# Establishment of a CRISPR/Cas12a/13a-driven dual-detection platform for rapid diagnosis of swine influenza virus and porcine reproductive and respiratory syndrome virus infection

## TL;DR

A new CRISPR-based test can quickly and accurately detect two major pig viruses in the field.

## Contribution

A dual-detection platform combining RT-LAMP and CRISPR-Cas12a/13a for simultaneous, rapid diagnosis of SIV and PRRSV.

## Key findings

- The assay detected SIV at 5 copies/µL and PRRSV at 2 copies/µL with high specificity.
- The entire process took 25 minutes, including a 20-minute amplification and 5-minute readout.
- Validation showed high agreement with reference methods and resistance to non-target pathogens.

## Abstract

Swine influenza virus (SIV) and porcine reproductive and respiratory syndrome virus (PRRSV) are leading pathogens in pigs, whose co-infections exacerbate disease severity. Current diagnostics like RT-PCR lack suitability for rapid, on-site use, while CRISPR-based systems face challenges in convenient multiplex detection.

We developed an RT-LAMP-CRISPR-Cas12a/13a-LFD dual-detection platform that integrates reverse transcription loop-mediated isothermal amplification (RT-LAMP) with the orthogonal trans-cleavage activities of CRISPR-Cas12a and Cas13a, followed by lateral flow dipstick (LFD) visualization. This assay achieved detection limits of 5 copies/µL for SIV and 2 copies/µL for PRRSV, and exhibited high specificity against other common swine pathogens. The entire process, including a 20-minute amplification at 40 °C and 5-minute LFD readout, enables rapid and visual diagnosis. A preliminary validation was conducted using respiratory infection samples, demonstrating high concordance with reference methods and specificity against non-target pathogens.

The RT-LAMP-CRISPR-Cas12a/13a-LFD assay provides a sensitive, specific, and potentially field-adaptable tool for the simultaneous detection of SIV and PRRSV. It is ideally suited for early screening and precise control of these pathogens in resource-limited settings.

The online version contains supplementary material available at 10.1186/s12985-026-03085-6.