实验动物, 病原预警方法, 快速检测, CRISPR / Cas13a, MHV

To establish a rapid MHV monitoring method based on environmental media for early pathogen warning in laboratory animal barrier environments. Methods Conserved sequences of MHV were analyzed to design specific RAA primers and crRNA compatible with the LwCas13a reaction system. After optimization, a RAA-CRISPR / Cas13a detection system for MHV was developed, with validation of its specificity and sensitivity, followed by application in animal environmental dust sample detection. Results A rapid MHV detection method based on Recombinase aided amplification ( RAA) combined with CRISPR / Cas13a technology was established for dust samples from laboratory animal habitats. Specificity tests demonstrated no cross-reactivity with common murine viruses including Sendai  virus, Reovirus type 3, Pneumonia virus of mice, Encephalomyelitis virus, and Murine norovirus. The method showed high specificity for MHV detection with a superior sensitivity of 10 copies/ μL detection limit. Clinical validation using PCR as reference involved parallel testing of 160 random dust samples, revealing complete consistency between both methods and identifying 26 antigen-positive samples.Conclusion The RAA-CRISPR / Cas13a detection system based on environmental dust provides efficient and sensitive technical support for pathogen surveillance in laboratory animal facilities.