Detection of Mycobacterium tuberculosis from Paraffinembedded Tissues of Skin Tuberculosis by Nested - Polymerase Chain Reaction

Won, Young Ho; Oh, Jong Suk; Park, Keon; Chun, Inn Ki

Abstract

BACKGROUND: It is well known that the cutaneous tuberculosis especially skin tuberculids is difficult to detect early and make a confirm diagnosis because the organism is seldom detected in a lesion by conventional methods such as AFB stain and culture. The polymerase chain reaction is a very powerful molecular biological tool in diagnosing infectious, neoplastic and genetic diseases. It has the merit of employing various samples, and to improving sensitivity by the modification of PCR techniques. Usage of the routinely formalin-fixed paraffin-embedded tissue as a sample for PCR is very useful in clinical examination, however the sensitivity and specificity have been known to be decreased.
OBJECTIVE
To evaluate the efficacy of the nested-PCR in the diagnosis of different types of cutaneous tuberculosis, we tried to detect the DNA of Mycobacterium tuberculosis from routinely formalin-fixed paraffin-embedded tissue.
METHODS
The various cutaneous tuberculosis and infectious granulomatous disease were retrospecitively selected by the findings of clinical, histopathological and bacterial culture. There were 9 cases of lupus vulgaris, 4 tuberculosis verrucosa cutis, 2 papulonecrotic tuberculid, 2 erythema induratum and 4 lupus miliaris disseminatus faciei. To amplify the 123bp DNA fragment of a repeated insertion sequence IS6110, two pairs of primers composing outside and inside primers were applied.
RESULTS
The positive band was found in 10(91% ) of lupus vulgaris, 3 of tuberculosis verrucosa cutis, 1 papulonecrotic tuberculid and 1 erythema induratum. Four of the LMDF and 5 of the infectious granulomatous diseases were negative.
CONCLUSION
The nested-PCR using routinely formalin-fixed paraffin-embedded tissue is a useful diagnostic tool if combined with the result of other clinical data. Also it may provide further insight into pathogenesis of skin tuberculids and the differential diagnosis of granulomatous skin diseases.