Evaluation of the invader assay with the BACTEC MGIT 960 system for prompt isolation and identification of mycobacterial species from clinical specimens.
Rapid and accurate identification of mycobacterial species is essential for patient management. We describe the use of the Invader assay in conjunction with the BACTEC MGIT 960 system that together provide an efficient procedure for clinical use. This assay discriminates single-base differences (e.g., genotyping single-nucleotide polymorphisms) under homogeneous and isothermal conditions and can measure directly on genomic DNA without prior target DNA amplification. To identify a wide variety of mycobacterial species, 20 Invader probes were designed to target the 16S rRNA gene and the 16S-23S rRNA gene internal transcribed spacer 1 (ITS-1) region. To validate the Invader probes, we used 78 ATCC strains, and 607 clinical mycobacterial strains, which were identified by DNA sequencing of the 16S rRNA gene and ITS-1. The Invader assay could accurately identify and differentiate these strains according to target sequences. Moreover, it could detect and identify 116 (95.1%) of 122 positive liquid cultures from the BACTEC MGIT 960 system and did not react to 83 contaminated MGIT cultures. Species identification takes 6.5 h by the Invader assay: 2.0 h for DNA extraction, 0.5 h for handling, and up to 4 h for the Invader reaction. The Invader assay has the speed, ease of use, and accuracy to be an effective procedure for the bacteriological diagnosis of mycobacterial infections.
Ichimura
Sadahiro
S
Department of Microbiology, BML, Inc., 1361-1 Matoba, Kawagoe, Saitama 350-1101, Japan. ichi-s@bml.co.jp
Nagano
Makoto
M
Ito
Nobuko
N
Shimojima
Masahiro
M
Egashira
Toru
T
Miyamoto
Chikara
C
Ohkusu
Kiyofumi
K
Ezaki
Takayuki
T
D004269:DNA, Bacterial / Q000737:chemistry*; D006801:Humans; D009161:Mycobacterium / Q000235:genetics / Q000302:isolation & purification*; D020641:Polymorphism, Single Nucleotide; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods*
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S