Title
In situ hybridization method for studies of cell wall deficient M-paratuberculosis in tissue samples
Abbreviated Journal Title
Vet. Microbiol.
Keywords
In situ hybridization; M. paratuberculosis; Crohn's disease; spheroplast; POLYMERASE CHAIN-REACTION; CROHNS-DISEASE; MYCOBACTERIUM-PARATUBERCULOSIS; SPHEROPLASTS; ACID; DNA; Microbiology; Veterinary Sciences
Abstract
Cell wall deficient forms of mycobacteria may be important in the pathogenesis of Crohn's disease and sarcoidosis. However, no method has been available to localize this type of organisms in tissue sections. We developed an in situ hybridization method for the demonstration of Mycobacterium paratuberculosis spheroplasts (cell wall deficient forms) in paraffin embedded tissue sections. M. paratuberculosis spheroplasts were prepared by treatment with glycine and lysozyme. Pieces of beef were injected with the prepared spheroplasts. The samples were fixed in buffered formalin and paraffin embedded. A M. paratuberculosis-specific probe derived from the IS900 gene was used. Specificity was controlled by using an irrelevant probe and by hybridizing sections with spheroplasts from other bacteria. Beef samples injected with M. paratuberculosis spheroplasts were the only samples that hybridized with the probe. Beef samples containing acid-fast or spheroplast forms of M. smegmatis and M. tuberculosis Bs well as the acid-fast forms of M. paratuberculosis did not hybridize with the probe. Unrelated bacterial controls, i.e. Helicobacter pylori and Escherichia coli were also negative in the assay. In situ hybridization with the IS900 probe provides a specific way to localize M. paratuberculosis spheroplasts in tissue sections and may be useful for studies of the connection between M. paratuberculosis and Crohn's disease and sarcoidosis. The assay may also be valuable for studies on Johne's diseased animals. (C) 2000 Elsevier Science B.V. All rights reserved.
Journal Title
Veterinary Microbiology
Volume
77
Issue/Number
3-4
Publication Date
1-1-2000
Document Type
Article; Proceedings Paper
Language
English
First Page
513
Last Page
518
WOS Identifier
ISSN
0378-1135
Recommended Citation
"In situ hybridization method for studies of cell wall deficient M-paratuberculosis in tissue samples" (2000). Faculty Bibliography 2000s. 2619.
https://stars.library.ucf.edu/facultybib2000/2619