Singpanomchai N, Akeda Y, Tomono K, et al. - TB control is seriously threatened by multidrug-resistant tuberculosis (MDR-TB) and hence its early diagnosis and proper treatment are essential factors to limit the spread of the disease. Researchers herein developed an allele-specific isothermal recombinase polymerase amplification (allele-specific RPA) that may aid in simultaneous detection of the most common mutations in the rpoB gene at codons 516, 526, and 531, which are linked with rifampicin resistance, and in the katG gene at codon 315, which is linked with isoniazid resistance. They constructed allele-specific primers targeting four major mutations, rpoB516, rpoB526, rpoB531, and katG315, and used these in individual RPA reactions. Evaluation of the optimised RPA assay was done with the Mycobacterium tuberculosis wild-type strain H37Rv and 141 clinical M. tuberculosis isolates. Per findings, allele-specific RPA combined with SYBR Green I detection (AS-RPA/SYBR) led to detection of these four major mutations with 100% sensitivity and specificity relative to DNA sequencing. Integration of AS-RPA/SYBR is supported for the molecular diagnosis of MDR-TB, especially in low-resource settings, given its outstanding performance, including its easy setup, speed, lack of a specific instrument requirement, and lack of cross-reaction with other bacteria.