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Rapid Detection of Ethambutol-resistant Mycobacterium tuberculosis Directly from Sputum Samples by Radioisotope (32P)-based PCR Dot Blot Hybridization and Sequencing Methods

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Aim: to develop an assay system of a radioisotope (32P)-based PCR dot-blot hybridization technique and evaluation of the assay directly for TB sputum samples to detect mutation at codon 306 of embB gene of Mycobacterium tuberculosis related with ethambutol (EMB) resistance.

Methods: one hundred and sixteen of sputum samples were used in this study. Bacterial genome in sputum samples was extracted and tested for mutation at codon 306 of embB gene by the developed PCR dot blot assay using a radioisotope (32P)-labeled oligonucleotide. The positive results were confirmed by DNA sequencing.

Results: all 116 sputum samples were PCR positive for M. tuberculosis. Of 116 samples, three (2.59%) were EMB resistant-M. tuberculosis (MTB) and showed a substitution mutation (ATG/Met’!GTG/Val) at codon 306 of embB gene. None of mutation was detected at codon 299 of embB gene.

Conclusion: we successfully developed a radioisotope (32P)-based PCR dot blot hybridization technique for detection of mutation at codon 306 of embB gene related with EMB resistant M. tuberculosis. The assay can detect a large number of samples that is suitable for monitoring, surveillance, and epidemiology studies.

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Detail Information

Series Title

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Call Number

Actamedica Indonesia, 43(1)2011: 34-38

Publisher

Jakarta : Indonesia Journal of Medicine., 2011

Collation

4p.

Language

English

ISBN/ISSN

0125-9326

Classification

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Content Type

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Media Type

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Carrier Type

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Edition

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Subject(s)

MYCOBACTERIUM TUBERCULOSIS
radioisotopes
SPUTUM
ETHAMBUTOL

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Statement of Responsibility

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