A rapid and visual detection for canine Adenovirus-2 using CRISPR-Cas13a-based SHERLOCK technology

Area of Science:

• Veterinary Diagnostics
• Molecular Biology
• CRISPR Technology

Background:

• Canine adenovirus type 2 (CAdV-2) causes significant respiratory and enteric diseases in puppies.
• Current CAdV-2 diagnostic methods are slow, complex, expensive, and require specialized personnel.
• These limitations hinder rapid diagnosis and control of CAdV-2, particularly in resource-limited areas.

Purpose of the Study:

• To develop a novel, rapid, sensitive, and visually interpreted diagnostic method for CAdV-2.
• To overcome the limitations of existing CAdV-2 detection techniques for field applications.

Main Methods:

• Integrated CRISPR/Cas13a collateral cleavage with HUDSON nucleic acid extraction and recombinase-aided amplification (RAA).
• Utilized a lateral flow strip for visual, naked-eye result interpretation.
• The assay is isothermal, requiring minimal equipment and personnel expertise.

Main Results:

• Achieved a sensitivity of 10^2 copies/μL, with visual detection on lateral flow strips.
• Demonstrated excellent specificity, with no cross-reactivity against five other common canine viruses.
• Showed 95% concordance with conventional simplex PCR on 20 clinical samples, delivering results within 90 minutes.

Conclusions:

• The developed CRISPR/Cas13a-based assay provides a simple, rapid, and accurate point-of-care diagnostic tool for CAdV-2.
• This method has significant potential for field-based surveillance and control of canine infectious diseases.
• Advances the application of CRISPR diagnostics in veterinary medicine.