CRISPR-Cas12a-Based Detection of SARS-CoV-2 Harboring the E484K Mutation

Area of Science:

• Molecular Biology
• Virology
• Epidemiology

Background:

• The SARS-CoV-2 virus is continuously evolving, leading to variants with increased transmissibility and immune escape properties.
• The E484K mutation, identified in the Beta variant, is a significant escape mutation that poses challenges for public health interventions.

Purpose of the Study:

• To develop and apply a CRISPR-Cas12a based diagnostic system for the rapid detection of SARS-CoV-2 harboring the E484K mutation.
• To evaluate the utility of CRISPR diagnostics in identifying escape mutations in clinical samples for epidemiological purposes.

Main Methods:

• Utilized the CRISPR-Cas12a system for sequence-specific detection of SARS-CoV-2 E484K mutation without requiring prior sequencing.
• Exploited the E484K mutation's effect on the protospacer adjacent motif for differential Cas12a recognition.
• Analyzed fecal samples from hospitalized patients to identify infections with the E484K mutation.

Main Results:

• Successfully detected SARS-CoV-2 genomes with the E484K mutation using the CRISPR-Cas12a system.
• Identified one fecal sample from a hospitalized patient in Valencia, Spain, positive for SARS-CoV-2 with the E484K mutation.
• Confirmed the presence of the E484K mutation in the identified sample through subsequent sequencing.

Conclusions:

• CRISPR-Cas12a diagnostics offer a rapid and effective method for detecting specific SARS-CoV-2 mutations, including escape variants.
• This technology holds significant potential as a valuable tool for epidemiological surveillance and monitoring the spread of viral mutations.