Improved specificity for detection of Mycobacterium bovis in fresh tissues using IS6110 real-time PCR

BMC Vet Res. 2011 Aug 25;7:50. doi: 10.1186/1746-6148-7-50.

Abstract

Background: Culture of M. bovis from diagnostic specimens is the gold standard for bovine tuberculosis diagnostics in the USA. Detection of M. bovis by PCR in tissue homogenates may provide a simple rapid method to complement bacterial culture. A significant impediment to PCR based assays on tissue homogenates is specificity since mycobacteria other than M. bovis may be associated with the tissues.

Results: Previously published IS6110 based PCR diagnostic assays, along with one developed in house, were tested against environmental mycobacteria commonly isolated from diagnostic tissues submitted to the National Veterinary Services Laboratory. A real-time PCR assay was developed (IS6110_T) that had increased specificity over other IS6110 based assays. Of the 13 non-tuberculous mycobacteria tested with IS6110_T only M. wolinskyi was positive. Thirty M. bovis infected tissue homogenates and 18 control tissues were used to evaluate the potential for the assay as a diagnostic test. In this small sample, IS6110_T detected 20/30 samples from M. bovis infected animals and 0/18 control tissues.

Conclusions: The IS6110_T assay provides a PCR based assay system that is compatible with current diagnostic protocols for the detection of M. bovis in the USA and compliments current testing strategies.

MeSH terms

  • Animals
  • Cattle
  • DNA Transposable Elements / genetics
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / genetics
  • Mycobacterium bovis / genetics
  • Mycobacterium bovis / isolation & purification*
  • Real-Time Polymerase Chain Reaction / veterinary*
  • Sensitivity and Specificity
  • Sequence Analysis, DNA
  • Tuberculosis, Bovine / diagnosis
  • Tuberculosis, Bovine / microbiology*

Substances

  • DNA Transposable Elements
  • DNA, Bacterial