Disease: CRISPR SARS-CoV-2 Detection platform for Coronavirus Disease (COVID-19)

Disease info:

COVID-19 is the infectious disease caused by the coronavirus, SARS-CoV-2, which is a respiratory pathogen.

The most common symptoms of COVID-19 are: fever, dry cough, fatigue

Other symptoms that are less common and may affect some patients include: loss of taste or smell, nasal congestion, conjunctivitis (also known as red eyes), sore throat, headache, muscle or joint pain, different types of skin rash, nausea or vomiting, diarrhea, chills or dizziness.

Most people (about 80%) recover from the disease without needing hospital treatment. About 20% of those who get COVID-19 become seriously ill and require oxygen, with 5% becoming critically ill and needing intensive care.

Complications leading to death may include respiratory failure, acute respiratory distress syndrome (ARDS), sepsis and septic shock, thromboembolism, and/or multiorgan failure, including injury of the heart, liver or kidneys.

In rare situations, children can develop a severe inflammatory syndrome a few weeks after infection.

In most situations, a molecular test is used to detect SARS-CoV-2 and confirm COVID-19. Polymerase chain reaction (PCR) is the most commonly used molecular test. Samples are collected from the nose and/or throat with a swab. Molecular tests detect virus in the sample by amplifying viral genetic material to detectable levels. For this reason, a molecular test is used to confirm an active infection, usually within a few days of exposure and around the time that symptoms may begin. 

Rapid tests (sometimes known as a rapid diagnostic test – RDT) detect viral proteins (known as antigens). Samples are collected from the nose and/or throat with a swab. These tests are cheaper than PCR and will offer results more quickly, although they are generally less accurate. 

Frequency:
Due to the ongoing pandemic, frequency data of COVID-19 cases are monitored via the WHO COVID-19 Dashboard. Globally, as of 10:33am CEST, 17 October 2020, there have been 39,023,292 confirmed cases of COVID-19, including 1,099,586 deaths.
Official title:
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Partners:
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Method:
DNA Endonuclease-Targeted CRISPR- Cas12 Trans Reporter (DETECTR)
Condition:
Coronavirus Disease (COVID-19)
Target gene:
Nucleocapsid (N)
Safety updates:

Status:

Description

The SARS-CoV-2 RNA DETECTR Assay is a molecular diagnostic test intended for the qualitative detection of nucleic acid from the SARS-CoV-2 in upper respiratory specimens (nasopharyngeal swabs, oropharyngeal (throat) swabs, mid-turbinate nasal swabs, anterior nasal swabs, nasopharyngeal wash/aspirate or nasal aspirate) from individuals suspected of COVID-19 by their healthcare provider.

The assay, SARS-CoV-2 RNA DNA Endonuclease-Targeted CRISPR Trans Reporter (DETECTR), performs simultaneous reverse transcription and isothermal amplification using loop-mediated amplification (RT-LAMP) for RNA extracted from nasopharyngeal or oropharyngeal swabs in universal transport media (UTM), followed by the addition of the Cas12 enzyme, after which cleavage of a reporter molecule confirms detection of the virus. The assay uses fluorescence measurements on the ABI 7500 Fast Dx instrument with a total run time of 45 minutes (30 minutes RT-LAMP, followed by 15 minutes Cas12 detection) as the test readout.

This test uses primer sets and CRISPR guide RNAs (gRNAs) specific for the detection SARS-CoV-2 using reverse transcription-loop-mediated isothermal amplification (RTLAMP) with the Bst DNA polymerase and Cas12-mediated cleavage of the amplified product. The target is the N gene, along with primers and gRNAs for the human RNaseP (RP) gene as a sample internal control. The target region of the N gene overlaps with the N2 region of the FDA Emergency Use Authorization (EUA)-authorized CDC assay.

Controls for the assay include positive control (PC), human specimen control (HSC), universal transport medium (UTM) and negative template/extraction water control (NTC).

NTC, PC, HSC and UTM are extracted with each extraction run along with patient samples.

The PC consists of AccuPlex SARS-CoV-2 Verification Panel reference material (SeraCare, Catalog # 0505-0129) spiked at 18,000 copies/mL in negative patient matrix (corresponding to 30 copies/µL or 1.5X the claimed LoD of 20 copies/µL. The PC serves as a positive control for amplification of the N gene target.

The AccuPlex SARS-CoV-2 Verification Panel reference material consists of a target synthetic viral RNA, including the N gene, encapsulated in a viral protein coat. As the reference material spiked into negative matrix is fully extractable, this reference material serves as a fullprocess control for the DETECTR assay.

The HSC serves as a positive control for the presence of human nucleic acid in the sample, which is targeted by the RP gene.

The UTM serves as a control for contamination in the extraction oramplification.

The NTC serves as a control for contamination during both the extraction and PCR reagent preparation.

The RP target also serves as an internal control for each patient sample, ensuring that human nucleic acid is present in the sample.

Last updated: Apr. 5, 2021
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