Molecular characterization of mycobacteria isolated from seals

Tuberculosis (TB) was diagnosed in 10 seals from three species (Arctocephal us australis, Arctocephalus tropicalis and Otaria flavescens) found in Sout h America. The mycobacteria isolated from these cases belonged to the Mycob acterium tuberculosis complex, as determined by RFLP using an IS6110 probe, spoligotyping, analysis of the 16S rRNA gene sequence and by PCR-restricti on analysis of hsp65. Polymorphisms in gyrA, katG, oxyR and pncA were inves tigated in some of the isolates, as well as the presence of the MPB70 antig en. The insertion sequence IS6110 was present in three to seven copies in t he genome of the mycobacteria isolated from seals. Using the IS6110 probe, six patterns (designated A, B, C, D, E and F) were identified from 10 diffe rent isolates. Patterns A and B were found for the mycobacteria isolated fr om two and four seals, respectively, indicating an epidemiological relation ship between isolates grouped according to their IS6110 RFLP. The mycobacte ria isolated from seals shared the majority of their IS6110 DNA-containing restriction fragments, and nine isolates had an identical spoligotype; only one isolate showed a minor difference in its spoligotype. In addition, non e of these spoligotypes were found in other M. tuberculosis complex strains . These results suggest that the isolates from seals constitute a unique gr oup of closely related strains. The mycobacteria isolated from seals showed polymorphisms at gyrA codon 95 and katG codon 463, as do group 1 M. tuberc ulosis, and M. bovis. Group 1 mycobacteria are associated with cluster case s. The spoligotypes found in the mycobacteria isolated from seals lack spac ers 39-43, as does M. bovis, but the MPB70 antigen, which is highly express ed in M, bovis and minimally expressed in M. tuberculosis, was not detected in these mycobacteria. The mycobacteria isolated from seals also showed ox yR and pncA polymorphisms specific to M. tuberculosis. In conclusion, the m ycobacteria that cause TB in seals in the South-Western Atlantic are a rela ted group, and based on the combination of genetic characteristics, belong to a unique genotypic group within the M. tuberculosis complex.