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Journal of Clinical Microbiology, January 2001, p. 57-65, Vol. 39, No. 1
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.1.57-65.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Genetic Diversity of Mycobacterium africanum Clinical Isolates Based on IS6110-Restriction Fragment Length Polymorphism Analysis, Spoligotyping, and Variable Number of Tandem DNA Repeats

Cristina Viana-Niero,1 Cristina Gutierrez,1 Christophe Sola,2 Ingrid Filliol,2 Fadila Boulahbal,1 Véronique Vincent,1 and Nalin Rastogi2,*

Centre National de Référence des Mycobactéries, Institut Pasteur, 75724-Paris Cedex 15, France,1 and Unité de la Tuberculose et des Mycobactéries, Institut Pasteur de Guadeloupe, F-97165 Pointe-à-Pitre Cedex, Guadeloupe2

Received 23 June 2000/Returned for modification 23 August 2000/Accepted 7 October 2000

A collection of 105 clinical isolates originally identified as Mycobacterium africanum were characterized using both phenotypic and genotyping methods. The phenotypic methods included routine determination of cultural properties and biochemical tests used to discriminate among the members of the M. tuberculosis complex, whereas genotypic characterization was based on IS6110-restriction fragment length polymorphism (IS6110-RFLP) analysis, IS1081-RFLP analysis, direct repeat-based spacer oligonucleotide typing (spoligotyping), variable number of tandem DNA repeats (VNTR), and the polymorphism of the oxyR, pncA, and mtp40 loci. The results obtained showed that a majority of M. africanum isolates were characterized by a specific spoligotyping pattern that was intermediate between those of M. tuberculosis and M. bovis, which do not hybridize with spacers 33 to 36 and spacers 39 to 43, respectively. A tentative M. africanum-specific spoligotyping signature appeared to be absence of spacers 8, 9, and 39. Based on spoligotyping, as well as the polymorphism of oxyR and pncA, a total of 24 isolates were excluded from the final study (19 were identified as M. tuberculosis, 2 were identified as M. canetti, and 3 were identified as M. bovis). The remaining 81 M. africanum isolates were efficiently subtyped in three distinct subtypes (A1 to A3) by IS6110-RFLP analysis and spoligotyping. The A1 and A2 subgroups were relatively more homogeneous upon spoligotyping than A3. Further analysis of the three subtypes by VNTR corroborated the highly homogeneous nature of the A2 subtype but showed significant variations for subtypes A1 and A3. A phylogenetic tree based on a selection of isolates representing the three subtypes using VNTR and spoligotyping alone or in combination confirmed the subtypes described as well as the heterogeneity of subtype A3.

* Corresponding author. Mailing address: Unité Tuberculose et Mycobactéries, Institut Pasteur de Guadeloupe, Morne Jolivière, BP 484, F-97165 Pointe-à-Pitre Cedex, Guadeloupe. Phone: 590-893-881. Fax: 590-893-880. E-mail: rastogi{at}pasteur.gp.

Journal of Clinical Microbiology, January 2001, p. 57-65, Vol. 39, No. 1
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.1.57-65.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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