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Veterinary Medicine International
Volume 2011, Article ID 267102, 5 pages
Research Article

Detection of Mycobacterium avium subsp. paratuberculosis by a Direct In Situ PCR Method

1Instituto de Patobiología, CICVyA, INTA, De Los Reseros y Dr. Nicolás Repetto S/N, Hurlingham, 1686 Buenos Aires, Argentina
2Experimental Pathology Section, Department of Pathology, Instituto Nacional de la Nutrición “Salvador Zubirán”, Vasco de Quiroga 15, Colonia Seccion XVI, Tlalpan,14000 Mexico City, DF, Mexico
3Instituto de Biotecnología, CICVyA, INTA, De Los Reseros y Dr. Nicolás Repetto S/N, Hurlingham, 1686 Buenos Aires, Argentina

Received 12 January 2011; Revised 3 March 2011; Accepted 26 April 2011

Academic Editor: Jesse M. Hostetter

Copyright © 2011 Fernando Delgado et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


In situ detection of Mycobacterium avium subsp. paratuberculosis is useful for diagnosis and research of paratuberculosis. The aim of this paper was to detect this agent in formalin-fixed, paraffin-embedded tissue samples by a direct in situ PCR. The technique was performed on ileum or ileocaecal lymph node samples from 8 naturally infected cattle and 1 healthy calf, by using p89 and p92 primers for amplification of IS900 sequence. Moderate positive signal was detected in all positive samples and not in negative control, but tissues resulted were affected in many cases due to the enzymatic treatment and the high temperature exposition. Although the technique was useful for Map detection, the signal was lower than immunohistochemistry probably because of the fixation process. In one case, signal was higher, which might be due to the detection of spheroplasts. Thus, the described method should be recommended when others resulted negative or for spheroplasts detection.