Development of a polymerase chain reaction dot-blotting system for detecting cutaneous tuberculosis

For a definitive diagnosis of cutaneous tuberculosis the demonstration of mycobacteria is essential, but this is generally not possible in skin lesions, Routinely available techniques have poor sensitivity and are time consuming therefore delaying the institution of timely therapy. The high sensitivity and speed of polymerase chain reaction (PCR) for the detection of infectious agents has prompted investigators to use this technique for the detection of Mycobacterium tuberculosis in body fluids such as cerebrospinal fluid or pleural fluid. In the present study, PCR was used to examine punch biopsy specimens from the affected skin of 10 patients with clinical diagnoses of tuberculosis verrucosa cutis, lupus vulgaris, scrofuloderma, papulonecrotic tuberculide and erythema induratum. A control group of 20 patients included individuals having skin manifestations with definite clinical diagnoses other than cutaneous tuberculosis, such as leprosy, fungal mycetoma, chronic bullous disease of childhood and pemphigus vulgaris, The PCR amplified products were dot hybridized with a probe which was random prime labelled with (32)p, The results were compared with routine microbiological and histological findings, Among the test group, six of 10 (60%) were positive for M. tuberculosis by PCR, although their histopathology showed non-specific chronic inflammation with no definite diagnosis. Microbiological investigations, including acid-fast bacillus smear and culture, were positive in a single case of scrofuloderma. All patients in the control group were negative by PCR for ill, tuberculosis. The data indicate that the combination of dot: hybridization with PCR markedly increased the sensitivity and specificity of PCR. This may be a useful tool in the diagnosis of tuberculosis when conventional methods fail.