Development of an engineered bioluminescent reporter phage for the sensitive detection of viable Salmonella typhimurium.

Abstract

Because foodborne illnesses continuously threaten public health, rapid and sensitive detection of pathogens in food has become an important issue. As an alternative to time-consuming and laborious conventional detection methods, a technique using recombinant reporter phages has been developed. Here, we developed an advanced bioluminescent reporter phage SPC32H-CDABE by inserting a bacterial luxCDABE operon into the Salmonella temperate phage SPC32H genome. Whole SPC32H genome sequencing enabled the selection of nonessential genes, which can be replaced with approximately 6-kb luxCDABE operon, which provides both luciferase (LuxAB) and its substrate, fatty aldehyde, as generated by fatty acid reductase (LuxCDE). Thus, the SPC32H-CDABE detection assay is simpler and more efficient compared to the luxAB-based assay because the substrate addition step is excluded. At least 20 CFU/mL of pure S. Typhimurium culture was detectable using SPC32H-CDABE within 2 h, and the signals increased proportionally to the number of cells contaminated in lettuce, sliced pork, and milk. These results thereby demonstrate that this phage successfully detects live Salmonella without appreciable interference from food components. Furthermore, the presented data suggest that SPC32H-CDABE represents a promising easy-to-use diagnostic tool for the detection of Salmonella contamination in food.