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Botulinum Toxin as an Ultrasensitive Reporter for Bacterial and SARS-CoV-2 Nucleic Acid Diagnostics

Song, F;Shen, Y;Wei, Y;Yang, C;Ge, X;Wang, A;Li, C;Wan, Y;Li, J;

The rapid identification of pathogenic microorganisms plays a crucial role in the timely diagnosis and treatment strategies during a global pandemic, especially in resource-limited area. Herein, we present a sensitive biosensor strategy depended on botulinum neurotoxin type A light chain (BoNT/A LC) activated complex assay (BACA). BoNT/A LC, the surrogate of BoNT/A which embodying the most potent biological poisons, could serve as an ultrasensitive signal reporter with high signal-to-noise ratio to avoid common strong background response, poor stability and low intensity of current biosensor methods. A nanoparticle hybridization system, involving specific binding probes that recognize pathogenic 16S rRNAs or SARS-CoV-2 gene site, was developed to measure double-stranded biotinylated target DNA containing a single-stranded overhang using Fluorescence Resonance Energy Transfer (FRET)-based assay and colorimetric method. The method is validated widely by six different bacteria strains and severe acute respiratory related coronavirus 2 (SARS-CoV-2) nucleic acid, demonstrating a single cell or 1 aM nucleic acid detecting sensitivity. This detection strategy offers a solution for general applications and has a great prospect to be a simple instrument-free colorimetric tool, especially when facing public health emergency.