Evaluation of surgical tissue from patients with Crohn's disease for the presence of Mycobacterium avium subspecies paratuberculosis DNA by in situ hybridization and nested polymerase chain reaction

Authors

  • Claudia Romero MS,

    1. Department of Molecular Biology and Microbiology and Biomolecular Science Center, The Burnett College of Biomedical Sciences, University of Central Florida, Orlando, Florida
    Search for more papers by this author
  • Amal Hamdi BS,

    1. Department of Molecular Biology and Microbiology and Biomolecular Science Center, The Burnett College of Biomedical Sciences, University of Central Florida, Orlando, Florida
    Search for more papers by this author
  • John F. Valentine MD,

    1. Department of Medicine, University of Florida and the Gainesville VAMC, Gainesville, Florida
    Search for more papers by this author
  • Saleh A. Naser MD, PhD

    Corresponding author
    1. Department of Molecular Biology and Microbiology and Biomolecular Science Center, The Burnett College of Biomedical Sciences, University of Central Florida, Orlando, Florida
    • Department of Molecular Biology and Microbiology. University of Central Florida, Orlando, Florida 32816
    Search for more papers by this author

Abstract

Crohn's disease (CD) is a chronic inflammatory bowel disease (IBD) with tissue granuloma and histopathological alteration that resembles aspects in tuberculosis, leprosy, and paratuberculosis. Mycobacterium avium subsp paratuberculosis (MAP) is the causative agent of paratuberculosis, with a suspected role in the etiology of CD. We investigated the presence of MAP DNA in 31 surgical tissue samples from 20 subjects using fluorescence in situ hybridization (FISH) with the aid of confocal scanning laser microscopy and nested polymerase chain reaction (PCR) using the IS900 sequence unique to MAP. MAP DNA was detected by PCR in tissue from 10 of 12 (83%) patients with CD: 7/12 (58%) in inflamed, 6/11 (55%) in noninflamed and in 10 (83%) of either tissue and by FISH in 8 of 12 (67%) patients with CD: 7 of 12 (58%) in inflamed, 4 of 11 (36%) in noninflamed, and in 8(67%) of either tissue. In non-IBD subjects, MAP DNA was detected in the tissue of only 1 of 6 patients (17%) by PCR and 0 of 6 patients (0%) by FISH. MAP DNA was identified by PCR in inflamed tissue from 2 of 2 patients with ulcerative colitis. The detection of MAP DNA by either technique in tissue from subjects with CD is significant compared with non-IBD subjects (P < 0.005). Identification of MAP DNA in both inflamed and noninflamed tissue by both techniques suggests that MAP infection in patients with CD may be systemic. The data add more evidence toward a possible association of MAP in the pathogenesis of CD.

Ancillary