Detection of M. paratuberulosis in Crohn's Disease

描述（由申请人提供）：克罗恩病（CD）是一种慢性胃肠道炎症性疾病，有强有力的证据表明，在CD和分枝杆菌感染的易感位点之间有相当大的重叠。在世界范围内，乳糜泻呈上升趋势，据估计，美国有40万至60万乳糜泻患者。反刍动物的临床和组织病理学上相似的疾病，称为约翰病（JD），是由鸟分枝杆菌亚种副结核（MAP）引起的。然而，尽管长期假设，MAP和CD之间的关系仍未得到证实。这在很大程度上是因为缺乏敏感和特定的诊断测试，这些测试是可靠的，可以

DESCRIPTION (provided by applicant): 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.