Low cost in-house PCR for the routine diagnosis of extra-pulmonary tuberculosis

SETTING: Conventional methods for the identification of mycobacteria are slow and labour intensive. DNA amplification methods offer rapid sensitive and specific diagnosis. OBJECTIVE: To determine the feasibility of an in-house polymerase chain reaction (PCR) method to detect Mycobacterium tuberculosis in clinical samples. DESIGN: The present study focused mainly on diagnosing extra-pulmonary tuberculosis (EPTB) using an in-house PCR method in 465 clinical samples. This study also compared the efficacy of a standard phenol-chloroform (PC) extraction procedure and the guanidine thiocyanate with diatomaceous silica (GTCS) method of DNA extraction and purification. A subsample of patients was used for the validation of results based on the final diagnosis. RESULTS: Among 373 patients with suspected EPTB, 75 specimens were positive by PCR, four by microscopy and six by culture. Of the 25 PCR-positive patients, 95% had a final diagnosis of TB. Globally, the GTCS method was found to be superior to the PC method for DNA extraction and removal of inhibitors from clinical specimens. CONCLUSION: The DNA amplification method was found to be significantly more sensitive and rapid compared to culture and microscopy for a reliable final diagnosis of EPTB.