Evaluation of reverse transcription loop-mediated isothermal amplification in conjunction with ELISA-hybridization assay for molecular detection of Mycobacterium tuberculosis

Traditional culture, followed by a panel of biochemical tests for the diagnosis of tuberculosis (TB), is time-consuming, and rapid identification of Mycobacterium tuberculosis is crucial for the early administration of appropriate therapy. In this study, the reverse transcription loop-mediated isothermal amplification combined with enzyme-linked immunosorbent hybridization (RT-LAMP-ELISA-hybridization) assay has been designed for the rapid detection of 16S rRNA in clinical isolates of M. tuberculosis. This assay reproducibly detected a single copy, as opposed to 2000 copies of MTB; 16S rRNA detected by conventional gel electrophoresis. Among the 150 specimens of sputum analysed, RT-LAMP-ELISA-hybridization assay had a sensitivity of 94.1% in the culture method, compared to the Amplified M. tuberculosis Direct Test (AMTD), 91.1% and the 88.2% sensitivity of acid-fast staining. Furthermore, RT-LAMP-ELISA-hybridization assay is more cost-effective when compared to the real-time TaqMan RT-PCR and AMTD assays. In conclusion, our results suggest that the RT-LAMP-ELISA-hybridization assay is a highly sensitive, low cost diagnostic tool useful for the rapid and accurate direct diagnosis of sputum specimens, and is suitable for routine clinical use. Crown Copyright (c) 2008 Published by Elsevier B.V. All rights reserved.