Submitted to: International Veterinary Vaccines and Diagnostics Conference
Publication Type: Abstract Only
Publication Acceptance Date: July 27, 1997
Publication Date: N/A
Technical Abstract: Mycobacterium paratuberculosis (Mptb) is the etiologic agent of paratuberculosis (Johne's disease). In humans, Mptb has been implicated as a possible cause of Crohn's disease. Currently, there is a need for improved diagnostics because of the lack of accurate, rapid, and reliable identification of paratuberculosis (paratb) infection. An Mptb genetic element was cloned, sequenced, and a 30 bp species-specific oligonucleotid (oligo) was synthesized. As an internal control for all mycobacterial species, a 33 bp oligo for the mycobacterial recA gene was synthesized. Dioligo hybridization analysis (DOHA) identified mycobacterial species and specifically identified reference (ATCC 19698), bovine (cattle with Johne's disease), and human (patients with Crohn's disease) isolates of Mptb. The Mptb-specific oligo distinguished Mptb isolates from related mycobacteria, including all closely related M. avium and M. intracellulare strains tested din this study. The Mptb genetic element was also used in the development o a duplex polymerase chain reaction (dPCR) assay. Primers specific to the Mptb sequence were synthesized. As an internal control to confirm that the organisms were mycobacteria, a second set of primers were synthesized based on the conserved 5' terminus of the mycobacterial recA gene. The experiments indicate that both the DOHA and the dPCR assay are useful diagnostic tools to detect mycobacterial infection, specifically paratb.