title: Quantification of Mycobacterium avium subspecies in pig tissues by real-time quantitative PCR.
abstract: BACKGROUND
  Mycobacterioses in animals cause economical losses and certain Mycobacterium avium subspecies are regarded as potential zoonotic agents. The evaluation of the zoonotic risk caused by M. avium subspecies requires information about the quantities of Mycobacterium strains in infected animals. Because M. avium subspecies in pig tissues are difficult or even impossible to quantify by culturing, we tested the suitability of a culture-independent real-time quantitative PCR (qPCR) assay for this purpose.
  
  
  METHODS
  Mycobacterial DNA was extracted from porcine tissues by a novel method and quantified by Mycobacterium genus specific qPCR assay targeting the 16S rRNA gene.
  
  
  RESULTS
  The response of the qPCR assay to the amount of M. avium subspecies avium mixed with porcine liver was linear in the range of approximately log105 to log107 Mycobacterium cells per 1 g of liver. The assay was validated with three other M. avium subspecies strains. When the assay was applied to porcine lymph nodes with or without visible lesions related to Mycobacterium avium subspecies infections, around 104-107 mycobacterial genomes per gram of lymph nodes were detected.
  
  
  CONCLUSIONS
  The qPCR assay was found to be suitable for the quantification of Mycobacterium avium subspecies in porcine lymph nodes and liver.
authors.affiliation: Department of Production Animal Medicine, University of Helsinki, POB 66, FIN 00014, Helsinki, Finland. taneli.tirkkonen@a-tuottajat.fi
authors.forename: Taneli
authors.initials: T
authors.lastname: Tirkkonen
authors.forename: Timo
authors.initials: T
authors.lastname: Nieminen
authors.forename: Terhi
authors.initials: T
authors.lastname: Ali-Vehmas
authors.forename: Olli A T
authors.initials: OA
authors.lastname: Peltoniemi
authors.forename: Gerard J
authors.initials: GJ
authors.lastname: Wellenberg
authors.forename: Jaakko
authors.initials: J
authors.lastname: Pakarinen
mesh_terms: D000818:Animals; D008099:Liver / Q000382:microbiology; D008853:Microscopy; D009162:Mycobacterium avium / Q000145:classification* / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary*; D013552:Swine