Differentiation of clinical Mycobacterium tuberculosis complex isolates by their GyrB polymorphism.
OBJECTIVE
To evaluate the reliability of the gyrB PCR-RFLP technique in differentiating clinical Mycobacterium tuberculosis complex isolates.


METHODS
A primer pair MTUB-f and MTUB-r for M. tuberculosis complex (MTBC) was used to differentiate 79 mycobacterial isolates by specific amplification of the 1,020-bp fragment of the gyrB gene (gyrB-PCR1). The MTBC isolates were further differentiated using a set of specific primers MTUB-756-Gf and MTUB-1450-Cr that allowed selective amplification of the gyrB fragment specific for M. tuberculosis (gyrB-PCR2). The DNA polymorphisms in the 1,020-bp gyrB fragment for 7 M. tuberculosis strains confirmed by PCR as well as 2 reference strains; M. tuberculosis H37Rv and M. bovis BCG were analyzed with the restriction enzyme Rsa1.


RESULTS
Seventy-seven (97.5%) isolates were positive for gyrB-PCR1 and thus identified as members of M. tuberculosis complex (MTBC) and two (2.6%) isolates were negative and identified as Mycobacteria other than tuberculosis (MOTT). All the M. tuberculosis isolates showed the typical M. tuberculosis specific Rsa1 RFLP patterns (100, 360, 560-bp) while 360 and 480-bp fragments were generated from M. bovis BCG.


CONCLUSIONS
The gyrB PCR-RFLP using the endonuclease Rsa1 can be used to differentiate M. tuberculosis from M. bovis in clinical isolates.
Abass
N A
NA
Department of Preventive Medicine, College of Veterinary Medicine and Animal Production, Sudan University of Science and Technology, PO Box 204, Khartoum North, Sudan.
Suleiman
K M
KM
El Jalii
I M
IM
D001426:Bacterial Proteins / Q000235:genetics*; D015373:Bacterial Typing Techniques / Q000379:methods*; D016172:DNA Fingerprinting / Q000379:methods*; D027081:DNA Gyrase / Q000235:genetics*; D017931:DNA Primers / Q000235:genetics; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D006801:Humans; D009163:Mycobacterium bovis / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000145:classification* / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D012150:Polymorphism, Restriction Fragment Length; D012680:Sensitivity and Specificity; D014376:Tuberculosis / Q000382:microbiology
D001426:Bacterial Proteins; D017931:DNA Primers; D015252:Deoxyribonucleases, Type II Site-Specific; C040158:GTAC-specific type II deoxyribonucleases; D027081:DNA Gyrase