title: Detection of Mycobacterium immunogenum by real-time quantitative Taqman PCR.
abstract: A quantitative real-time 5'-nuclease (Taqman) PCR technique was developed to specifically detect Mycobacterium immunogenum. rpoB-specific primers and Taqman probe were evaluated for detection of M. immunogenum DNA extracted from pure cultures and from industrial metal working fluids (MWFs). Specificity was confirmed and the sensitivity of detection of M. immunogenum genomic DNA was shown to be approximately 9 fg (2 cell equivalents). When tested on industrial metal working fluids from the UK and USA from which no M. immunogenum CFU were recovered, the assay detected between 3.4x10(1) and 1.9x10(4) cell equivalents (CE) per ml, and increased the detection rate over culture to 37.5% (12 of 32 samples). This assay provides a specific, sensitive and rapid method for the detection of M. immunogenum and is applicable within industry for the early detection of this human pathogen and to the possible prevention of hypersensitivity pneumonitis (HP) in workers.
authors.affiliation: Centre for Ecology and Hydrology, Lancaster Environment Centre, Library Avenue, Bailrigg, Lancaster. LA1 4AP, United Kingdom. glenn@ceh.ac.uk
authors.forename: Glenn
authors.initials: G
authors.lastname: Rhodes
authors.forename: Alexandra
authors.initials: A
authors.lastname: Fluri
authors.forename: Marco
authors.initials: M
authors.lastname: Gerber
authors.forename: Alan
authors.initials: A
authors.lastname: Henderson
authors.forename: Andrea
authors.initials: A
authors.lastname: Ruefenacht
authors.forename: Roger W
authors.initials: RW
authors.lastname: Pickup
mesh_terms: D000542:Alveolitis, Extrinsic Allergic / Q000517:prevention & control*; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000235:genetics / Q000302:isolation & purification*; D009784:Occupational Diseases / Q000517:prevention & control*; D016133:Polymerase Chain Reaction / Q000379:methods*; D016415:Sequence Alignment; D006113:United Kingdom; D014481:United States; D014871:Water Microbiology
chemical_list: D001426:Bacterial Proteins; D012321:DNA-Directed RNA Polymerases; C105078:RNA polymerase beta subunit