[Study on isoniazid-resistant Mycobacterium tuberculosis isolates by multiple-polymerase chain reaction-single strand conformation polymorphism].
OBJECTIVE
To develop a new multiple-polymerase chain reaction-single strand conformation polymorphism (multi-PCR-SSCP) system for detecting the aphC promoter, inhA, and katG gene mutations in isoniazid-resistant Mycobacterium tuberculosis isolates in the single reaction, and for the quick diagnosis of isoniazid-resistant Mycobacterium tuberculosis isolates.


METHODS
Three pairs of oligonucleotide primers were designed according to the aphC promoter, inhA, and katG genes of Mycobacterium tuberculosis to examine isoniazid-resistance by multi-PCR-SSCP.


RESULTS
Isoniazid-sensitivity and resistance were analyzed with general PCR and multi-PCR at the same time, and H(37) Rv was used as a control. These two protocols amplified the anticipated fragments, the rate of consistency being 100%. By single gene PCR-SSCP, the mutation rates of aphC promoter, inhA, and katG gene were 17%, 20%, and 66%, respectively. The mutation rate detected by multi-PCR-SSCP was 83%.


CONCLUSIONS
Multi-PCR-SSCP is a sensitive and specific method for rapid detection of aphC promoter, inhA, and katG gene mutations in isoniazid-resistant Mycobacterium tuberculosis isolates. Drug-resistant gene detection may be clinically useful in the therapy of tuberculosis.
Cheng
Xiao-dong
XD
Department of Clinical Laboratories, Xijing Hospital, Fouth Military Medical University, Xi'an 710032, China.
Yu
Wen-bin
WB
Bie
Liang-feng
LF
Su
Ming-quan
MQ
Zhang
Rong
R
Hao
Xiao-ke
XK
D000995:Antitubercular Agents / Q000494:pharmacology; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D006801:Humans; D007538:Isoniazid / Q000494:pharmacology; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods; D018807:Polymorphism, Single-Stranded Conformational
D000995:Antitubercular Agents; D004269:DNA, Bacterial; D007538:Isoniazid