Sensitive differential detection of genetically related mycobacterial pathogens in archival material

A polymerase chain reaction (PCR) assay targeted to the immunogenic protein MPB64 gene was used to detect members of the Mycobacterium tuberculosis co mplex, and an outward-primed PCR (OPPCR) designed on the IS6110 element all owed differentiation between Mycobacterium bovis and Mycobacterium tubercul osis. Additionally, the amplification of IS1110 and 16S ribosomal RNA seque nces combined with a dot blotting assay were able to differentially detect Mycobacterium avium, Mycobacterium intracellulare, and Mycobacterium paratu berculosis. The validity of the experimental procedure was tested on refere nce material and formalin-fixed paraffin-embedded samples from patients wit h tuberculosis, sarcoidosis, or Crohn disease. We demonstrated mycobacteria l DNA in 59 of 75 cases,with histologic lesions typical of tuberculosis; we detected M tuberculosis and M paratuberculosis in 6 of 25 sarcoidosis case s and in 7 of 20 Crohn disease specimens, respectively. The proposed diagno stic procedure is directly applicable to archival material and allows diffe rentiation of genetically related mycobacterial pathogens in more detail th an other molecular methods. It provides a tool for the diagnostic study, of tuberculosis, sarcoidosis, and Crohn disease.