Development of a recombinase polymerase amplification-based chromogenic assay for rapid detection of Salmonella

Salmonella spp. is a leading global cause of foodborne illnesses and poses a significant threat to public health. Rapid and reliable detection of this pathogen is critical for implementing effective prevention and control measures.

A label-free, equipment-free detection method was developed by integrating recombinase polymerase amplification (RPA) with G-quadruplex (G4)/hemin DNAzyme catalysis. Tandem cytosine repeat-modified primers targeting the invA gene were employed to enable the generation of G4 structures after RPA amplification; the resulting G4 amplicons complex with hemin to form a DNAzyme that catalyzes H₂O₂-mediated oxidation of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), producing a naked-eye-detectable color change.

The entire detection process was completed within 1 hour, with a limit of detection (LOD) as low as 12 CFU/ml. Specificity testing confirmed no cross-reactivity with non-target bacteria, including Shigella flexneri (S. flexneri), Enterobacter cloacae (E. cloacae), Listeria monocytogenes (L. monocytogenes), and Escherichia coli (E. coli).

This accurate, portable, and user-friendly detection platform offers promising applications for Salmonella spp. monitoring in food safety surveillance, particularly in resource-limited settings.