Background.

M. avium subspecies paratuberculosis (MAP) is the cause of Johne's disease and has been implicated in Crohn's disease. Cell wall deficient (CWD) forms have been suggested as the pathogenic form of the organism in humans. It is uncertain if CWD forms are present in tissue of animals with Johne's disease. We used a previously developed an in situ hybridization assay for CWD MAP to study animals.

Purpose.

To determine whether CWD MAP were present in diseased animals and to validate the in situ hybridization assay in animal tissue.

Methods.

Paraffin-embedded tissues (intestines, liver and lymph nodes) from 36 coded archival specimens (representing 14 cows and one sheep) were tested by in situ hybridization assay using a digoxigenin-labeled IS900 probe. Positive and negative controls were beef tissues injected with MAP CWD forms and acid fast bacilli, respectively.

Results.

Six of the 10 JD infected animals were positive for MAP (60% sensitivity); 4 intestinal and 2 intestinal/lymph nodes. The single positive liver specimen originated from a cow with JD whose intestinal specimen was also positive. None of the 5 control cows was positive (100% specificity).

Conclusion.

The presence of CWD Map in animals with JD may explain why MAP isn't readily cultured from some animals and/or why pathology is seen in tissues like the liver when no acid fast bacilli are detected. CWD forms may represent a highly evolved survival strategy as the lack of the bacterium's outermost structural layer could assist in evading or delaying the host's defense system, thereby allowing it to quietly establish itself within the host.