Detection and genotyping of Mycobacterium species from clinical isolates and specimens by oligonucleotide array.
Identification of pathogenic Mycobacterium species is important for a successful diagnosis of mycobacteriosis. The purpose of this study was to develop an oligonucleotide array which could detect and differentiate mycobacteria to the species level by using the internal transcribed spacer (ITS) sequence. Using a genus-specific probe and 20 species-specific probes including two M. avium-intracellulare complex (MAC)-specific probes, we have developed an ITS-based oligonucleotide array for the rapid and reliable detection and discrimination of M. tuberculosis, MAC, M. fortuitum, M. chelonae, M. abscessus, M. kansasii, M. gordonae, M. scrofulaceum, M. szulgai, M. vaccae, M. xenopi, M. terrae, M. flavescens, M. smegmatis, M. malmoense, M. simiae, M. marinum, M. ulcerans, M. gastri, and M. leprae. All mycobacteria were hybridized with a genus-specific probe (PAN-03) for detection of the genus Mycobacterium. Mycobacterial species were expected to show a unique hybridization pattern with species-specific probes, except for M. marinum and M. ulcerans, which were not differentiated by ITS-based probe. Among the species-specific probes, two kinds of species-specific probes were designed for MAC in which there were many subspecies. The performance of the oligonucleotide array assay was demonstrated by using 46 reference strains, 149 clinical isolates, and 155 clinical specimens. The complete procedure (DNA extraction, PCR, DNA hybridization, and scanning) was carried out in 4.5 h. Our results indicated that the oligonucleotide array is useful for the identification and discrimination of mycobacteria from clinical isolates and specimens in an ordinary clinical laboratory.
Park
Heekyung
H
Department of Biochemistry, College of Medicine, Pusan National University, #10 Ami-Dong 1-Ga Seo-Gu, Busan 602-739, Korea.
Jang
Hyunjung
H
Song
Eunsil
E
Chang
Chulhun L
CL
Lee
Minki
M
Jeong
Seokhoon
S
Park
Junhyung
J
Kang
Byeongchul
B
Kim
Cheolmin
C
D015373:Bacterial Typing Techniques; D015342:DNA Probes; D021903:DNA, Ribosomal Spacer / Q000032:analysis*; D005838:Genotype; D006801:Humans; D009161:Mycobacterium / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D009164:Mycobacterium Infections / Q000382:microbiology*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012015:Reference Standards; D012680:Sensitivity and Specificity; D013045:Species Specificity
D015342:DNA Probes; D021903:DNA, Ribosomal Spacer