This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Rajakumar, K. Articles by Barer, M. R. Search for Related Content PubMed PubMed Citation Articles by Rajakumar, K. Articles by Barer, M. R.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, May 2004, p. 1890-1896, Vol. 42, No. 5
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.5.1890-1896.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Use of Genome Level-Informed PCR as a New Investigational Approach for Analysis of Outbreak-Associated Mycobacterium tuberculosis Isolates

Kumar Rajakumar,^1,2, Jamila Shafi,^1, Rebecca J. Smith,^1, Richard A. Stabler,^3, Peter W. Andrew,¹ Deborah Modha,³ Gerry Bryant,⁴ Philip Monk,⁴ Jason Hinds,³ Philip D. Butcher,³ and Michael R. Barer^1,2*

Department of Infection, Immunity and Inflammation, Leicester Medical School, University of Leicester, Leicester LE1 9HN,¹ Department of Clinical Microbiology, University Hospitals of Leicester NHS Trust, Leicester LE1 5WW,² Leicestershire Health Authority, Leicester,⁴ Department of Cellular and Molecular Medicine (Medical Microbiology), St. George's Hospital Medical School, London SW17 0RE, United Kingdom³

Received 15 October 2003/ Returned for modification 29 November 2003/ Accepted 7 January 2004

Mycobacterium tuberculosis strain CH, the index isolate linked to a major tuberculosis outbreak associated with high levels of transmissibility and virulence, was characterized by microarray analysis by use of a PCR product array representative of the genome of M. tuberculosis strain H37Rv. Seven potential genomic deletions were identified in CH, five of which were confirmed by PCR analysis across the predicted deletion points. The panel of five PCRs required to individually interrogate these loci was collectively referred to as the genome level-informed PCR (GLIP) assay. GLIP analysis was performed with CH, 12 other epidemiologically linked isolates, and 43 recent, non-outbreak-associated isolates derived from patients within the local area. All 13 outbreak-linked isolates showed a profile corresponding to the presence of all five deletions. These 13 isolates were also found to share common variable-number tandem repeat and mycobacterial interspersed repetitive unit profiles. None of the 43 non-outbreak-associated isolates exhibited the five-deletion profile. Although three individual deletions were present in upwards of 44% of the non-outbreak-associated isolates, no single-deletion isolates were detected. Interestingly, none of these deletions had been previously recognized, and sequence analysis of the immediate flanking regions in CH failed to identify a likely mechanism of deletion for four of the five loci. The GLIP assay also proved valuable in ongoing surveillance of the outbreak, rapidly identifying a further two outbreak-associated cases months after the initial cluster and, importantly, dismissing a further 12 epidemiologically suspect cases, which allowed the optimum deployment of public health resources.

---

* Corresponding author. Mailing address: Department of Infection, Immunity and Inflammation, Leicester Medical School, University of Leicester, Maurice Shock Building, University Rd., P.O. Box 138, Leicester LE1 9HN, United Kingdom. Phone: 44 (0) 116 2522933. Fax: 44 (0) 116 2525030. E-mail: mrb19{at}leicester.ac.uk.

K.R., J.S., R.J.S., and R.A.S. contributed equally to this work.

---

Journal of Clinical Microbiology, May 2004, p. 1890-1896, Vol. 42, No. 5
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.5.1890-1896.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Ashworth, M., Horan, K. L., Freeman, R., Oren, E., Narita, M., Cangelosi, G. A. (2008). Use of PCR-Based Mycobacterium tuberculosis Genotyping To Prioritize Tuberculosis Outbreak Control Activities. J. Clin. Microbiol. 46: 856-862 [Abstract] [Full Text]  
• Newton, S. M., Smith, R. J., Wilkinson, K. A., Nicol, M. P., Garton, N. J., Staples, K. J., Stewart, G. R., Wain, J. R., Martineau, A. R., Fandrich, S., Smallie, T., Foxwell, B., Al-Obaidi, A., Shafi, J., Rajakumar, K., Kampmann, B., Andrew, P. W., Ziegler-Heitbrock, L., Barer, M. R., Wilkinson, R. J. (2006). A deletion defining a common Asian lineage of Mycobacterium tuberculosis associates with immune subversion. Proc. Natl. Acad. Sci. USA 103: 15594-15598 [Abstract] [Full Text]  
• Huard, R. C., Fabre, M., de Haas, P., Claudio Oliveira Lazzarini, L., van Soolingen, D., Cousins, D., Ho, J. L. (2006). Novel Genetic Polymorphisms That Further Delineate the Phylogeny of the Mycobacterium tuberculosis Complex.. J. Bacteriol. 188: 4271-4287 [Abstract] [Full Text]  
• Freeman, R., Kato-Maeda, M., Hauge, K. A., Horan, K. L., Oren, E., Narita, M., Wallis, C. K., Cave, D., Nolan, C. M., Small, P. M., Cangelosi, G. A. (2005). Use of Rapid Genomic Deletion Typing To Monitor a Tuberculosis Outbreak within an Urban Homeless Population. J. Clin. Microbiol. 43: 5550-5554 [Abstract] [Full Text]  
• Stabler, R. A., Marsden, G. L., Witney, A. A., Li, Y., Bentley, S. D., Tang, C. M., Hinds, J. (2005). Identification of pathogen-specific genes through microarray analysis of pathogenic and commensal Neisseria species. Microbiology 151: 2907-2922 [Abstract] [Full Text]  
• Ou, H.-Y., Smith, R., Lucchini, S., Hinton, J., Chaudhuri, R. R., Pallen, M., Barer, M. R., Rajakumar, K. (2005). ArrayOme: a program for estimating the sizes of microarray-visualized bacterial genomes. Nucleic Acids Res 33: e3-e3 [Abstract] [Full Text]