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Journal of Clinical Microbiology, November 2001, p. 3883-3888, Vol. 39, No. 11
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.11.3883-3888.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Luciferase Reporter Mycobacteriophages for Detection, Identification, and Antibiotic Susceptibility Testing of Mycobacterium tuberculosis in Mexico

N. Banaiee,1,* M. Bobadilla-del-Valle,2 S. Bardarov Jr.,3 P. F. Riska,3 P. M. Small,1 A. Ponce-de-Leon,2 W. R. Jacobs Jr.,3 G. F. Hatfull,4 and J. Sifuentes-Osornio2

Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California1; Department of Infectious Diseases, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubiran, Mexico City, Mexico2; Howard Hughes Medical Institute, Albert Einstein College of Medicine, Bronx, New York3; and Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania4

Received 14 March 2001/Returned for modification 13 May 2001/Accepted 13 August 2001

The utility of luciferase reporter mycobacteriophages (LRPs) for detection, identification, and antibiotic susceptibility testing of Mycobacterium tuberculosis was prospectively evaluated in a clinical microbiology laboratory in Mexico City, Mexico. Five hundred twenty-three consecutive sputum samples submitted to the laboratory during a 5-month period were included in this study. These specimens were cultivated in Middlebrook 7H9 (MADC), MGIT, and Löwenstein-Jensen (LJ) media. Of the 71 mycobacterial isolates recovered with any of the three media, 76% were detected with the LRPs, 97% were detected with the MGIT 960 method, and 90% were detected with LJ medium. When contaminated specimens were excluded from the analysis, the LRPs detected 92% (54 of 59) of the cultures. The median time to detection of bacteria was 7 days with both the LRPs and the MGIT 960 method. LRP detection of growth in the presence of p-nitro-alpha -acetylamino-beta -hydroxypropiophenone (NAP) was used for selective identification of M. tuberculosis complex (MTC) and compared to identification with BACTEC 460. Using the LRP NAP test, 47 (94%) out of 50 isolates were correctly identified as tuberculosis complex. The accuracy and speed of LRP antibiotic susceptibility testing with rifampin, streptomycin, isoniazid, and ethambutol were compared to those of the BACTEC 460 method, and discrepant results were checked by the conventional proportion method. In total, 50 MTC isolates were tested. The overall agreement between the LRP and BACTEC 460 results was 98.5%. The median LRP-based susceptibility turnaround time was 2 days (range, 2 to 4 days) compared to 10.5 days (range, 7 to 16 days) by the BACTEC 460 method. Phage resistance was not detected in any of the 243 MTC isolates tested. Mycobacteriophage-based approaches to tuberculosis diagnostics can be implemented in clinical laboratories with sensitivity, specificity, and rapidity that compare favorably with those of the MGIT 960 and BACTEC 460 methods. The phages currently provide the fastest phenotypic assay for susceptibility testing.

* Corresponding author. Present address: Department of Laboratory Medicine, UCSF, L518 Box 0134, San Francisco, CA 94143-0134. Phone: (415) 502-5324. Fax: (415) 476-9625. E-mail: niaz{at}itsa.ucsf.edu.

Journal of Clinical Microbiology, November 2001, p. 3883-3888, Vol. 39, No. 11
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.11.3883-3888.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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Copyright © 2001 by the American Society for Microbiology. All rights reserved.