Related Experiment Video
Updated: May 25, 2026
Field-Deployable Candidatus Liberibacter asiaticus Detection Using Recombinase Polymerase Amplification Combined with CRISPR-Cas12a
Published on: December 23, 2022
A Rapid and Ultrasensitive Detection of Coxsackievirus A16 Using Reverse Transcription Multiple Cross Displacement
Qi Liang1,2,3, Yumei Cao2,4, Xujian Zhang2,5
1Department of Basic Clinical Laboratory Medicine, School of Clinical Laboratory Science, Guizhou Medical University, Guiyang 550001 Guizhou, P. R. China.
Insights
A new diagnostic platform, CVA16-RT-MCDA-CRISPR, offers rapid and sensitive detection of Coxsackievirus A16 (CVA16), a key cause of hand, foot, and mouth disease (HFMD) in young children.
Area of Science:
- Molecular Biology
- Virology
- Biotechnology
Background:
- Coxsackievirus A16 (CVA16) is a major cause of hand, foot, and mouth disease (HFMD) in children.
- Accurate and timely CVA16 detection is vital for patient management and epidemic control, especially in resource-limited settings.
Purpose of the Study:
- To develop a novel diagnostic platform for the rapid and sensitive detection of CVA16.
- To combine reverse transcription multiple cross displacement amplification (RT-MCDA) with CRISPR-Cas12a for CVA16 detection.
Main Methods:
- Designed MCDA primers, an engineered CP1 primer, and a guide RNA (gRNA) targeting the CVA16 VP1 gene.
- Utilized RT-MCDA for preamplification of CVA16 RNA and CRISPR-Cas12a for specific detection of amplicons.
- Validated the assay using CVA16 RNA standards and 96 clinical samples.
Main Results:
- The CVA16-RT-MCDA-CRISPR assay demonstrated a limit of detection of 2.8 × 10-1 copies/µL for CVA16 RNA.
- The assay exhibited high specificity, with no cross-reactivity against other common pathogens.
- Feasibility was confirmed through successful validation with clinical samples.
Conclusions:
- The CVA16-RT-MCDA-CRISPR assay is a reliable, rapid, and sensitive tool for CVA16 detection.
- This platform holds promise for improving diagnostics in settings with limited resources.
- Effective CVA16 detection is crucial for managing HFMD outbreaks.
Abstract:
Coxsackievirus A16 (CVA16) is one of the primary viral etiological agents of hand, foot, and mouth disease (HFMD) in infants and children under five years of age. Prompt and reliable detection of CVA16 is crucial for guiding immediate therapeutic interventions and for implementing effective epidemic prevention and control strategies, particularly in settings with limited resources. Herein, a diagnostic platform for CVA16 (CVA16-RT-MCDA-CRISPR) was developed by combining reverse transcription multiple cross displacement amplification (RT-MCDA) with CRISPR-Cas12a-based detection. In this system, the CVA16 VP1 gene was preamplified using RT-MCDA technology. The resulting amplicons were then specifically recognized and cleaved by the CRISPR-Cas12a-based detection system. MCDA primers, an engineered CP1 primer, and a specific guide RNA (gRNA) were designed to target the VP1 gene of CVA16. The assay achieved a limit of detection of 2.8 × 10-1 copies per microliter for CVA16 RNA standard templates and showed no cross-reactivity against non-CVA16 pathogens. Furthermore, the CVA16-RT-MCDA-CRISPR assay's feasibility was validated using 96 clinical samples. Taken together, these results demonstrate that the CVA16-RT-MCDA-CRISPR assay is a reliable diagnostic tool for rapidly and sensitively detecting CVA16.
Related Concept Videos
01:59CRISPR
01:23The Antiviral System of Bacteria and Archaea: CRISPR