Use of fluorescence resonance energy transfer for rapid detection of isoniazid resistance in Mycobacterium tuberculosis clinical isolates.
METHODS
Rapid detection of drug resistance in Mycobacterium tuberculosis is important to select effective treatment and prevent transmission of resistant isolates.


OBJECTIVE
To evaluate the use of fluorescence resonance energy transfer (FRET) for rapid detection of isoniazid (INH) resistance in M. tuberculosis clinical isolates.


METHODS
One hundred INH-resistant and 50 INH-susceptible isolates of M. tuberculosis were included in the study. The drug susceptibility of all isolates was determined by the standard agar proportion method, and all isolates were then tested by FRET. Three genes associated with INH resistance, katG, inhA and ahpC, were analysed. All isolates were amplified with three pairs of primers. Three pairs of fluorescently labelled DNA probes specific to codon 315 of katG, nucleotide 209 in the regulatory region of inhA and a frequent mutation site in the intergenic region of oxyR-ahpC, were used for mutation detection.


RESULTS
The results obtained using FRET were compared with those from the proportion method. The sensitivity and specificity of FRET were respectively 76% and 100%. The frequencies of mutations were 48% in katG, 17% in inhA, 8% in ahpC, 2% in inhA-ahpC and 1% in inhA-katG.


CONCLUSIONS
FRET is a rapid, specific method that can be useful to detect INH resistance in M. tuberculosis clinical isolates.
Saribas
Z
Z
Department of Microbiology and Clinical Microbiology, Hacettepe University Faculty of Medicine, Ankara, Turkey. zsaribas@superonline.com
Yurdakul
P
P
Alp
A
A
Gunalp
A
A
D024881:Drug Resistance, Bacterial / Q000235:genetics; D031541:Fluorescence Resonance Energy Transfer; D007538:Isoniazid / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D007538:Isoniazid