Construction of recA Dual Luciferase Reporter System and Its Application on Screening of Dietary Inhibitors for Shiga Toxin-Converting Bacteriophage Induction

Abstract

Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen that seriously threats the public and animal health. The major virulence of STEC is Shiga toxin (Stx) which is encoded by Stx prophage. Stx prophage induction leads to the production of Stx. Our recent research has shown that dietary functional sugars can differently regulate Shiga toxin type 2 (Stx2) prophage induction. However, high-throughput assays are required to find more dietary inhibitors for Stx prophage induction. This study aims to establish the recA dual-luciferase reporter system for rapidly screening the functional sugars that can inhibit Stx prophage induction. Results showed that the recA dual-luciferase reporter plasmid was successfully constructed as confirmed by DNA sequencing. Established recA dual-luciferase reporter system showed similar screening results when compared with traditional temperate phage determination method (i.e., qPCR) and suggested that L-arabinose is a dietary functional Stx prophage induction inhibitor. Supplementation with L-arabinose in culture medium reduces Stx production in STEC isolated from fecal samples of diarrhea piglets. Collectively, the recA dual-luciferase reporter system is a promising tool for convenient and rapid screening of dietary Stx prophage induction inhibitor, especially suitable for large-scale screening experiments.