Clinical efficacy of direct DNA sequencing analysis on sputum specimens for early detection of drug-resistant Mycobacterium tuberculosis in a clinical setting.
BACKGROUND
Early detection of drug-resistant Mycobacterium tuberculosis is important for the control and prevention of disease transmission. However, conventional drug susceptibility tests for drug-resistant M tuberculosis take at least 3 to 8 weeks. Here, we report the clinical efficacy of direct DNA sequencing analysis for detecting drug-resistant TB on sputum specimens in a clinical setting.


METHODS
A total of 113 sputum specimens from 111 patients, who were suspected of having drug-resistant TB by clinicians, were used for DNA sequencing of katG, rpoB, embB, and pncA genes for isoniazid (INH), rifampin (RIF), ethambutol (EMB), and pyrazinamide (PZA) resistance, respectively, and the results were compared with drug susceptibility tests. The optimization of antituberculosis drugs according to the results of DNA sequencing and the treatment outcomes of the patients were also analyzed.


RESULTS
Turnaround time of the direct DNA sequencing analysis was 3.8 +/- 1.8 days. We found mutations related to drug resistance in 30 clinical specimens for katG, 39 for rpoB, 13 for embB, and 24 for pncA. The sensitivity and specificity of the assay were 63.6% and 94.6% for INH, 96.2 and 93.9% for RIF, 69.2% and 97.5% for EMB, and 100% and 92.6% for PZA, respectively. Of the patients with RIF resistance, including multidrug-resistant TB by the assay, 92.5% of the patients with initial first-line antituberculosis drugs were changed to second-line antituberculosis drugs, and treatment was successful in 61.9% of these cases.


CONCLUSIONS
Direct DNA sequencing analysis of clinical sputum specimens is a rapid and useful method for the detection and treatment of drug-resistant TB.
Choi
Jeong-Hee
JH
Division of Pulmonary, Allergy and Critical Care Medicine, Department of Internal Medicine, Hallym University Medical Center, Gangwon-do, Korea.
Lee
Kyung Wha
KW
Kang
Hye-Ryun
HR
Hwang
Yong Il
YI
Jang
SeungHun
S
Kim
Dong-Gyu
DG
Kim
Cheol-Hong
CH
Hyun
In-Gyu
IG
Shin
Tae-Rim
TR
Park
Sang-Myeon
SM
Lee
Myung-Goo
MG
Lee
Chang-Youl
CY
Park
Yong-Bum
YB
Jung
Ki-Suck
KS
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000581:Amidohydrolases / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D002374:Catalase / Q000235:genetics; D004252:DNA Mutational Analysis / Q000379:methods*; D004269:DNA, Bacterial / Q000032:analysis*; D012321:DNA-Directed RNA Polymerases; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D010430:Pentosyltransferases / Q000235:genetics; D016133:Polymerase Chain Reaction; D012189:Retrospective Studies; D012680:Sensitivity and Specificity; D013183:Sputum / Q000382:microbiology*; D018088:Tuberculosis, Multidrug-Resistant / Q000175:diagnosis* / Q000382:microbiology; D055815:Young Adult
D001426:Bacterial Proteins; D004269:DNA, Bacterial; C505805:rpoB protein, Mycobacterium tuberculosis; D002374:Catalase; C480353:katG protein, Mycobacterium tuberculosis; C540269:EmbB protein, Mycobacterium tuberculosis; D010430:Pentosyltransferases; D012321:DNA-Directed RNA Polymerases; D000581:Amidohydrolases; C487496:PncA protein, Mycobacterium tuberculosis