Bacteriophage-based bioluminescent bioreporter for the detection of Escherichia coli 0157:H7

The rapid detection of pathogenic bacteria in food and water is vital for the prevention of foodborne illness. In this study, the lux reporter genes were used in a new bioassay that allows pathogen monitoring without multiple sample manipulations or the addition of exogenous substrate. A recombinant...

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Bibliographic Details
Published in:Journal of food protection 2007-06, Vol.70 (6), p.1386
Main Authors: Brigati, Jennifer R, Ripp, Steven A, Johnson, Courtney M, Iakova, Polina A, Jegier, Patricia, Sayler, Gary S
Format: Article
Language:English
Subjects:
4-Butyrolactone - analogs & derivatives
4-Butyrolactone - metabolism
Aliivibrio fischeri - genetics
Bacterial Proteins - genetics
Bacteriophages
Colony Count, Microbial
Escherichia coli O157 - genetics
Escherichia coli O157 - virology
Food Contamination - analysis
Food Microbiology
Homoserine - analogs & derivatives
Humans
Lactones
Luminescent Measurements
Operon
Recombinant Proteins
Species Specificity
Transcription Factors - genetics
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Summary:The rapid detection of pathogenic bacteria in food and water is vital for the prevention of foodborne illness. In this study, the lux reporter genes were used in a new bioassay that allows pathogen monitoring without multiple sample manipulations or the addition of exogenous substrate. A recombinant phage specific for Escherichia coli 0157:H7 was constructed that, upon infection, catalyzes the synthesis of N-(3-oxohexanoyl)-L-homoserine lactone (OHHL). This phage PP01 derivative carries the luxI gene from Vibrio fischeri under the control of the phage promoter PL. OHHL produced by infected E. coli 0157:H7 induces bioluminescence in bioreporter cells carrying the V. fischeri lux operon. The ability of phage PP0-luxl to detect several strains of E. coli 0157:H7 was confirmed in a 96-well plate assay. In this assay, luxCDABE bioreporter cells capable of detecting OHHL were mixed with phage PPOI-luxl and E. coli 0157:H7, and luminescence was monitored. Reporter phages induced light in bioreporter cells within I h when exposed to 10(4) CFU/ml of E. coli 0157:H7 and were able to detect 10 CFU/ml in pure culture with a preincubation step (total detection time, 4 h). The detection method was also applied to contaminated apple juice and was able to detect 10(4) CFU/ml of E. coli 0157:H7 in 2 h after a 6-h preincubation.
ISSN:0362-028X