Genetic biodiversity of Mycobacterium tuberculosis complex strains from patients with pulmonary tuberculosis in Cameroon

Abstract

We analyzed DNA polymorphisms in 455 Mycobacterium tuberculosis complex isolates from 455 patients to evaluate the biodiversity of tubercle bacilli in Ouest province, Cameroon. The phenotypic and genotypic identification methods gave concordant results for 99.5% of M. tuberculosis isolates (413 strains) and for 90% of Mycobacterium africanum isolates (41 strains). Mycobacterium bovis was isolated from only one patient. Analysis of regions of difference (RD4, RD9, and RD10) proved to be an accurate and rapid method of distinguishing between unusual members of the M. tuberculosis complex. Whereas M. africanum strains were the etiologic agent of tuberculosis in 56% of cases 3 decades ago, our results showed that these strains now account for just 9% of cases of tuberculosis. We identified a group of closely genetically related M. tuberculosis strains that are currently responsible for >40% of smear-positive pulmonary tuberculosis cases in this region of Cameroon. These strains shared a spoligotype lacking spacers 23, 24, and 25 and had highly related IS6110 ligation-mediated (LM) PCR patterns. They were designated the "Cameroon family." We did not find any significant association between tuberculosis-causing species or strain families and patient characteristics (sex, age, and human immunodeficiency virus status). A comparison of the spoligotypes of the Cameroon strains with an international spoligotype database (SpolDB3) containing 11,708 patterns from >90 countries, showed that the predominant spoligotype in Cameroon was limited to West African countries (Benin, Senegal, and Ivory Coast) and to the Caribbean area.

FIG.1.

Dendrogram and schematic representation of the 95 spoligotype patterns obtained from the 455 M. tuberculosis complex strains. The degree of similarity of the spoligotypes was calculated by the 1-Jaccard index. The relationships between patterns were assessed by the unweighted pair group method of averages. The spoligotypes listed are the designations in the SpolDB3 database. Spoligotypes 836 to 862 were first described in this study. Orphan patterns did not have matches in the SpolDB3 database.

FIG. 2.

The 11 different banding patterns (A to K) obtained by IS6110 LM-PCR for the 193 strains included in the Cameroon family. Lane 1, 100-bp ladder (Gibco). Strains with large DR deletions, i.e., spoligotypes 847 and 852, showed LM-PCR pattern C, G, or J.

FIG. 3.

Spoligotype patterns of the 193 Cameroon family strains.