ArticlePLoS neglected tropical diseases2022
Degenerate sequence-based CRISPR diagnostic for Crimean-Congo hemorrhagic fever virus.
Article in PLoS neglected tropical diseases, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 16 papers.
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Who cites it
16 citing papers in PubMed, 22 citations in OpenAlex.
- Simultaneous Detection of Human Norovirus GI, GII and Hepatitis A Virus Using CRISPR-Cas12a-Based RT-RPA and Lateral Flow Strip Method.Food and environmental virology · 2026Article
- Review
- CRISPR/Cas14a combined with RPA for visual detection of Marek's disease virus.Microbiology spectrum · 2026Article
- Review
- Evolving Landscape of Emerging Virus Diagnosis: Challenges and Innovations.Molecular biotechnology · 2026Review
- Viral RNA persistence and long-term consequences following acute infection with Risk Group 4 zoonotic viruses.Frontiers in cellular and infection microbiology · 2026Review
- CRISPR/Cas Technology for the Diagnosis of Animal Infectious Diseases.Microorganisms · 2025Review
- CRISPR-Cas12a/RPA Dual-Readout Assay for Rapid Field Detection of Porcine Rotavirus with Visualization.Viruses · 2025Article
- Recent Advances in Crimean-Congo Hemorrhagic Fever Virus Detection, Treatment, and Vaccination: Overview of Current Status and Challenges.Biological procedures online · 2024Review
- CRISPR-Cas Technology as a Revolutionary Genome Editing tool: Mechanisms and Biomedical Applications.Iranian biomedical journal · 2023Review
- Strategies to Improve Multi-enzyme Compatibility and Coordination in One-Pot SHERLOCK.Analytical chemistry · 2023Article
- Comparative characterization of Crimean-Congo hemorrhagic fever virus cell culture systems with application to propagation and titration methods.Virology journal · 2023Article
- Latest Advances in Arbovirus Diagnostics.Microorganisms · 2023Review
- Machine learning for design of degenerate Cas13a crRNAs using lassa virus as a model of highly variable RNA target.Scientific reports · 2023Article
- Henipavirus zoonosis: outbreaks, animal hosts and potential new emergence.Frontiers in microbiology · 2023Review
- CRISPR-Cas13a system: A novel tool for molecular diagnostics.Frontiers in microbiology · 2022Review
Corrections and comments
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Authors and funding
6 authors at 3 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
CRISPR (clustered regularly interspaced short palindromic repeats), an ancient defense mechanism used by prokaryotes to cleave nucleic acids from invading viruses and plasmids, is currently being harnessed by researchers worldwide to develop new point-of-need diagnostics. In CRISPR diagnostics, a CRISPR RNA (crRNA) containing a "spacer" sequence that specifically complements with the target nucleic acid sequence guides the activation of a CRISPR effector protein (Cas13a, Cas12a or Cas12b), leading to collateral cleavage of RNA or DNA reporters and enormous signal amplification. CRISPR function can be disrupted by some types of sequence mismatches between the spacer and target, according to previous studies. This poses a potential challenge in the detection of variable targets such as RNA viruses with a high degree of sequence diversity, since mismatches can result from target variations. To cover viral diversity, we propose in this study that during crRNA synthesis mixed nucleotide types (degenerate sequences) can be introduced into the spacer sequence positions corresponding to viral sequence variations. We test this crRNA design strategy in the context of the Cas13a-based SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) technology for detection of Crimean-Congo hemorrhagic fever virus (CCHFV), a biosafety level 4 pathogen with wide geographic distribution and broad sequence variability. The degenerate-sequence CRISPR diagnostic proves functional, sensitive, specific and rapid. It detects within 30-40 minutes 1 copy/μl of viral RNA from CCHFV strains representing all clades, and from more recently identified strains with new mutations in the CRISPR target region. Also importantly, it shows no cross-reactivity with a variety of CCHFV-related viruses. This proof-of-concept study demonstrates that the degenerate sequence-based CRISPR diagnostic is a promising tool of choice for effective detection of highly variable viral pathogens.
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