[The 69th annual meeting president lecture. Application of molecular biology for rapid detection of mycobacteria].
The application of molecular biology techniques for the rapid detection of mycobacteria in clinical specimens was evaluated. The DNA probe method was found to be accurate and rapid for identification of mycobacteria, but its low sensitivity needs improving. The nested polymerase chain reaction (PCR) targeting Pab gene was specific and sensitive enough for rapid detection of Mycobacterium tuberculosis in clinical specimens. The overall sensitivity and specificity of this method were excellent, 97% and 92%, respectively. Additionally, a novel method for the rapid detection of mycobacteria other than M. tuberculosis was developed using a combined method of PCR and DNA probe. The gene encoding 16S ribosomal RNA in mycobacteria was first amplified by PCR, and then PCR products were identified using an acridinium-ester labeled DNA probe. This method was as useful as that of the PCR for M. tuberculosis described above. The DNA probe method was also applied for drug sensitivity testing, which needs three to four weeks by conventional methods. The quantity of ribosomal RNA in mycobacteria cultured with or without the addition of drugs was compared. Three days after the start of culture, the strains sensitive for isoniazid or rifampicin showed remarkable decreases of ribosomal RNA, compared with the drug-free samples, while the resistant strains showed no difference. Another method for determining the drug resistance of mycobacteria consists of detection of the gene which is related to the resistance. The deletion of catalase-peroxidase gene suggested a gene related to isoniazid resistance was observed in 15% of isoniazid-resistant strains.(ABSTRACT TRUNCATED AT 250 WORDS)
Hara
K
K
Second Department of Internal Medicine, Nagasaki University School of Medicine, Japan.
D004269:DNA, Bacterial / Q000032:analysis; D004352:Drug Resistance, Microbial / Q000235:genetics; D006801:Humans; D009161:Mycobacterium / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial