SARS-CoV-2 / COVID-19 Wastewater Testing

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As COVID-19 infection rates continue to rise, the need for fast, sensitive, reliable tests to monitor cases and reduce the spread of COVID-19 has also become increasingly important. Even as vaccines roll out across the globe, new COVID-19-related challenges continue to arise. In particular, scientists and public health officials have raised the alarm about novel variants of the SARS-CoV-2 coronavirus that have emerged in the UK (B.1.1.7), South Africa (B.1.351), Brazil (P.1), California (B.1.429) and New York (B.1.526). Each variant is characterized by a set of specific, known mutations in its genomic sequence, some of which can increase transmissibility, virulence, or the viruses’ ability to evade immune responses. To fully understand these changes and respond accordingly, researchers need to reliably identify where the variants are, when they appear, and how prevalent they become over time.

  • Wastewater testing can help detect SARS-CoV-2 viral RNA shed in the stool of symptomatic and asymptomatic infected individuals, providing near real-time surveillance of SARS-CoV-2 viral spread at the community level.

  • In this webinar, a team of scientists from Colorado State University (CSU) describe their experiences building a successful wastewater surveillance system to monitor viral loads in wastewater.

Wastewater Surveillance with Droplet Digital PCR

Since symptomatic and asymptomatic SARS-CoV-2 infected individuals shed viral RNA in sewer systems, analysis of SARS-CoV-2 in sewage networks provides information about COVID-19 infection within the community. Wastewater based epidemiology (WBE) for detecting SARS-CoV-2 has been carried out in a range of countries, and a good correlation has been found between the number of COVID-19 cases and SARS-CoV-2 gene concentration in wastewater, forming a basis for the epidemiological role of wastewater testing (Medema et al. 2020). Wastewater based epidemiology has previously been used to track infectious diseases like polio, hepatitis A, and norovirus. The shed viruses are detectable in wastewater samples days before symptoms appear. Researchers can test these samples for specific viral genetic signatures without needing to perform extensive sequencing.

Wastewater surveillance using Droplet Digital PCR (ddPCR) is a comprehensive, cost-effective, highly sensitive way to detect specific variants of the SARS-CoV-2 coronavirus in communities, allowing scientists and public health officials to monitor spread and take action. Droplet Digital PCR takes wastewater testing to the next level with the sensitivity and precision needed to reliably detect the presence of specific variants in wastewater samples, even at extremely low concentrations. Using ddPCR for WBE enables scientists to detect virus 6 days before clinical testing at a sensitivity of 1 infected individual in 10,000, which is extremely important because only 32% of infected individuals seek medical attention. When identifying SARS-CoV-2 variants, ddPCR can accurately discriminate and quantify multiple variants in a sample using a single-well test. Importantly, multiplexing using ddPCR is less prone to artifacts and has simpler data analysis compared to multiplexing using other technologies; therefore, ddPCR can give an accurate measure of the copy number of both wild-type and variant genomes.

# OF COVID WASTEWATER TESTERS
>215 ...and growing
ONLY
32%
of individuals infected
with SARS-CoV-2
seek medical attention1
WASTEWATER SURVEILLANCE FOR SARS-CoV-2:
Detects virus
6 days4
before clinical testing
Detects 1
infected individual in 10,0005
Saves billions of
$$$
when combined
Wastewater surveillance detects SARS-CoV-2
Clinical testing detects SARS-CoV-2
DAYS AFTER
INFECTION
0
1
2
3
4
5
6
7
8
9
10
Infected
Infected person
Contagious2
Contagious person
Symptomatic3
Symptomatic person
Testing
Clinical testing and results
Results
COMMUNITY SPREAD
Infected couple
Infected family
Infected group
Infected community
UNIVERSITY
Wasterwater sampling from a university to track community spread of SARS-CoV-2
Water Sampling
Water sampling
Dorm Sample
Dormatory samples prepared for ddPCR detection of SARS-CoV-2 infections in wastewater
RESIDENTIAL NEIGHBORHOOD
Wasterwater sampling from residential neighborhood to track community spread of SARS-CoV-2
Water Sampling
Water sampling
Community
Sample
Community samples prepared for ddPCR detection of SARS-CoV-2 infections in wastewater
WASTEWATER TREATMENT FACILITY
Wasterwater treatment facility
ddPCR Analysis
ddPCR instrument to detect community-level SARS-CoV-2 infections in wastewater
ddPCR detects community-level SARS-CoV-2 infections in wastewater
ddPCR plate
 

A recent study (Gonzalez 2020) used ddPCR to track SARS-CoV-2 viral shedding in weekly wastewater samples from nine wastewater treatment plants over several months. Increased wastewater SARS-CoV-2 RNA levels predicted increases in confirmed case numbers, highlighting the ability of wastewater surveillance to identify municipality outbreaks before they occur.

Wastewater testing can also be employed by the transport industry to monitor SARS-CoV-2 transmission among aircraft and cruise liner passengers. Another study used ddPCR to analyze wastewater from large transport vessels with their own sanitation systems with implications for contact tracing of disembarking passengers if SARS-CoV-2 is detected in wastewater. SARS-CoV-2 RNA was detected in aircraft and cruise ship wastewater, concluding that wastewater testing has the potential to aid clinical testing and contact tracing for disembarking passengers (Ahmed et el. 2020).

As colleges and universities transition from remote learning to in-person education, administrators must face tough decisions on how to support a safe learning environment. Campus WBE is a crucial component of a comprehensive COVID-19 prevention plan. Academic laboratories are beginning to leverage wastewater testing to prevent campus COVID-19 outbreaks.

Testimonials

Learn how scientists from around the world are monitoring the
community-level spread of SARS-CoV-2 through wastewater testing.

 

End-to-End Solution for SARS-CoV-2 Molecular Testing

With over 5,000+ peer-reviewed publications and thousands of placements worldwide, Bio-Rad’s Droplet Digital PCR is a proven molecular technology for the detection and quantitation of rare targets in complex backgrounds.

Wastewater Assays

SARS-CoV-2, N1, SARS-CoV-2 N2, and BRSV assays are formatted for wastewater testing and come in singleplex, duplex, or triplex formats. Bio-Rad’s Droplet Digital PCR Assays are designed for optimal ddPCR performance and manufactured to reduce the incidence of false positives.

System Feature Comparison

 
QX200 Manual System
QX200 Manual System
QX200 AutoDG System
QX200 AutoDG System
QX ONE System
QX ONE System
Hands-on Time
50 min
10 min
0 min
Sample Throughput
96
96
480
Number of Color Channels
2
2
4
Thermal Cycler
External
External
Integrated

Benefits of Droplet Digital PCR for SARS-CoV-2 Molecular Testing

  • Absolute Quantification

    Directly count DNA molecules and eliminate standard curves

  • Inhibitor Tolerant

    End-point PCR uncouples quantification from amplification efficiency

  • High Sensitivity

    Partitioning allows for enrichment of rare targets

The PREvalence ddPCR SARS-CoV-2 Wastewater Quantification Kit, enables SARS-CoV-2 virus detection in wastewater (sewage).

PREvalence ddPCR SARS-CoV-2 Wastewater Quantification Kit

The new PREvalence ddPCR SARS-CoV-2 Wastewater Quantification Kit includes assay, controls, and supermix to detect SARS-CoV-2 virus in wastewater (sewage). The kit contains reagents sufficient for 200 reactions. An automated software tool is also available for analysis.

See ddPCR Kit Details

CDC Wastewater Surveillance Testing Guidance

The CDC has published guidance on wastewater surveillance testing methods and laboratory workflows to detect SARS-CoV-2 across the U.S.A. In the guidance, droplet digital PCR has been recommended as the detection method for RNA measurement of SARS-CoV-2 in wastewater.

A 1-step RT-ddPCR protocol is advantageous for wastewater because RT is performed in individual droplets, which can reduce RT inhibition compared to RT in bulk solution, as in a 2-step process and in RT-qPCR.

 

Learn More about CDC Wastewater Guidelines

 
Aaron Bivins, University of Notre Dame and Raul Gonzalez, Hampton Roads Sanitation District

COVID-19 Wastewater-Based Epidemiology Webinar

Methods and Insights for SARS-CoV-2 Community Surveillance

Aaron Bivins, University of Notre Dame
Raul Gonzalez, Hampton Roads Sanitation District

Raul Gonzalez and Aaron Bivins discuss their ongoing wastewater surveillance efforts aimed at tracking community spread of SARS-CoV-2. Bio-Rad’s ddPCR platform plays a crucial role in these efforts by offering ultrasensitive SARS-CoV-2 detection in complex wastewater samples.

Watch the Webinar to Learn More

Contact a Specialist

Questions on COVID-19 Surveillance?

Contact a Specialist

More COVID-19 Surveillance Solutions

COVID-19 surveillance is needed to better inform public health decisions. Obstacles to surveillance work include large sample numbers, supply shortages, and complex sample types. Bio-Rad offers a range of solutions for COVID-19 surveillance.

Learn More about Our COVID-19 Surveillance Solutions »

References

  1. Silverman JD et al. (2020). Using influenza surveillance networks to estimate state-specific prevalence of SARS-CoV-2 in the United States. Science Translational Medicine, 12(554), eabc1126
  2. Wei WE et al. (2020). Presymptomatic Transmission of SARS-CoV-2 - Singapore, January 23-March 16, 2020. MMWR. Morbidity and Mortality Weekly Report, 69(14), 411–415
  3. Garg S et al. (2020). Hospitalization Rates and Characteristics of Patients Hospitalized with Laboratory-Confirmed Coronavirus Disease 2019 - COVID-NET, 14 States, March 1-30, 2020. MMWR. Morbidity and Mortality Weekly Report, 69(15), 458–464
  4. Medema G et al. (2020). Presence of SARS-Coronavirus-2 RNA in Sewage and Correlation with Reported COVID-19 Prevalence in the Early Stage of the Epidemic in the Netherlands. Environmental Science & Technology Letters, acs.estlett.0c00357
  5. Hart OE & Halden RU. (2020). Computational analysis of SARS-CoV-2/COVID-19 surveillance by wastewater-based epidemiology locally and globally: Feasibility, economy, opportunities and challenges. Science of the Total Environment, 730, 138875