International journal of medical sciences · 2012
Rapid detection of rpoB mutations in rifampin resistant M. tuberculosis from sputum samples by denaturing gradient gel electrophoresis.
Оценочное исследованиеСтатьяВалидационное исследованиеНазвание и аннотация: Перевод пока не подготовлен
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Li Jun, Xin Jiaojiao, Zhang Liyuan, Jiang Longyan, Cao Hongcui, Li Lanjuan
Аннотация
OBJECTIVE: To establish a rapid detection method for identifying rpoB mutations associated with rifampin (RIF) resistance in sputum specimens.
METHODS: We detected rpoB mutations directly in 90 sputum specimens collected from suspected tuberculosis patients using PCR-based denaturing gradient gel electrophoresis (DGGE) and compared these results with those obtained by rpoB sequencing and conventional drug susceptibility testing.
RESULTS: The positive detection rate of Mycobacterium tuberculosis (M. tuberculosis) was 52.2% by Acid-Fast Bacilli staining and 72.2% by conventional mycobacterial culture. In contrast, the positive rate was significantly higher (93.3%) by PCR-based detection of the rpoB gene in the same specimens. Furthermore, 75% of the tested specimens presented abnormal patterns compared with the wild-type pattern (standard H37Rv strain) analysed by DGGE. A total of 12 different patterns, representing 12 different rpoB mutations, were observed in the 63 abnormal patterns. The match rate of rpoB mutations detected by DGGE reached 96.9% when compared to DNA sequencing.
CONCLUSION: Our findings indicate that PCR-based DGGE is a rapid and reliable bio-technique for direct detection of rpoB mutations associated with RIF resistance in the sputum of suspected tuberculosis patients.
METHODS: We detected rpoB mutations directly in 90 sputum specimens collected from suspected tuberculosis patients using PCR-based denaturing gradient gel electrophoresis (DGGE) and compared these results with those obtained by rpoB sequencing and conventional drug susceptibility testing.
RESULTS: The positive detection rate of Mycobacterium tuberculosis (M. tuberculosis) was 52.2% by Acid-Fast Bacilli staining and 72.2% by conventional mycobacterial culture. In contrast, the positive rate was significantly higher (93.3%) by PCR-based detection of the rpoB gene in the same specimens. Furthermore, 75% of the tested specimens presented abnormal patterns compared with the wild-type pattern (standard H37Rv strain) analysed by DGGE. A total of 12 different patterns, representing 12 different rpoB mutations, were observed in the 63 abnormal patterns. The match rate of rpoB mutations detected by DGGE reached 96.9% when compared to DNA sequencing.
CONCLUSION: Our findings indicate that PCR-based DGGE is a rapid and reliable bio-technique for direct detection of rpoB mutations associated with RIF resistance in the sputum of suspected tuberculosis patients.