title: Rapid quantification of Escherichia coli in food and media using bacteriophage T7 amplification and liquid chromatography-multiple reaction monitoring tandem mass spectrometry.
abstract: Conventional microbiological assays have been a valuable tool for specific enumeration of indicative bacteria of relevance to food and public health, but these culture-based methods are time-consuming and require tedious biochemical and morphological identification. In this work, we exploit the ability of bacteriophage T7 to specifically infect Escherichia coli and amplify nearly a 100-fold in 1–2 h. Bacteriophage amplification is integrated with liquid chromatography-multiple reaction monitoring tandem mass spectrometry (LC-MRM–MS/MS) for quantitation of phage-specific peptides. Heavy isotopic 15N labeled T7 is introduced as the inoculum phage and internal standard. Quantification is performed by determining the ratio of phage-specific peptides over the internal standard which value is proportional to E. coli numbers. A broad dynamic range of 6-log orders ranging from 3.0 × 10(3) to 3.0 × 10(9) CFU/ml is attained in LB, while between 4.1 × 10(4)–2.7 × 10(9) CFU/ml and 1.9 × 10(3)–3.0 × 10(7) CFU/ml was enumerated respectively in coconut water and apple juice. With this method, viable E. coli are quantified in 4 h with a detection limit of 3.0 × 10(3) CFU/ml, 4.1 × 10(4) CFU/ml and 1.9 × 10(3) CFU/ml in LB, coconut water and apple juice, respectively. This method has potential as a rapid tool for detection of fecal contamination during food bioprocessing and distribution to safeguard public health.
authors.forename: Mazlina
authors.initials: M
authors.lastname: Banu
authors.forename: Daniel
authors.initials: D
authors.lastname: Ng
authors.forename: Lu
authors.initials: L
authors.lastname: Zheng
authors.forename: Lin-Tang
authors.initials: LT
authors.lastname: Goh
authors.forename: Xuezhi
authors.initials: X
authors.lastname: Bi
authors.forename: Dave Siak-Wei
authors.initials: DS
authors.lastname: Ow
mesh_terms: D017123:Bacteriophage T7 / Q000502:physiology*; D001628:Beverages / Q000382:microbiology*; D002853:Chromatography, Liquid; D003059:Cocos; D015169:Colony Count, Microbial; D004926:Escherichia coli / Q000254:growth & development / Q000302:isolation & purification* / Q000821:virology; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology / Q000379:methods*; D005638:Fruit; D027845:Malus; D009587:Nitrogen Isotopes; D009842:Oligopeptides / Q000032:analysis; D053719:Tandem Mass Spectrometry; D014764:Viral Proteins / Q000032:analysis / Q000737:chemistry
chemical_list: D009587:Nitrogen Isotopes; D009842:Oligopeptides; D014764:Viral Proteins