Rapid detection of rifampin-resistant clinical isolates of Mycobacterium tuberculosis by reverse dot blot hybridization.

Qian Guo; Yan YU; Yan Ling Zhu; Xiu Qin Zhao; Zhi Guang Liu; Yuan Yuan Zhang; Gui Lian LI; Jian Hao Wei; Yi Mou WU; Kang Lin Wan

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Rapid detection of rifampin-resistant clinical isolates of Mycobacterium tuberculosis by reverse dot blot hybridization.

Author: Qian GUO ^{1
,

2
,

3} ; Yan YU ⁴ ; Yan Ling ZHU ⁵ ; Xiu Qin ZHAO ⁴ ; Zhi Guang LIU ⁴ ; Yuan Yuan ZHANG ⁴ ; Gui Lian LI ⁴ ; Jian Hao WEI ⁴ ; Yi Mou WU ⁶ ; Kang Lin WAN ¹ ;
Author Information

1. Pathogenic Biology Institute, University of South China, Hengyang 421000, Hunan, China
2. State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
3. Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Hangzhou 310003, Zhejiang, China.
4. State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
5. State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China.
6. Pathogenic Biology Institute, University of South China, Hengyang 421000, Hunan, China.
Publication Type:Journal Article
Keywords: Mycobacterium tuberculosis; Reverse dot blot hybridization; Rifampin-resistance
MeSH: Antitubercular Agents; pharmacology; Drug Resistance, Bacterial; Genotype; Immunoblotting; methods; Microbial Sensitivity Tests; Mycobacterium tuberculosis; drug effects; genetics; Polymerase Chain Reaction; methods; Rifampin; pharmacology; Sensitivity and Specificity; Time Factors
From: Biomedical and Environmental Sciences 2015;28(1):25-35
CountryChina
Language:English
Abstract:
OBJECTIVEA PCR-reverse dot blot hybridization (RDBH) assay was developed for rapid detection of rpoB gene mutations in 'hot mutation region' of Mycobacterium tuberculosis (M. tuberculosis).
METHODS12 oligonucleotide probes based on the wild-type and mutant genotype rpoB sequences of M. tuberculosis were designed to screen the most frequent wild-type and mutant genotypes for diagnosing RIF resistance. 300 M. tuberculosis clinical isolates were detected by RDBH, conventional drug-susceptibility testing (DST) and DNA sequencing to evaluate the RDBH assay.
RESULTSThe sensitivity and specificity of the RDBH assay were 91.2% (165/181) and 98.3% (117/119), respectively, as compared to DST. When compared with DNA sequencing, the accuracy, positive predictive value (PPV) and negative predictive value (NPV) of the RDBH assay were 97.7% (293/300), 98.2% (164/167), and 97.0% (129/133), respectively. Furthermore, the results indicated that the most common mutations were in codons 531 (48.6%), 526 (25.4%), 516 (8.8%), and 511 (6.6%), and the combinative mutation rate was 15 (8.3%). One and two strains of insertion and deletion were found among all strains, respectively.
CONCLUSIONOur findings demonstrate that the RDBH assay is a rapid, simple and sensitive method for diagnosing RIF-resistant tuberculosis.