The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the virus responsible for the COVID-19 epidemic. Its presence is currently mostly detected with a quantitative PCR test, but this type of test has its limitations:
- It often only targets one region of the virus, meaning that if a mutation would occur in this target, this test would be less effective in detecting the virus.
- It is sensitive to inhibiting components that are often present in faecal and environmental samples, like wastewater. Consequently the test will not detect the virus efficiently.
- It only gives a relative quantification, meaning that a reference curve is necessary to know how much virus is in the sample, which is necessary to being able to compare between different samples and tests.
Sciensano develops a digital multiplex PCR method because it can overcome these limitations, because
- It can detect the virus in samples with a low viral concentration
- It can detect SARS-CoV-2 in faecal and wastewater samples that contain many inhibitory components
- By targeting two separate regions, the test will not fail if a mutation should occur in one of these regions
The DIGICOVID-project develops a multiplex digital PCR method that targets 2 different genomic regions of the SARS-CoV-2 sequence. This allows a more sensitive detection and more precise quantification of SARS-CoV-2 in humans as well as in environmental samples like wastewater. Sciensano develops and validates this method using purified RNA controls and then tested on human samples and environmental samples.
Currently, quantitative PCR (qPCR) is widely used for the detection of SARS-CoV-2. However, these methods target often one region of the viral genome and they encounter several limitations that a two-target digital PCR (ddPCR) may overcome.
A digital PCR test has the following advantages:
- A ddPCR test usually presents a lower limit of detection and is less sensitive to inhibition compared to the qPCR.
- A ddPCR test results in an absolute quantification of a target sequence, thus it does not require a reference curve and therefore facilitates its quantifications.
- Targeting two different parts of the genome allows to deal with the high mutation rate that is often present in viruses and thus may affect the efficiency of the SARS-CoV-2 assay. The use of ddPCR for efficient detection and quantification may be particularly interesting for a fast diagnosis, resulting in the quarantine of patients with a low viral load, and for the detection of SARS-CoV-2 in faecal and environmental samples like wastewater that contain a lot of inhibitory components.