Crohn's disease (CD) is a chronic inflammatory disorder of the gastrointestinal tract, and there is strong evidence of considerable overlap between susceptibility loci for CD and mycobacterial infections. Increasing trends of CD are observed worldwide, and it is estimated that there are 400,000-600,000 patients with CD in the US. A clinically and histopathologically similar disease in ruminants, called Johne's disease (JD), is caused by Mycobacterium avium subspecies paratuberculosis (MAP). However, although long hypothesized, the association between MAP and CD remains unproven. This is, in large part, because of a lack of sensitive and specific diagnostic tests that are robust and can be easily applied to screen a sufficiently large number of tissue or clinical samples. After many years of effort, preliminary studies have finally led to the development of a monoclonal antibody (17A12) that specifically recognizes MAP. Hence, we here propose exploratory studies to meet the well-recognized need to develop highly specific and sensitive assays to detect MAP in tissues from CD patients.
Two specific aims are proposed.
In Aim 1, we will develop and establish performance characteristics of an immunohistochemistry (IHC) assay for the detection of MAP in tissues using the MAP-specific monoclonal antibody, 17A12. The IHC assay will be developed, standardized and validated on a collection of paraffin embedded tissue samples from animals with pluribacillary or pauciacillary infections, and from mice experimentally inoculated with other mycobacteria (M. avium and M. tuberculosis). Performance characteristics such as analytical sensitivity, specificity, precision (repeatability and reproducibility), and accuracy of the newly developed assay will be determined.
In Aim 2, we will apply the newly developed IHC assay as a preliminary screen to test intestinal tissue from a convenience sample of CD patients and healthy controls for the presence of MAP and establish initial estimates of analytical and clinical sensitivity and specificity of the newly developed assay. Together, these studies will provide us with a long-needed robust toolkit to specifically detect MAP in tissues from patients with CD, and enable future large scale epidemiologic and mechanistic investigations that can address the association of MAP with CD. In the long-term, the availability of this highly specific and sensitive assay may have important clinical implications for the treatment of CD by enabling the identification of individuals that may benefit from targeted anti- mycobacterial therapy, as well as stimulate the development of control strategies to prevent the contamination of milk and the human food supply with MAP.

Public Health Relevance

Crohn's disease (CD) is a chronic inflammatory bowel disease of major public health significance. Increasing evidence suggests that the development of CD may involve an environmental trigger such as infection with a Mycobacterium. The similarity between CD and bovine Johne's disease (JD), a chronic intestinal disease of cattle caused by infection with Mycobacterium avium subspecies paratuberculosis (MAP) has long been recognized, but attempts at testing the hypothesis that there is an association between MAP and CD have been hampered by a lack of robust specific and sensitive diagnostic assays. We here propose studies to use a newly characterized and highly specific monoclonal antibody to develop a robust immunohistochemistry assay to detect the presence of MAP in tissues of patients with CD. In the long-term, our studies are likely to have important implications for bette understanding disease pathogenesis and developing new treatment options for patients with CD.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Exploratory/Developmental Grants (R21)
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Clinical Research and Field Studies of Infectious Diseases Study Section (CRFS)
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Mills, Melody
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Pennsylvania State University
Veterinary Sciences
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University Park
United States
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Li, Lingling; Katani, Robab; Schilling, Megan et al. (2016) Molecular Epidemiology of Mycobacterium avium subsp. paratuberculosis on Dairy Farms. Annu Rev Anim Biosci 4:155-76