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Free Content Efficient mycobacterial DNA extraction from clinical samples for early diagnosis of tuberculosis

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Abstract:

BACKGROUND: Polymerase chain reaction (PCR) detection of Mycobacterium tuberculosis in clinical samples requires the use of an extraction method that can efficiently lyse mycobacterial cells and recover small amounts of DNA.

OBJECTIVE: To evaluate the use of a benzyl-alcohol guanidine hydrochloride (DNA extraction) method (GuHClM) on blood samples.

DESIGN: GuHClM was evaluated in quantitatively spiked blood samples with M. tuberculosis. We assessed the insertion sequence (IS) 6110 region of M. tuberculosis to evaluate the efficacy of the method. The method was also applied on 102 clinical samples of suspected tuberculosis (TB) individuals and compared with smear microscopy of sputum specimens and the results of cultures.

RESULTS: This method reproducibly detected as low as 4–6 bacilli. Of 102 clinical samples, 84 were human immunodeficiency virus (HIV) negative, while 18 were HIV-positive. Among the HIV-negative individuals, 58.3% were TB-positive using PCR, while respectively 47.6% and 45.2% were sputum- and culture-positive. Among the HIV-positive individuals, 55.6% were PCR-positive, whereas only 38.9% were sputum-positive and 50% were culture-positive.

CONCLUSION: These results demonstrate that the identification of mycobacteria by PCR using GuHClM is very sensitive and therefore may have wide utility in the diagnosis of TB.

Keywords: DNA extraction; M. tuberculosis; PCR; benzyl alcohol guanidine hydrochloride; tuberculosis diagnosis

Document Type: Regular Paper

Affiliations: Clinical Microbiology Division, Indian Institute of Integrative Medicine (Council of Scientific & Industrial Research), Jammu, India

Publication date: July 1, 2010

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  • The International Journal of Tuberculosis and Lung Disease publishes articles on all aspects of lung health, including public health-related issues such as training programmes, cost-benefit analysis, legislation, epidemiology, intervention studies and health systems research. The IJTLD is dedicated to the continuing education of physicians and health personnel and the dissemination of information on tuberculosis and lung health world-wide.

    Certain IJTLD articles are selected for translation into French, Spanish, Chinese or Russian. They are available on the Union website

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