A rapid and sensitive CRISPR-Cas12a for the detection of Fusobacterium nucleatum

Fusobacterium nucleatum (Fn), as a conditional pathogen, can cause a range of oral and gastrointestinal diseases. However, existing clinical detection methods require expensive equipment and complex procedures, which are inconvenient for large-scale screening in epidemiological research. The purpose of this study was to establish a reliable, rapid, and inexpensive detection method based on CRISPR/Cas12a technology for the detection of Fn. Specific recombinase polymerase amplification (RPA) primer sequences and crRNA sequences were designed based on the nusG gene of Fn. Subsequently, a fluorescence assay and a lateral flow immunoassay were established using the RPA and CRISPR-Cas12a system (RPA-CRISPR-Cas12a). Sensitivity validation revealed a limit of detection of 5 copies/mu L. This method could distinguish Fn from other pathogens with excellent specificity. Furthermore, the RPA-CRISPR-Cas12a assay was highly consistent with the classical quantitative real-time PCR method when testing periodontal pocket samples. This makes it a promising method for the detection of Fn and has the potential to play an increasingly important role in infectious disease testing.