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Journal of Clinical Microbiology, September 2000, p. 3231-3234, Vol. 38, No. 9
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Differentiation of Clinical Mycobacterium tuberculosis Complex Isolates by gyrB DNA Sequence Polymorphism Analysis

Stefan Niemann,1,* Dag Harmsen,2 Sabine Rüsch-Gerdes,1 and Elvira Richter1

Forschungszentrum Borstel, National Reference Center for Mycobacteria, D-23845 Borstel,1 and Institute for Hygiene & Microbiology, University of Würzburg, D-97080 Würzburg,2 Germany

Received 20 April 2000/Returned for modification 7 June 2000/Accepted 30 June 2000

The discriminatory power of gyrB DNA sequence polymorphisms for differentiation of the species of the Mycobacterium tuberculosis complex (MTBC) was evaluated by sequencing and restriction fragment length polymorphism (RFLP) analysis of a 1,020-bp fragment amplified from clinical isolates of M. tuberculosis, Mycobacterium bovis (pyrazinamide [PZA] resistant as well as PZA susceptible), Mycobacterium africanum subtypes I and II, and Mycobacterium microti types vole and llama. We found sequence polymorphisms in four regions described previously and at one additional position. These differences in the gyrB sequences allow an accurate discrimination of M. bovis, M. microti, and M. africanum subtype I. The PZA-susceptible subtypes of M. bovis shared the M. bovis-specific substitution at position 756 with the PZA-resistant strains, but can be unambiguously differentiated by a characteristic substitution at position 1311. As a drawback, M. tuberculosis and M. africanum subtype II showed an identical gyrB sequence that facilitates discrimination from the other species, but not from each other. A PCR-RFLP technique applying three restriction enzymes could be shown to be a rapid and easy-to-perform tool for the differentiation of the members of the MTBC. Based on these results, we present a clear diagnostic algorithm for the differentiation of species of the MTBC.

* Corresponding author. Mailing address: Forschungszentrum Borstel, National Reference Center for Mycobacteria, Parkallee 18, D-23845 Borstel, Germany. Phone: (49)-4537-188658. Fax: (49)-4537-188311. E-mail: sniemann{at}fz-borstel.de.

Journal of Clinical Microbiology, September 2000, p. 3231-3234, Vol. 38, No. 9
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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