigate its utility for detecting a wide range of infectious diseases from rhinovirus to HIV to Dengue.

We believe that wastewater, as a composite biological sample, can be used to gain deep insight into circulating diseases and be used to inform the use of therapeutics that may only work for specific variants of diseases; recommendations for non-pharmaceutical interventions such as masking and social distancing; hospital resources and testing centers; and even vaccine development. We are excited to be a part of the research and translation of this technology.