title: PCR restriction fragment length polymorphism analysis (PRA)-algorithm targeting 644 bp Heat Shock Protein 65 (hsp65) gene for differentiation of Mycobacterium spp.
abstract: A method based on PCR-restriction fragment length polymorphism analysis (PRA) using a novel region of the hsp65 gene was developed for the rapid and exact identification of mycobacteria to the species level. A 644 bp region of hsp65 in 62 mycobacteria reference strains, and 4 related bacterial strains were amplified, and the amplified DNAs were subsequently digested with restriction enzymes, namely, AvaII, HphI, and HpaII. Most of the mycobacteria species were easily differentiated at the species level by the developed method. In particular, the method enabled the separation of M. avium, M. intracellulare and M. tuberculosis to the species level by AvaII digestion alone. An algorithm was constructed based on the results and a blind test was successfully performed on 251 clinical isolates, which had been characterized by conventional biochemical testing. Our results suggest that this novel PRA offers a simple, rapid, and accurate method for the identification of mycobacteria culture isolates at the species level.
authors: Department of Microbiology and Liver Research Institute, College of Medicine, Seoul National University, 28 Yongon-dong, Chongno-gu, Seoul 110-799, South Korea.
authors: Hong
authors: H
authors: Kim
authors: Sun-Hyun
authors: SH
authors: Kim
authors: Tae-Sun
authors: TS
authors: Shim
authors: Mi-na
authors: MN
authors: Kim
authors: Gill-Han
authors: GH
authors: Bai
authors: Young-Gil
authors: YG
authors: Park
authors: Sueng-Hyun
authors: SH
authors: Lee
authors: Chang-Yong
authors: CY
authors: Cha
authors: Yoon-Hoh
authors: YH
authors: Kook
authors: Bum-Joon
authors: BJ
authors: Kim
mesh_terms: D000465:Algorithms; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics*; D018834:Chaperonin 60; D018833:Chaperonins / Q000737:chemistry / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D006801:Humans; D009161:Mycobacterium / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D009164:Mycobacterium Infections / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D017422:Sequence Analysis, DNA
chemical_list: D001426:Bacterial Proteins; D018834:Chaperonin 60; D004269:DNA, Bacterial; C074119:heat-shock protein 65, Mycobacterium; D018833:Chaperonins