Skip to main content
NCBI home page
Journal of Clinical Microbiology logoLink to Journal of Clinical Microbiology
. 1997 Dec;35(12):3225–3231. doi: 10.1128/jcm.35.12.3225-3231.1997

Specific identification of Mycobacterium tuberculosis with the luciferase reporter mycobacteriophage: use of p-nitro-alpha-acetylamino-beta-hydroxy propiophenone.

P F Riska 1, W R Jacobs Jr 1, B R Bloom 1, J McKitrick 1, J Chan 1
PMCID: PMC230152  PMID: 9399524

Abstract

We have previously described a luciferase reporter mycobacteriophage (LRP) assay that can detect Mycobacterium tuberculosis and characterize mycobacterial drug susceptibility patterns within 24 to 48 h in positive cultures. One drawback of this LRP protocol is the ability of the recombinant mycobacteriophage phAE40 to infect a variety of Mycobacterium species, thus limiting its specificity for the detection of M. tuberculosis. In this study, we have (i) explored the host range of phAE40, (ii) developed a modified LRP assay that exploits the selective inhibitory effect of the compound p-nitro-alpha-acetylamino-beta-hydroxy propiophenone (NAP) against members of the M. tuberculosis complex to differentiate between the tubercle bacillus and other mycobacterial species, and (iii) tested over 300 samples, including primary clinical isolates and drug-resistant strains of M. tuberculosis, demonstrating the ability of the NAP-modified LRP assay to identify M. tuberculosis complex organisms with high degrees of sensitivity and specificity.

Full Text

The Full Text of this article is available as a PDF (183.3 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barnes P. F., Bloch A. B., Davidson P. T., Snider D. E., Jr Tuberculosis in patients with human immunodeficiency virus infection. N Engl J Med. 1991 Jun 6;324(23):1644–1650. doi: 10.1056/NEJM199106063242307. [DOI] [PubMed] [Google Scholar]
  2. Barsom E. K., Hatfull G. F. Characterization of Mycobacterium smegmatis gene that confers resistance to phages L5 and D29 when overexpressed. Mol Microbiol. 1996 Jul;21(1):159–170. doi: 10.1046/j.1365-2958.1996.6291342.x. [DOI] [PubMed] [Google Scholar]
  3. Benson C. A., Ellner J. J. Mycobacterium avium complex infection and AIDS: advances in theory and practice. Clin Infect Dis. 1993 Jul;17(1):7–20. doi: 10.1093/clinids/17.1.7. [DOI] [PubMed] [Google Scholar]
  4. Bodmer T., Gurtner A., Schopfer K., Matter L. Screening of respiratory tract specimens for the presence of Mycobacterium tuberculosis by using the Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test. J Clin Microbiol. 1994 Jun;32(6):1483–1487. doi: 10.1128/jcm.32.6.1483-1487.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Body B. A., Warren N. G., Spicer A., Henderson D., Chery M. Use of Gen-Probe and Bactec for rapid isolation and identification of mycobacteria. Correlation of probe results with growth index. Am J Clin Pathol. 1990 Mar;93(3):415–420. doi: 10.1093/ajcp/93.3.415. [DOI] [PubMed] [Google Scholar]
  6. Bradley S. P., Reed S. L., Catanzaro A. Clinical efficacy of the amplified Mycobacterium tuberculosis direct test for the diagnosis of pulmonary tuberculosis. Am J Respir Crit Care Med. 1996 May;153(5):1606–1610. doi: 10.1164/ajrccm.153.5.8630609. [DOI] [PubMed] [Google Scholar]
  7. Carrière C., Riska P. F., Zimhony O., Kriakov J., Bardarov S., Burns J., Chan J., Jacobs W. R., Jr Conditionally replicating luciferase reporter phages: improved sensitivity for rapid detection and assessment of drug susceptibility of Mycobacterium tuberculosis. J Clin Microbiol. 1997 Dec;35(12):3232–3239. doi: 10.1128/jcm.35.12.3232-3239.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chan J., Xing Y., Magliozzo R. S., Bloom B. R. Killing of virulent Mycobacterium tuberculosis by reactive nitrogen intermediates produced by activated murine macrophages. J Exp Med. 1992 Apr 1;175(4):1111–1122. doi: 10.1084/jem.175.4.1111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Counsell S. R., Tan J. S., Dittus R. S. Unsuspected pulmonary tuberculosis in a community teaching hospital. Arch Intern Med. 1989 Jun;149(6):1274–1278. [PubMed] [Google Scholar]
  10. 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]
  11. EIDUS L., DIENA B. B., GREENBERG L. The use of p-nitro-alpha-acetylamino-beta-hydroxy-propiophenone (NAP) in the differentiation of mycobacteria. Am Rev Respir Dis. 1960 May;81:759–760. doi: 10.1164/arrd.1960.81.5.759. [DOI] [PubMed] [Google Scholar]
  12. Falkinham J. O., 3rd Epidemiology of infection by nontuberculous mycobacteria. Clin Microbiol Rev. 1996 Apr;9(2):177–215. doi: 10.1128/cmr.9.2.177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Foley-Thomas E. M., Whipple D. L., Bermudez L. E., Barletta R. G. Phage infection, transfection and transformation of Mycobacterium avium complex and Mycobacterium paratuberculosis. Microbiology. 1995 May;141(Pt 5):1173–1181. doi: 10.1099/13500872-141-5-1173. [DOI] [PubMed] [Google Scholar]
  14. Good R. C., Snider D. E., Jr Isolation of nontuberculous mycobacteria in the United States, 1980. J Infect Dis. 1982 Dec;146(6):829–833. doi: 10.1093/infdis/146.6.829. [DOI] [PubMed] [Google Scholar]
  15. Gross W. M., Hawkins J. E. Radiometric selective inhibition tests for differentiation of Mycobacterium tuberculosis, Mycobacterium bovis, and other mycobacteria. J Clin Microbiol. 1985 Apr;21(4):565–568. doi: 10.1128/jcm.21.4.565-568.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hobby G. L., Holman A. P., Iseman M. D., Jones J. M. Enumeration of tubercle bacilli in sputum of patients with pulmonary tuberculosis. Antimicrob Agents Chemother. 1973 Aug;4(2):94–104. doi: 10.1128/aac.4.2.94. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jacobs W. R., Jr, Barletta R. G., Udani R., Chan J., Kalkut G., Sosne G., Kieser T., Sarkis G. J., Hatfull G. F., Bloom B. R. Rapid assessment of drug susceptibilities of Mycobacterium tuberculosis by means of luciferase reporter phages. Science. 1993 May 7;260(5109):819–822. doi: 10.1126/science.8484123. [DOI] [PubMed] [Google Scholar]
  18. Kramer F., Modilevsky T., Waliany A. R., Leedom J. M., Barnes P. F. Delayed diagnosis of tuberculosis in patients with human immunodeficiency virus infection. Am J Med. 1990 Oct;89(4):451–456. doi: 10.1016/0002-9343(90)90375-n. [DOI] [PubMed] [Google Scholar]
  19. Laszlo A., Eidus L. Test for differentiation of M. tuberculosis and M. bovis from other mycobacteria. Can J Microbiol. 1978 Jun;24(6):754–756. doi: 10.1139/m78-124. [DOI] [PubMed] [Google Scholar]
  20. Laszlo A., Siddiqi S. H. Evaluation of a rapid radiometric differentiation test for the Mycobacterium tuberculosis complex by selective inhibition with p-nitro-alpha-acetylamino-beta-hydroxypropiophenone. J Clin Microbiol. 1984 May;19(5):694–698. doi: 10.1128/jcm.19.5.694-698.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Menzies D., Fanning A., Yuan L., Fitzgerald M. Tuberculosis among health care workers. N Engl J Med. 1995 Jan 12;332(2):92–98. doi: 10.1056/NEJM199501123320206. [DOI] [PubMed] [Google Scholar]
  22. Morgan M. A., Doerr K. A., Hempel H. O., Goodman N. L., Roberts G. D. Evaluation of the p-nitro-alpha-acetylamino-beta-hydroxypropiophenone differential test for identification of Mycobacterium tuberculosis complex. J Clin Microbiol. 1985 Apr;21(4):634–635. doi: 10.1128/jcm.21.4.634-635.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Peterson E. M., Lu R., Floyd C., Nakasone A., Friedly G., de la Maza L. M. Direct identification of Mycobacterium tuberculosis, Mycobacterium avium, and Mycobacterium intracellulare from amplified primary cultures in BACTEC media using DNA probes. J Clin Microbiol. 1989 Jul;27(7):1543–1547. doi: 10.1128/jcm.27.7.1543-1547.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rado T. A., Bates J. H., Fitzhugh J. K. Evidence for host-dependent modification and restriction of bacteriophage DNA in Mycobacterium tuberculosis. J Gen Virol. 1976 Jan;30(1):91–97. doi: 10.1099/0022-1317-30-1-91. [DOI] [PubMed] [Google Scholar]
  25. Sepkowitz K. A., Raffalli J., Riley L., Kiehn T. E., Armstrong D. Tuberculosis in the AIDS era. Clin Microbiol Rev. 1995 Apr;8(2):180–199. doi: 10.1128/cmr.8.2.180. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Siddiqi S. H., Hwangbo C. C., Silcox V., Good R. C., Snider D. E., Jr, Middlebrook G. Rapid radiometric methods to detect and differentiate Mycobacterium tuberculosis/M. bovis from other mycobacterial species. Am Rev Respir Dis. 1984 Oct;130(4):634–640. doi: 10.1164/arrd.1984.130.4.634. [DOI] [PubMed] [Google Scholar]
  27. 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]
  28. Turett G. S., Telzak E. E., Torian L. V., Blum S., Alland D., Weisfuse I., Fazal B. A. Improved outcomes for patients with multidrug-resistant tuberculosis. Clin Infect Dis. 1995 Nov;21(5):1238–1244. doi: 10.1093/clinids/21.5.1238. [DOI] [PubMed] [Google Scholar]
  29. de Wet J. R., Wood K. V., DeLuca M., Helinski D. R., Subramani S. Firefly luciferase gene: structure and expression in mammalian cells. Mol Cell Biol. 1987 Feb;7(2):725–737. doi: 10.1128/mcb.7.2.725. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES