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J Infect Chemother. 2013 Dec;19(6):1116-25. doi: 10.1007/s10156-013-0636-3. Epub 2013 Jun 23.

High resolution melting curve assay for rapid detection of drug-resistant Mycobacterium tuberculosis.

Author information

1
Mie Prefecture Health and Environment Research Institute, 3684-11 Sakura, Yokkaichi, Mie, 512-1211, Japan, nagaiy02@pref.mie.jp.

Abstract

We developed and evaluated a high resolution melting (HRM) curve assay by using real-time PCR for the detection of the most frequent mutations of Mycobacterium tuberculosis, which are responsible for the resistance of four anti-TB drugs: rifampicin, isoniazid, ethambutol, and streptomycin. The HRM assay was successfully used for the detection of dominant mutations: A516V, H526A, H526T, S531L, L533P, and A516G/S531L in rpoB; S315T, and S315A in katG; -15C/T, and -8T/C in mab-inhA; M306I in embB; K88Q and K43R in rpsL; and 513A/C in rrs. We were able to discriminate the mutant from the wild type by analyzing the melting-curve shape in 40 clinical M. tuberculosis isolates, and the results of the HRM assay were completely consistent with those of DNA sequencing. This HRM assay is a simple, rapid, and cost-effective method that can be performed in a closed tube. Therefore, our assay is a potentially useful tool for the rapid detection of drug-resistant M. tuberculosis.

PMID:
23793795
DOI:
10.1007/s10156-013-0636-3
[Indexed for MEDLINE]

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