Streamlined CRISPR-based assays for detection and subtyping of H5 and H7 avian influenza

Avian influenza viruses (AIVs) are zoonotic pathogens that pose an increasing global threat due to their potential for significant economic losses in agriculture, and the risk of a pandemic should human-to-human transmission occur. In light of this, we need rapid, sensitive, and specific tests to detect and differentiate circulating AIVs. Current AIV diagnostic methods require specialized laboratory equipment, limiting their use in the field and in low-resource settings. Here, we develop three CRISPR-based Streamlined Highlighting of Infections to Navigate Epidemics (SHINE) assays that address these limitations. We designed, optimized, and validated a SHINE assay for H5 AIV, achieving 100% specificity when tested on seasonal-influenza-positive clinical samples, and a limit of detection of 121.7 copies/μL. We also developed SHINE assays targeting the 2.3.4.4b A(H5N1) and Eurasian H7 lineages. Together, these SHINE assays offer a promising platform for AIV diagnosis and surveillance, particularly outside laboratory settings.