This Article
Right arrow Full Text (PDF)
Right arrow A correction has been published
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Radhakrishnan, I.
Right arrow Articles by Mundayoor, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Radhakrishnan, I.
Right arrow Articles by Mundayoor, S.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, April 2001, p. 1683-1683, Vol. 39, No. 4
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.4.1683.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

LETTERS TO THE EDITOR

Implications of Low Frequency of IS6110 in Fingerprinting Field Isolates of Mycobacterium tuberculosis from Kerala, India


    LETTER

Restriction fragment length polymorphism (RFLP) analysis of IS6110 in PvuII-digested genomic DNA has been advocated for the fingerprinting of Mycobacterium tuberculosis as an approved standard methodology (8). This has proven to be very useful in the developed world, including the tracing of the New York outbreak of multidrug-resistant M. tuberculosis (2). The success of the screening hinges on the extent and type of polymorphism of the isolates.

Kerala, a small state in southern India, has moderately good health care facilities and a high rate of tuberculosis. With a view to study IS6110 polymorphism, we screened 80 isolates obtained from patients at local tuberculosis clinics. The strains were isolated from sputum samples on L-J medium, acid-fast stained, and confirmed to be M. tuberculosis by standard biochemical tests. The DNA was isolated by the sand procedure (5) and further purified by phenol-chloroform extractions. The RFLP analysis was performed as described earlier (8) using the [alpha -32P]dCTP-labeled, amplified 245-bp fragment of IS6110. The strains showing an absence of IS6110 in the RFLP assay were confirmed to be IS6110 negative by PCR amplification of purified genomic DNA.

Among the 80 isolates, 19 had no copy of IS6110 and another 31 had only a single copy of the IS6110 sequence. All the single-copy isolates (except three) showed a band at the 1.3-kb position where an integration hot spot (ipl) has been reported earlier (4). Thus, 50 (62.5%) of the 80 strains were not typeable by IS6110-based fingerprinting. In addition to this, another eight isolates had only 2 to 5 copies, making definitive fingerprinting difficult. This study clearly shows that IS6110-based fingerprinting is not possible in Kerala. While small numbers of IS6110-deficient strains have been reported earlier (ranging from less than 1% in San Francisco [1] and 2% in Vietnam [6] to 4% in Chennai [3]), this study is the first of its kind to report a large number of IS6110-deficient strains while also demonstrating unequivocally the inability to use the only approved and accepted fingerprinting system for molecular epidemiology of tuberculosis at least in some areas where tuberculosis is endemic.

The ramifications of the study are manifold. There is a clear need for a usable fingerprinting system for areas where IS6110-deficient strains abound. While several typing systems such as polymorphic GC-rich sequence typing, variable number of tandem repeat typing, and spoligotyping (7) have been reported, none has the versatility or utility to replace IS6110-based fingerprinting nor have they been widely used in any area of endemicity. The second point is that most molecular epidemiology studies of M. tuberculosis have been done mainly in the developed world, where the extent and type of variants are likely to be limited, unlike in an area where the disease is endemic. This also raises the fundamental question of whether the strain variations are restricted to IS6110 distribution or the genetic differences are deeper. Thus, this modest study indicates a need to carry out systematic molecular epidemiological screenings in areas of endemicity to learn more about this pathogen.


    ACKNOWLEDGMENTS

Rajiv Gandhi Centre for Biotechnology is under the Science, Technology and Environment Department of the Government of Kerala and is a recipient of program mode support from the Department of Biotechnology, Government of India.

The technical assistance of Laiza K. Paul is gratefully acknowledged.


    FOOTNOTES

* Phone: 91-471-345 899 Fax: 91-471-329 472 E-mail: mundayoor_s{at}yahoo.com


    REFERENCES

1. Agasino, C. B., A. Ponce de Leon, R. M. Jasmer, and P. M. Small. 1998. Epidemiology of Mycobacterium tuberculosis that do not contain IS6110. Int. J. Tuberc. Lung Dis. 2:518-520[Medline].
2. Bifani, P. J., B. B. Plikaytis, V. Kapur, K. Stockbauer, X. Pan, M. L. Lutley, S. L. Moghazeh, W. Eisner, T. M. Daniel, M. H. Kaplan, J. T. Crawford, J. M. Musser, and B. N. Kreisworth. 1996. Origin and interstate spread of a New York City multidrug resistant Mycobacterium tuberculosis clone family. JAMA 275:452-457[Abstract/Free Full Text].
3. Das, S., C. N. Paramasivan, D. B. Lowrie, R. Prabhakar, and P. R. Narayanan. 1995. IS6110 restriction fragment length polymorphism typing of clinical isolates of Mycobacterium tuberculosis in Madras, South India. Tuber. Lung Dis. 76:550-554[CrossRef][Medline].
4. Fang, Z., and K. J. Forbes. 1997. A Mycobacterium tuberculosis IS6110 preferential locus for insertion into the genome. J. Clin. Microbiol. 35:479-481[Abstract].
5. Kumar, R. A., I. Radhakrishnan, and S. Mundayoor. 2000. A single-step, ecofriendly method to extract DNA from Mycobacterium tuberculosis for polymerase chain reaction. Anal. Biochem. 286:310-312[CrossRef][Medline].
6. Park, Y. K., G. H. Bai, and S. J. Kim. 2000. Restriction fragment length polymorphism analysis of Mycobacterium tuberculosis isolated from countries in the Western Pacific Region. J. Clin. Microbiol. 38:191-197[Abstract/Free Full Text].
7. Saunders, N. A. 1999. Review article: strain typing of Mycobacterium tuberculosis. J. Infect. 18:80-86.
8. van Embden, J. D. A., M. D. Cave, J. T. Crawford, J. W. Dale, K. D. Eisenach, B. Giquel, P. W. M. Hermans, C. Martin, R. McAdam, T. Shinnick, and P. M. Small. 1993. Strain identification of Mycobacterium tuberculosis by DNA fingerprinting: recommendations for a standard methodology. J. Clin. Microbiol. 31:406-409[Abstract/Free Full Text].
Indulakshmi Radhakrishnan
Manju Y. K.
R. Ajay Kumar
Sathish Mundayoor*
Mycobacterial Research Group
Rajiv Gandhi Centre for Biotechnology
Jagathy
Trivandrum 695 014
Kerala, India

Journal of Clinical Microbiology, April 2001, p. 1683-1683, Vol. 39, No. 4
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.4.1683.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Richardson, E. T., Samson, D., Banaei, N. (2009). Rapid Identification of Mycobacterium tuberculosis and Nontuberculous Mycobacteria by Multiplex, Real-Time PCR. J. Clin. Microbiol. 47: 1497-1502 [Abstract] [Full Text]  
  • Haldar, S., Sharma, N., Gupta, V. K., Tyagi, J. S. (2009). Efficient diagnosis of tuberculous meningitis by detection of Mycobacterium tuberculosis DNA in cerebrospinal fluid filtrates using PCR. J Med Microbiol 58: 616-624 [Abstract] [Full Text]  
  • Phyu, S., Stavrum, R., Lwin, T., Svendsen, O. S., Ti, T., Grewal, H. M. S. (2009). Predominance of Mycobacterium tuberculosis EAI and Beijing Lineages in Yangon, Myanmar. J. Clin. Microbiol. 47: 335-344 [Abstract] [Full Text]  
  • Haldar, S., Chakravorty, S., Bhalla, M., De Majumdar, S., Tyagi, J. S. (2007). Simplified detection of Mycobacterium tuberculosis in sputum using smear microscopy and PCR with molecular beacons. J Med Microbiol 56: 1356-1362 [Abstract] [Full Text]  
  • Quan, C., Lu, C.-Z., Qiao, J., Xiao, B.-G., Li, X. (2006). Comparative evaluation of early diagnosis of tuberculous meningitis by different assays.. J. Clin. Microbiol. 44: 3160-3166 [Abstract] [Full Text]  
  • Balamurugan, R., Venkataraman, S., John, K. R., Ramakrishna, B. S. (2006). PCR Amplification of the IS6110 Insertion Element of Mycobacterium tuberculosis in Fecal Samples from Patients with Intestinal Tuberculosis.. J. Clin. Microbiol. 44: 1884-1886 [Abstract] [Full Text]  
  • Banu, S., Gordon, S. V., Palmer, S., Islam, R., Ahmed, S., Alam, K. M., Cole, S. T., Brosch, R. (2004). Genotypic Analysis of Mycobacterium tuberculosis in Bangladesh and Prevalence of the Beijing Strain. J. Clin. Microbiol. 42: 674-682 [Abstract] [Full Text]  
  • Mistry, N. F., Iyer, A. M., D'souza, D. T. B., Taylor, G. M., Young, D. B., Antia, N. H. (2002). Spoligotyping of Mycobacterium tuberculosis Isolates from Multiple-Drug-Resistant Tuberculosis Patients from Bombay, India. J. Clin. Microbiol. 40: 2677-2680 [Abstract] [Full Text]  
  • Brosch, R., Gordon, S. V., Marmiesse, M., Brodin, P., Buchrieser, C., Eiglmeier, K., Garnier, T., Gutierrez, C., Hewinson, G., Kremer, K., Parsons, L. M., Pym, A. S., Samper, S., van Soolingen, D., Cole, S. T. (2002). A new evolutionary scenario for the Mycobacterium tuberculosis complex. Proc. Natl. Acad. Sci. USA 99: 3684-3689 [Abstract] [Full Text]  

This Article
Right arrow Full Text (PDF)
Right arrow A correction has been published
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Radhakrishnan, I.
Right arrow Articles by Mundayoor, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Radhakrishnan, I.
Right arrow Articles by Mundayoor, S.

Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»