Multiple fluorescent saltatory rolling circle amplification (SRCA) for simultaneous and sensitive detection of Salmonella spp. and Shigella spp. in food

Salmonella spp. and Shigella spp. are known as the leading causes of food poisoning worldwide. In this study, we developed a new multiple fluorescent saltatory rolling circle amplification (MF-SRCA) assay for simultaneous and sensitive detection of Salmonella spp. and Shigella spp. in food based on the generation of real-time fluo-rescence curves that distinguish the positive from the negative samples. In particular, the MF-SRCA can simul-taneously detect two or more targets in one reaction, thus reducing time, reagents, and expenditure, and improving detection efficiency. We showed that MF-SRCA assay was capable of detecting DNA at concentrations as low as 20 fg/mu L for each pathogen. Using the MF-SRCA method, the limit of detection (LOD) for Salmonella spp. and Shigella spp. was 10 CFU/mL in artificially inoculated skimmed milk, which was 100 times lower than the LOD of multiple qPCR. Moreover, the MF-SRCA assay was shown to reliably and effectively detect Salmonella spp. and Shigella spp. in 62 samples with higher positive detection rates (9.7% and 6.5%), compared with multiple qPCR (8.0% and 4.8%) and the national standard method (6.5% and 3.2%), respectively. Therefore, this MF-SRCA assay provides an accurate, reliable and cost-effective alternative method for simultaneous detection of multiple microorganisms in food.

Automated summary

A new MF-SRCA assay was developed for simultaneous detection of Salmonella spp. and Shigella spp. in food. The assay showed high sensitivity and efficiency, with the ability to detect low DNA concentrations and higher positive detection rates compared to other methods.