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Journal of Clinical Microbiology, September 2008, p. 3005-3011, Vol. 46, No. 9
0095-1137/08/$08.00+0     doi:10.1128/JCM.00437-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Utility of New 24-Locus Variable-Number Tandem-Repeat Typing for Discriminating Mycobacterium tuberculosis Clinical Isolates Collected in Bulgaria{triangledown} ,{dagger}

Violeta Valcheva,1 Igor Mokrousov,2 Olga Narvskaya,2 Nalin Rastogi,3* and Nadya Markova1*

Department of Pathogenic Bacteria, The Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Sciences, Sofia 1113, Bulgaria,1 Laboratory of Molecular Microbiology, St. Petersburg Pasteur Institute, 197101 St. Petersburg, Russia,2 Unité de la Tuberculose et des Mycobactéries, Institut Pasteur de Guadeloupe, Abymes 97183, Guadeloupe, France3

Received 5 March 2008/ Returned for modification 8 April 2008/ Accepted 2 July 2008

The present study evaluated new markers for molecular typing of Mycobacterium tuberculosis with a collection of strains circulating in Bulgaria. A study sample included 133 strains from epidemiologically unlinked patients from different regions of the country. Spoligotyping was used as a primary typing tool; it subdivided these strains into 37 types, including 15 clusters and 22 singletons. Traditional IS6110-restriction fragment length polymorphism (RFLP) typing and novel 24-locus variable number tandem-repeat (VNTR) typing methods were applied to the selection of 73 strains. Discriminatory power (Hunter-Gaston index [HGI]) of these methods was found to be 0.983 and 0.997, respectively. The 73 strains were subdivided into 66 types by a 24-locus mycobacterial interspersed repetitive unit (MIRU)-VNTR scheme, 62 types by a classical 12-locus MIRU-VNTR scheme, 51 types by IS6110-RFLP typing, and 31 types by spoligotyping. A combination of the five most polymorphic loci (MIRU40, Mtub04, Mtub21, QUB-11b, and QUB-26) was shown to achieve a high discrimination (HGI = 0.984). To conclude, a complete 24-locus scheme excellently differentiated strains in our study, whereas a reduced 5-locus set provided a sufficiently high differentiation and may be preliminarily suggested for the first-line typing of M. tuberculosis isolates in Bulgaria.

* Corresponding authors. Mailing address for N. Markova: Department of Pathogenic Bacteria, The Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Sciences, Sofia 1113, Bulgaria. Phone: 3592 9793168. Fax: 3592 8700109. E-mail: nadya.markova{at}gmail.com. Mailing address for N. Rastogi: Unité de la Tuberculose et des Mycobactéries, Institut Pasteur de Guadeloupe, Abymes 97183, Guadeloupe, France. Phone: (590) 590-893881. Fax: (590) 590-893880. E-mail: nrastogi{at}pasteur-guadeloupe.fr

{triangledown} Published ahead of print on 9 July 2008.

{dagger} Supplemental material for this article may be found at http://jcm.asm.org/.

Journal of Clinical Microbiology, September 2008, p. 3005-3011, Vol. 46, No. 9
0095-1137/08/$08.00+0     doi:10.1128/JCM.00437-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



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