title: One-tube multiplex PCR method for rapid identification of mycobacterium tuberculosis.
abstract: A rapid, inexpensive, simple, and accurate multiplex polymerase chain reaction (PCR) was developed in a single tube for identification of Mycobacterium tuberculosis. Assessment of sensitivity and specificity of simple PCR was performed with 116 strains of M. tuberculosis complex (MTC) and 144 strains of nontuberculous mycobacteria (NTM) compared with the biochemical method. Specific amplification of KS4, MTC-specific DNA fragment, was found in 98% (114/116) of MTC and not detected in 99% (143/144) of NTM. Amplification of the mtp40 gene revealed 95% sensitivity (100/105 strains of M. tuberculosis) and 77% specificity (not found in 119/155 mycobacterial strains). A multiplex PCR method based on the combination of KS4- and mtp40-derived primers was used for identification of M. tuberculosis. Crude DNA from slow growing mycobacteria with cream rough colonies that showed both 768-bp amplified product for KS4 and 396-bp for mtp40 was identified as M. tuberculosis whereas that from MTC gave only the 768-bp product.
authors.affiliation: Molecular Biology of Fungi and Mycobacteria Laboratory, Department of Microbiology, Faculty of Medicine at Siriraj Hospital, Mahidol University, Bangkok, Thailand. siacp@mahidol.ac.th
authors.forename: Angkana
authors.initials: A
authors.lastname: Chaiprasert
authors.forename: Therdsak
authors.initials: T
authors.lastname: Prammananan
authors.forename: Nipa
authors.initials: N
authors.lastname: Tingtoy
authors.forename: Preeyawis
authors.initials: P
authors.lastname: Na-Ubol
authors.forename: Somboon
authors.initials: S
authors.lastname: Srimuang
authors.forename: Kittipan
authors.initials: K
authors.lastname: Samerpitak
authors.forename: Watcharin
authors.initials: W
authors.lastname: Rangsipanuratn
mesh_terms: D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics*; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000145:classification* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*
chemical_list: D004269:DNA, Bacterial