Tuberculoid-type lesions occur in the early stages of paratuberculosis, in which it becomes difficult to demonstrate acid-fast forms of MAP. A supersensitive in situ hybridization (ISH) technique was employed to detect the MAP IS900 gene in the tuberculoid-type lesions. Paraffin-embedded sections of mesenteric lymph nodes were obtained from symptomatic and asymptomatic cows. Slides were incubated with a hybridization solution containing a FITC-labeled PCR probe targeted for MAP IS900 for 16 hr at 50ºC. Immunological detection and visualization of the hybridized probe were performed with a commercial kit (GenPointTM Fluorescein, DakoCytomation, Tokyo). Specific signals for MAP IS900 were detected in the lesions of symptomatic cows in macrophages, epithelioid cells and multinucleate giant cells as well as acid-fast forms of MAP. Positive signals were also detected in tuberculoid-type lesions of asymptomatic cows in which no acid-fast bacilli were found. Thus, this ISH procedure provides a way to detect both acid-fast forms of MAP and cell wall-deficient forms of MAP.
(Research Team for Paratuberculosis, Kyushu Research Station, TEL +81-99-268-2078)
Reference:
Tanaka et al. (2005) Vet. Pathol. 42:579-588.
