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- Title
High resolution melting curve assay for rapid detection of drug-resistant <i>Mycobacterium tuberculosis</i>.
- Authors
Nagai, Yuhki; Iwade, Yoshito; Hayakawa, Eri; Nakano, Manabu; Sakai, Takashi; Mitarai, Satoshi; Katayama, Masahiko; Nosaka, Tetsuya; Yamaguchi, Tetsuo
- 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.
- Subjects
MYCOBACTERIUM tuberculosis; DETECTION of microorganisms; DRUG resistance in bacteria; POLYMERASE chain reaction; GENETIC mutation; NUCLEOTIDE sequence
- Publication
Journal of Infection & Chemotherapy (Springer Science & Business Media B.V.), 2013, Vol 19, Issue 6, p1116
- ISSN
1341-321X
- Publication type
Article
- DOI
10.1007/s10156-013-0636-3