Advancements in Programmable prokaryotic Argonautes-assisted sensitive diagnosis of foodborne bacteria: functionalities and multiplexing applications

The global spread of infectious diseases caused by pathogenic microorganisms poses significant challenges to public health. Therefore, it is imperative to develop diagnostic techniques that are sensitive, selective, and user-friendly to effectively control the transmission and progression of these infections. The Argonaute system, recognized as an advanced technique, has been successfully employed in biosensing applications beyond the extensively studied CRISPR/Cas system. While CRISPR/Cas has been widely explored, its reliance on PAM sequence recognition remains relatively limited. Argonaute, recognized for its role as a target-activated nuclease and its programmability, is being repurposed to develop novel sensing techniques. By leveraging its design versatility, the stability of guide DNA, its capacity for multiple-turnover activity, superior nucleic acid cleavage efficiency, and high specificity, Argonaute has been employed in biosensing applications, thereby opening new horizons in analytical chemistry. This paper primarily presents the working mechanism and structure of common Argonaute proteins. The advantages of Argonaute compared to the CRISPR/Cas system in bacterial analysis are outlined and examined. Subsequently, Argonaute-derived sensing approaches for the diagnosis of pathogenic bacteria are comprehensively reviewed, with an emphasis on their design principles. Finally, the prospects and current challenges in this field are critically discussed.