The clonal composition of Mycobacterium tuberculosis in clinical specimens could be modified by culture.
BACKGROUND
The application of molecular tools has revealed that infection by Mycobacterium tuberculosis (MTB) is more complex than initially assumed. Genotyping is generally performed on cultures. However, there is no information about bacterial clonal complexity in clinical specimens or whether standard culture procedures can modify this complexity.


METHODS
An in vitro assay was performed to determine whether culture can modify the clonal complexity of the MTB population in clinical specimens. Pairs of MTB strains (10 pairs) or stain-positive sputa (4 pairs) were mixed in different volumetric proportions. The DNA extracted from the mixtures before and after culture was genotyped using mycobacterial interspersed repetitive unit-variable-number tandem repeat analysis to detect potential changes in the proportion of the mixed strains.


RESULTS
In 6/10 pairs of MTB strains and 2/4 pairs of sputa, marked changes were observed in clonal composition after culture, even in mixtures of strains differing in their drug-susceptibility patterns. In some cases, only one of the mixed strains was detected after culture.


CONCLUSIONS
The initial clonal composition in bacteriologically complex clinical specimens could be underestimated if genotyping analysis is performed after culture. Genotyping strategies aimed at analyzing clinical samples must be optimized to reveal the real dimension of clonal complexity in infection by MTB.
Martín
Ana
A
Servicio de Microbiología Clínica y Enfermedades Infecciosas, Hospital General Universitario Gregorio Marañón. Madrid, Spain.
Herranz
Marta
M
Ruiz Serrano
María Jesús
MJ
Bouza
Emilio
E
García de Viedma
Darío
D
D015373:Bacterial Typing Techniques; D002478:Cells, Cultured; D017635:Clonal Anergy / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics* / Q000378:metabolism; D014644:Genetic Variation; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D020080:Tandem Repeat Sequences / Q000235:genetics*; D014397:Tuberculosis, Pulmonary / Q000235:genetics* / Q000382:microbiology
D004269:DNA, Bacterial