Tuberc Respir Dis.  2007 Aug;63(2):128-138. 10.4046/trd.2007.63.2.128.

Mutations of katG and inhA in MDR M. tuberculosis

• Lin HH
• Kim HY
• Yun YJ
• Park CG
• Kim BJ
• Park YG
• Kook YH

• KMID: 1910091
• DOI: http://doi.org/10.4046/trd.2007.63.2.128

Abstract

BACKGROUNDS: Mutations of katG and inhA (ORF and promoter) are known to be related to isoniazid (INH) resistance of Mycobacterium tuberculosis. Because reports on these mutations in Korean isolates are limited (i.e. only the frequency of katG codon 463 was evaluated.), we tried to know the kinds of mutations of two genes and their frequencies in INH resistant Korean M. tuberculosis strains.
METHODS
PCR was performed to amplify katG (2,223 bp), inhA ORF (-77~897, 975 bp), and inhA promoter (-168~80, 248 bp) from 29 multidrug resistant M. tuberculosis (MDR-TB) DNAs prepared by bead beater-phenol method. Their sequences were determined and analyzed by ABI PRISM 3730 XL Analyzer and MegAlign package program, respectively.
RESULTS
All of the isolates had more than one mutation in katG or inhA gene. Twenty seven (93%) of 29 tested strains had katG mutations, which suggests that katG is a critical gene determining INH resistance of M. tuberculosis. Amino acid substitutions, such as Arg463Leu and Ser315Thr, due to point mutations of the katG were the most frequent (62.1% and 55.2%) mutations. In addition, deletion of the katG gene was frequently observed (17.2%). Analyzed Korean MDR-TB isolates also had variable inhA mutations. Point mutation of inhA promoter region, such as -15 (C-->T) was frequently found. Substitution of amino acid (Lsy8Asn) due to point mutation (AAA-->AAC) of inhA ORF was found in 1 isolate. Interestingly, 14 point mutated types that were not previously reported were newly found. While four types resulted in amino acid change, the others were silent mutations.
CONCLUSIONS
Although it is not clear that the relationship of these newly found mutations with INH resistance, they show marked diversity in Korean MDR-TB strains. It also suggests their feasibility as a molecular target to supplement determining the INH resistance of clinical isolates because of the possible existence of low-level INH resistant strains.

Keyword

M. tuberculosis; MDR-TB; INH; katG; inhA; Mutation

MeSH Terms

Amino Acid Substitution
Animals
Codon
DNA
Ecthyma, Contagious
Isoniazid
Mycobacterium tuberculosis
Point Mutation
Polymerase Chain Reaction
Promoter Regions, Genetic
Tuberculosis*
Codon
DNA
Isoniazid

Figure

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  Figure 1 Location of the primers on the katG gene

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  Figure 2 Location of the primers on the inhA gene

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  Figure 3 Diagram showing the distribution of mutant strains and their frequencies

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