Molecular characterization of Mycobacterium tuberculosis isolates presenting various drug susceptibility from Greece using three DNA typing methods.
OBJECTIVE
The purpose of the present study was the molecular characterization of Mycobacterium tuberculosis clinical isolates using three DNA typing methods.


METHODS
One hundred nineteen independent (77 susceptible to all antituberculous drugs, 17 rifampin-resistant and 25 isoniazid-resistant), and nine related Mycobacterium tuberculosis isolates obtained over a 3-years period (1997-1999) from Greece were typed with restriction fragment length polymorphism (RFLP), using the non-radioactive IS6110 probe (IS6110-RFLP), and two PCR-based molecular methods: random amplification of polymorphic domains (RAPD) using four different primers and double repetitive element-PCR (DRE-PCR).


RESULTS
IS6110-RFLP and RAPD-PCR using IRIS primer were proved to be the most discriminatory methods, while DRE-PCR gave satisfactory results and RAPD-PCR methods using the other three primers (A1245, B1245 and Leg2) were not so effective. The related strains, isolated from affected members of four families, gave similar PCR and RFLP patterns, while the independent strains presented a high degree of polymorphism. In terms of cost effectiveness and technical simplicity, the PCR-based methods were found to be superior.


CONCLUSIONS
So, they may serve as screening methods to classify a large number of isolates into clusters for further subtyping and recognition of well-defined genotype families.
Department of Microbiology, Medical School, University of Ioannina, 45 110 Ioannina, Greece. gvrioni@cc.uoi.gr
Georgia
G
Vrioni
Stamatina
S
Levidiotou
Peggy
P
Matsiota-Bernard
Evangelos
E
Marinis
D000328:Adult; D000368:Aged; D015373:Bacterial Typing Techniques; D002648:Child; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D005260:Female; D006115:Greece; D006801:Humans; D008297:Male; D008875:Middle Aged; D009169:Mycobacterium tuberculosis / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length
D004269:DNA, Bacterial