Rapid clonal analysis of recurrent tuberculosis by direct MIRU-VNTR typing on stored isolates.
BACKGROUND
The application of molecular tools to the analysis of tuberculosis has revealed examples of clonal complexity, such as exogenous reinfection, coinfection, microevolution or compartmentalization. The detection of clonal heterogeneity by standard genotyping approaches is laborious and often requires expertise. This restricts the rapid availability of Mycobacterium tuberculosis (MTB) genotypes for clinical or therapeutic decision-making. A new PCR-based technique, MIRU-VNTR, has made it possible to genotype MTB in a time frame close to real-time fingerprinting. Our purpose was to evaluate the capacity of this technique to provide clinicians with a rapid discrimination between reactivation and exogenous reinfection and whether MIRU-VNTR makes it possible to obtain data directly from stored MTB isolates from recurrent episodes.


RESULTS
We detected differences, between the MIRUtypes of recurrent isolates in 38.5% (5/13) of the cases studied. These included cases of i) exogenous reinfection, often with more resistant strains, ii) likely examples of microevolution, leading to the appearance of new clonal variants and iii) a combination of microevolution, coinfection and competition.


CONCLUSIONS
MIRU-VNTR rapidly obtained clinically useful genotyping data in a challenging situation, directly from stored MTB isolates without subculturing them or purifying their DNA. Our results also mean that MIRU-VNTR could be applied for easy, rapid and affordable massive screening of collections of stored MTB isolates, which could establish the real dimension of clonal heterogeneity in MTB infection.
Servicio de Microbiología y Enfermedades Infecciosas, Hospital General Universitario Gregorio Marañón, CIBER Enfermedades Respiratorias (CIBERES), Universidad Complutense, Madrid, Spain. anamartsan@yahoo.es
Ana
A
Martín
Marta
M
Herránz
María Jesús Ruiz
MJ
Serrano
Emilio
E
Bouza
Darío
D
García de Viedma
D000328:Adult; D015373:Bacterial Typing Techniques; D005260:Female; D014644:Genetic Variation; D006801:Humans; D008297:Male; D018895:Microsatellite Repeats / Q000235:genetics; D008875:Middle Aged; D009169:Mycobacterium tuberculosis / Q000145:classification / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012008:Recurrence; D014376:Tuberculosis / Q000382:microbiology* / Q000473:pathology