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. 1997 Mar;35(3):719–723. doi: 10.1128/jcm.35.3.719-723.1997

Genotypic assessment of isoniazid and rifampin resistance in Mycobacterium tuberculosis: a blind study at reference laboratory level.

A Telenti 1, N Honoré 1, C Bernasconi 1, J March 1, A Ortega 1, B Heym 1, H E Takiff 1, S T Cole 1
PMCID: PMC229657  PMID: 9041419

Abstract

Progress in understanding the basis of resistance to isoniazid (INH) and rifampin (RMP) has allowed molecular tests for the detection of drug-resistant tuberculosis to be developed. Consecutive isolates (n = 95) of Mycobacterium tuberculosis, from a Spanish reference laboratory investigating outbreaks of multidrug-resistant tuberculosis, were coded and sent to two external laboratories for genotypic analysis of INH and RMP resistance by PCR-single-strand conformation polymorphism (SSCP) analysis of specific regions of four genes: part of the coding sequence of katG and the promoter regions of inhA and ahpC for INH and the RMP resistance region of rpoB. After correction for the presence of outbreak strains and multiple isolates from single patients, RMP resistance was detected successfully by PCR-SSCP in > 96% of the RMP-resistant strains. PCR-SSCP had a sensitivity of 87% for INH resistance detection, and mutations in katG, inhA, katG-inhA, ahpC, and katG-ahpC were identified in 36.8, 31.6, 2.6, 13.2, and 2.6%, respectively, of the unique strains. Specificity was 100%. Molecular detection of resistance to the two main antituberculous drugs, INH and RMP, can be accomplished accurately by using a strategy which limits analysis to four genetic regions. This may allow the expedient analysis of drug resistance by reference laboratories.

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Selected References

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  1. Banerjee A., Dubnau E., Quemard A., Balasubramanian V., Um K. S., Wilson T., Collins D., de Lisle G., Jacobs W. R., Jr inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosis. Science. 1994 Jan 14;263(5144):227–230. doi: 10.1126/science.8284673. [DOI] [PubMed] [Google Scholar]
  2. Cole S. T. Mycobacterium tuberculosis: drug-resistance mechanisms. Trends Microbiol. 1994 Oct;2(10):411–415. doi: 10.1016/0966-842x(94)90621-1. [DOI] [PubMed] [Google Scholar]
  3. Cole S. T. Rifamycin resistance in mycobacteria. Res Microbiol. 1996 Jan-Feb;147(1-2):48–52. doi: 10.1016/0923-2508(96)80203-8. [DOI] [PubMed] [Google Scholar]
  4. Cooksey R. C., Morlock G. P., McQueen A., Glickman S. E., Crawford J. T. Characterization of streptomycin resistance mechanisms among Mycobacterium tuberculosis isolates from patients in New York City. Antimicrob Agents Chemother. 1996 May;40(5):1186–1188. doi: 10.1128/aac.40.5.1186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. De Beenhouwer H., Lhiang Z., Jannes G., Mijs W., Machtelinckx L., Rossau R., Traore H., Portaels F. Rapid detection of rifampicin resistance in sputum and biopsy specimens from tuberculosis patients by PCR and line probe assay. Tuber Lung Dis. 1995 Oct;76(5):425–430. doi: 10.1016/0962-8479(95)90009-8. [DOI] [PubMed] [Google Scholar]
  6. Deretic V., Philipp W., Dhandayuthapani S., Mudd M. H., Curcic R., Garbe T., Heym B., Via L. E., Cole S. T. Mycobacterium tuberculosis is a natural mutant with an inactivated oxidative-stress regulatory gene: implications for sensitivity to isoniazid. Mol Microbiol. 1995 Sep;17(5):889–900. doi: 10.1111/j.1365-2958.1995.mmi_17050889.x. [DOI] [PubMed] [Google Scholar]
  7. Heym B., Alzari P. M., Honoré N., Cole S. T. Missense mutations in the catalase-peroxidase gene, katG, are associated with isoniazid resistance in Mycobacterium tuberculosis. Mol Microbiol. 1995 Jan;15(2):235–245. doi: 10.1111/j.1365-2958.1995.tb02238.x. [DOI] [PubMed] [Google Scholar]
  8. Heym B., Honoré N., Truffot-Pernot C., Banerjee A., Schurra C., Jacobs W. R., Jr, van Embden J. D., Grosset J. H., Cole S. T. Implications of multidrug resistance for the future of short-course chemotherapy of tuberculosis: a molecular study. Lancet. 1994 Jul 30;344(8918):293–298. doi: 10.1016/s0140-6736(94)91338-2. [DOI] [PubMed] [Google Scholar]
  9. Kapur V., Li L. L., Hamrick M. R., Plikaytis B. B., Shinnick T. M., Telenti A., Jacobs W. R., Jr, Banerjee A., Cole S., Yuen K. Y. Rapid Mycobacterium species assignment and unambiguous identification of mutations associated with antimicrobial resistance in Mycobacterium tuberculosis by automated DNA sequencing. Arch Pathol Lab Med. 1995 Feb;119(2):131–138. [PubMed] [Google Scholar]
  10. Musser J. M., Kapur V., Williams D. L., Kreiswirth B. N., van Soolingen D., van Embden J. D. Characterization of the catalase-peroxidase gene (katG) and inhA locus in isoniazid-resistant and -susceptible strains of Mycobacterium tuberculosis by automated DNA sequencing: restricted array of mutations associated with drug resistance. J Infect Dis. 1996 Jan;173(1):196–202. doi: 10.1093/infdis/173.1.196. [DOI] [PubMed] [Google Scholar]
  11. Ristow M., Möhlig M., Rifai M., Schatz H., Feldmann K., Pfeiffer A. New isoniazid/ethionamide resistance gene mutation and screening for multidrug-resistant Mycobacterium tuberculosis strains. Lancet. 1995 Aug 19;346(8973):502–503. doi: 10.1016/s0140-6736(95)91351-3. [DOI] [PubMed] [Google Scholar]
  12. Rouse D. A., Li Z., Bai G. H., Morris S. L. Characterization of the katG and inhA genes of isoniazid-resistant clinical isolates of Mycobacterium tuberculosis. Antimicrob Agents Chemother. 1995 Nov;39(11):2472–2477. doi: 10.1128/aac.39.11.2472. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Scorpio A., Zhang Y. Mutations in pncA, a gene encoding pyrazinamidase/nicotinamidase, cause resistance to the antituberculous drug pyrazinamide in tubercle bacillus. Nat Med. 1996 Jun;2(6):662–667. doi: 10.1038/nm0696-662. [DOI] [PubMed] [Google Scholar]
  14. Sherman D. R., Sabo P. J., Hickey M. J., Arain T. M., Mahairas G. G., Yuan Y., Barry C. E., 3rd, Stover C. K. Disparate responses to oxidative stress in saprophytic and pathogenic mycobacteria. Proc Natl Acad Sci U S A. 1995 Jul 3;92(14):6625–6629. doi: 10.1073/pnas.92.14.6625. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Stoeckle M. Y., Guan L., Riegler N., Weitzman I., Kreiswirth B., Kornblum J., Laraque F., Riley L. W. Catalase-peroxidase gene sequences in isoniazid-sensitive and -resistant strains of Mycobacterium tuberculosis from New York City. J Infect Dis. 1993 Oct;168(4):1063–1065. doi: 10.1093/infdis/168.4.1063. [DOI] [PubMed] [Google Scholar]
  16. Takiff H. E., Salazar L., Guerrero C., Philipp W., Huang W. M., Kreiswirth B., Cole S. T., Jacobs W. R., Jr, Telenti A. Cloning and nucleotide sequence of Mycobacterium tuberculosis gyrA and gyrB genes and detection of quinolone resistance mutations. Antimicrob Agents Chemother. 1994 Apr;38(4):773–780. doi: 10.1128/aac.38.4.773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Telenti A., Imboden P., Marchesi F., Lowrie D., Cole S., Colston M. J., Matter L., Schopfer K., Bodmer T. Detection of rifampicin-resistance mutations in Mycobacterium tuberculosis. Lancet. 1993 Mar 13;341(8846):647–650. doi: 10.1016/0140-6736(93)90417-f. [DOI] [PubMed] [Google Scholar]
  18. Telenti A., Imboden P., Marchesi F., Schmidheini T., Bodmer T. Direct, automated detection of rifampin-resistant Mycobacterium tuberculosis by polymerase chain reaction and single-strand conformation polymorphism analysis. Antimicrob Agents Chemother. 1993 Oct;37(10):2054–2058. doi: 10.1128/aac.37.10.2054. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Telenti A., Marchesi F., Balz M., Bally F., Böttger E. C., Bodmer T. Rapid identification of mycobacteria to the species level by polymerase chain reaction and restriction enzyme analysis. J Clin Microbiol. 1993 Feb;31(2):175–178. doi: 10.1128/jcm.31.2.175-178.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Telenti A., Persing D. H. Novel strategies for the detection of drug resistance in Mycobacterium tuberculosis. Res Microbiol. 1996 Jan-Feb;147(1-2):73–79. doi: 10.1016/0923-2508(96)80207-5. [DOI] [PubMed] [Google Scholar]
  21. Torrea G., Offredo C., Simonet M., Gicquel B., Berche P., Pierre-Audigier C. Evaluation of tuberculosis transmission in a community by 1 year of systematic typing of Mycobacterium tuberculosis clinical isolates. J Clin Microbiol. 1996 May;34(5):1043–1049. doi: 10.1128/jcm.34.5.1043-1049.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wilson T. M., Collins D. M. ahpC, a gene involved in isoniazid resistance of the Mycobacterium tuberculosis complex. Mol Microbiol. 1996 Mar;19(5):1025–1034. doi: 10.1046/j.1365-2958.1996.449980.x. [DOI] [PubMed] [Google Scholar]
  23. Zhang Y., Heym B., Allen B., Young D., Cole S. The catalase-peroxidase gene and isoniazid resistance of Mycobacterium tuberculosis. Nature. 1992 Aug 13;358(6387):591–593. doi: 10.1038/358591a0. [DOI] [PubMed] [Google Scholar]

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