A novel electrochemical biosensor based on SRCA-NEMA-G-quadruplex for sensitive detection of Listeria monocytogenes in food

Listeria monocytogenes (L. monocytogenes), as one of the most common foodborne pathogens, has caused many foodborne diseases, posing a major threat to food and public health safety. Therefore, a novel electrochemical biosensor was proposed based on the combination of saltatory rolling circle amplification (SRCA) and nicking enzyme mediated amplification (NEMA) for detecting Listeria monocytogenes. When L. monocytogenes was present, abundant double-stranded DNA (dsDNA) were produced by SRCA reaction. SRCA products could be specifically recognized and cleaved by Nb.BsmI. SRCA-NEMA was performed to generate abundant single-stranded DNA (ssDNA) with the action of Bst DNA polymerase and Nb.BsmI. The hydrosulfuryl-modified hairpin DNA (HShpDNA) was immobilized on the electrode surface and G-quadruplex was used as signaling molecule. The current signal increased with target DNA concentration and the ratio of the current could be calculated as the result. Subsequently, the detection linear range for L. monocytogenes genomic DNA was 5.4 x 100-5.4 x 107 fg/mu L, with the detection limit of 2.13 fg/mu L. In comparison with ISO 11290-1:2017 method, the relative sensitivity, relative specificity and relative accuracy of the developed method were 100 %, 97.8 % and 98 %, respectively. Overall, the developed method provides an outstanding platform for sensitive detection of L. monocytogenes in food.