LuxCDE-luxAB-based promoter reporter system to monitor the Yersinia enterocolitica O:3 gene expression in vivo

PubMed ID: 28235077 ; WOS: 000394688200160 ; It is crucial to understand the in vitro and in vivo regulation of the virulence factor genes of bacterial pathogens. In this study, we describe the construction of a versatile reporter system for Yersinia enterocolitica serotype O:3 (YeO3) based on the luxCDABE operon. In strain YeO3-luxCDE we integrated the luciferase substrate biosynthetic genes, luxCDE, into the genome of the bacterium so that the substrate is constitutively produced. The luxAB genes that encode the luciferase enzyme were cloned into a suicide vector to allow cloning of any promoter-containing fragment upstream the genes. When the obtained suicide-construct is mobilized into YeO3-luxCDE bacteria, it integrates into the recipient genome via homologous recombination between the cloned promoter fragment and the genomic promoter sequence and thereby generates a single-copy and stable promoter reporter. Lipopolysaccharide (LPS) O-antigen (O-ag) and outer core hexasaccharide (OC) of YeO3 are virulence factors necessary to colonization of the intestine and establishment of infection. To monitor the activities of the OC and O-ag gene cluster promoters we constructed the reporter strains YeO3-P-oc ; This work was supported by the Center for International Mobility (CIMO) fellowship grants KM -13-8757 and TM-14-9081 (http://www.cimo.fi/ frontpage), FEMS (http://tems-microbiology.org/ fems-work/grants/fems-research-grants/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. ; Center for International Mobility (CIMO) fellowship [KM -13-8757, TM-14-9081]; FEMS