CRISPR technology mobilized for rapid COVID-19 diagnosis

A group of authors from the United States published a study in the scientific journal Nature Biotechnology in which they have described a CRISPR-Cas-based approach for rapid and accurate diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection – without the need for complex laboratory infrastructure.

3D Rendering Crispr DNA Editing. Image Credit: Nathan Devery / Shutterstock
CRISPR–Cas12-based detection of SARS-CoV-2 - 3D Rendering Crispr DNA Editing. Image Credit: Nathan Devery / Shutterstock

Even though a majority of coronavirus disease (COVID-19) cases during the initial month of the epidemic were sourced back to the city of Wuhan in Chinese province Hubei, the advancing increase in cases worldwide is now driven by local community transmission. Hence, rapid diagnostic tests to detect the presence of SARS-CoV-2 are undoubtedly an urgent public health need.

In previous pandemics and outbreaks, the lack of rapid, convenient, and accurate molecular diagnostic testing has hampered the public health response to emerging viral threats. However, this time we may have the upper hand, as recently devised CRISPR-technology represents the unprecedented chance of reshaping epidemiological surveillance.

"The major pandemics and large-scale epidemics of the past half-century have all been caused by zoonotic viruses," explain authors of the Nature Biotechnology paper. "A diagnostic method that can be readily adapted to detect infection from emergent viruses is urgently needed," they add.

The power of genetic engineering

In this new study, a group of authors affiliated with the University of California San Francisco, Mammoth Biosciences Inc., UCSF-Abbott Viral Diagnostics and Discovery Center, and California Department of Public Health, reported the development of a rapid, uncomplicated and accurate CRISPR–Cas12-based lateral flow test to detect SARS-CoV-2 in respiratory swabs.

With the pervasive use of genetic engineering, CRISPR-Cas systems have been adjusted for use in humans. They are now being adapted and enhanced at a remarkable pace, enabling punctilious editing of basically any DNA or RNA molecule in the body. Consequently, in 2016 CRISPR-Cas systems were first used to detect nucleic acids for molecular diagnostics.

Novel Coronavirus SARS-CoV-2: This scanning electron microscope image shows SARS-CoV-2 (round gold objects) emerging from the surface of cells cultured in the lab.  Credit: NIAID-RML

Novel Coronavirus SARS-CoV-2: This scanning electron microscope image shows SARS-CoV-2 (round gold objects) emerging from the surface of cells cultured in the lab. SARS-CoV-2, also known as 2019-nCoV, is the virus that causes COVID-19. The virus shown was isolated from a patient in the U.S. Credit: NIAID-RML

Today, a plethora of successful CRISPR-Cas-based approaches to detect and diagnose various infectious and non-infectious diseases have been pioneered. Such revolutionary CRISPR-Cas editing has the propensity spread further into the domain of molecular diagnostics and replaces current gold-standard – polymerase chain reaction or PCR – in many applications.

To develop this novel test, the researchers combined CRISPR–Cas12 DETECTR technology with isothermal amplification; the resulting accuracy was comparable to currently used quantitative reverse transcription PCR (qRT–PCR) to detect SARS-CoV-2. However, it can use routine protocols and commercially available 'off-the-shelf' reagents without the need for expensive laboratory instrumentation required by qRT–PCR.

The methods mentioned above of isothermal amplification are especially crucial for molecular diagnostic approaches in remote areas and in laboratories that lack specially trained personnel. Still, the method is not without challenges, mostly linked to the low quality of the clinical sample when test sensitivity/specificity can be compromised.

Faster alternative, same results

The diagnostic approach for SARS-CoV-2 championed in this study was validated with the use of clinical samples from patients in the United States, as well as contrived reference samples. There were 36 study subjects with COVID-19 infection and 42 subjects with other viral respiratory infections. And the published results are encouraging.

"Our CRISPR-based DETECTR assay provides a visual and faster alternative to the US Centers for Disease Control and Prevention SARS-CoV-2 real-time RT–PCR assay, with 95% positive predictive agreement and 100% negative predictive agreement", study authors delineate in the paper.

Some crucial advantages of their approach over qRT–PCR include isothermal signal amplification (which obviates the need for thermocycling), quick turnaround time, single nucleotide target (which increases test specificity), integration with convenient reporting formats, as well as no need for complex laboratory infrastructure.

Transforming the field of molecular diagnostics

The reported cases of asymptomatic infection and subsequent transmission in patients with COVID-19 significantly increase the pool of individuals who should be screened. Moreover, viral titers in hospitalized patients may fluctuate day-to-day (without any correlation to disease severity), while a single negative qRT–PCR test does not exclude infection with SARS-CoV-2.

Tests such as the DETECTR assay reported in this study are amenable to repeat periodic testing of patient samples. This is why clinical validation of this diagnostic method (in response to recent draft guidance from the U.S. Food and Drug Administration) is ongoing in CLIA-certified microbiology laboratories.

"The time taken to develop and validate this SARS-CoV-2 DETECTR assay (which is less than two weeks for SARS-CoV-2) shows that this technology can be quickly mobilized to diagnose infections from emerging zoonotic viruses", study authors state in the paper.

In conclusion, the future development of transportable microfluidic-based cartridges and lyophilized reagents to run the assay could enable point-of-care testing outside of the clinical diagnostic laboratory – such as clinics, local emergency departments, airports, and other locations. CRISPR-based diagnostic tools may dramatically transform the field of molecular diagnostics, making them affordable and accessible basically anywhere in the world.

Journal reference:
Dr. Tomislav Meštrović

Written by

Dr. Tomislav Meštrović

Dr. Tomislav Meštrović is a medical doctor (MD) with a Ph.D. in biomedical and health sciences, specialist in the field of clinical microbiology, and an Assistant Professor at Croatia's youngest university - University North. In addition to his interest in clinical, research and lecturing activities, his immense passion for medical writing and scientific communication goes back to his student days. He enjoys contributing back to the community. In his spare time, Tomislav is a movie buff and an avid traveler.


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