Field-Deployable RPA-CRISPR/Cas12a Detection of Decapod Iridescent Virus 1 in Shrimp

Area of Science:

• Aquatic animal health
• Molecular diagnostics
• Viral disease detection

Background:

• Decapod iridovirus 1 (DIV1) causes significant mortality in farmed shrimp (Litopenaeus vannamei), leading to substantial economic losses.
• Early and accurate detection of DIV1 is crucial for effective disease management and preventing widespread outbreaks.

Purpose of the Study:

• To develop a rapid, sensitive, and field-deployable detection platform for Decapod iridovirus 1 (DIV1).
• To integrate recombinase polymerase amplification (RPA) with the CRISPR/Cas12a system for enhanced viral detection.

Main Methods:

• Utilized recombinase polymerase amplification (RPA) for isothermal amplification of the DIV1 ATPase gene.
• Employed the CRISPR/Cas12a system for specific recognition and trans-cleavage of amplified target DNA.
• Optimized reaction conditions for both RPA (38°C, 10 min) and CRISPR/Cas12a (37°C, 20 min) for a total workflow time of 40 minutes.

Main Results:

• Achieved a limit of detection of 2.3 × 10¹ copies/μL for DIV1.
• Demonstrated high specificity, with no cross-reactivity against other common shrimp pathogens.
• Validated the assay's reliable performance using clinical shrimp samples, confirming its practical utility.

Conclusions:

• The integrated RPA-CRISPR/Cas12a platform offers a robust tool for early warning and on-site screening of DIV1.
• Facilitates timely disease control and risk management strategies in shrimp farming.
• Supports the sustainable development of the aquaculture industry through effective pathogen surveillance.