title: Bacteriophage-amplified bioluminescent sensing of Escherichia coli O157:H7.
abstract: Escherichia coli O157:H7 remains a continuous public health threat, appearing in meats, water, fruit juices, milk, cheese, and vegetables, where its ingestion at concentrations of perhaps as low as 10 to 100 organisms can result in potent toxin exposure and severe damage to the lining of the intestine. Abdominal pain and diarrhea develop, which in the very young or elderly can progress towards hemolytic uremic syndrome and kidney failure. To assist in the detection of E. coli O157:H7, a recombinant bacteriophage reporter was developed that uses quorum sensing (luxI/luxR) signaling and luxCDABE-based bioluminescent bioreporter sensing to specifically and autonomously respond to O157:H7 serotype E. coli. The bacteriophage reporter, derived from phage PP01, was tested in artificially contaminated foodstuffs including apple juice, tap water, ground beef, and spinach leaf rinsates. In apple juice, detection of E. coli O157:H7 at original inoculums of 1 CFU mL(-1) occurred within approximately 16 h after a 6-h pre-incubation, detection of 1 CFU mL(-1) in tap water occurred within approximately 6.5 h after a 6-h pre-incubation, and detection in spinach leaf rinsates using a real-time Xenogen IVIS imaging system resulted in detection of 1 CFU mL(-1) within approximately 4 h after a 2-h pre-incubation. Detection in ground beef was not successful, however, presumably due to the natural occurrence of quorum sensing autoinducer (N-3-(oxohexanoyl)-L: -homoserine lactone; OHHL), which generated false-positive bioreporter signals in the ground beef samples.
authors.affiliation: Center for Environmental Biotechnology, University of Tennessee, 676 Dabney Hall, Knoxville, TN 37996-1605, USA. saripp@utk.edu
authors.forename: Steven
authors.initials: S
authors.lastname: Ripp
authors.forename: Patricia
authors.initials: P
authors.lastname: Jegier
authors.forename: Courtney M
authors.initials: CM
authors.lastname: Johnson
authors.forename: Jennifer R
authors.initials: JR
authors.lastname: Brigati
authors.forename: Gary S
authors.initials: GS
authors.lastname: Sayler
mesh_terms: D000368:Aged; D001426:Bacterial Proteins; D010582:Bacteriophage lambda / Q000235:genetics / Q000378:metabolism; D002648:Child; D019453:Escherichia coli O157 / Q000235:genetics / Q000302:isolation & purification*; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology / Q000379:methods*; D006714:Homoserine / Q000031:analogs & derivatives; D006801:Humans; D008163:Luminescent Measurements; D053038:Quorum Sensing; D014157:Transcription Factors / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; C085427:LuxI protein, Bacteria; D014157:Transcription Factors; D006714:Homoserine