ALLELE-SPECIFIC PCR METHOD BASED ON PNCA AND OXYR SEQUENCES FOR DISTINGUISHING MYCOBACTERIUM-BOVIS FROM MYCOBACTERIUM-TUBERCULOSIS - INTRASPECIFIC M-BOVIS PNCA SEQUENCE POLYMORPHISM

An allele-specific amplification method based on two genetic polymorph isms to differentiate Mycobacterium tuberculosis from Mycobacterium bo vis was tested. Based on the differences found at position 169 in the pncA genes from M. tuberculosis and M. bovis, a PCR system which was a ble to differentiate most of the 237 M. tuberculosis complex isolates tested in one of the two species was developed. All 121 M. tuberculosi s strains showed the expected base (cytosine) at position 169. Most of the M. bovis isolates had a guanine at the cited position. Neverthele ss, 18 of the 116 M. bovis isolates, all of them goat isolates, showed the pncA polymorphism specific to M tuberculosis. These results sugge st that goat M bovis may be the nicotinamidase-missing link at the ori gin of the M. tuberculosis species. Based on the polymorphism found at position 285 in the oxyR gene, the same system was used to differenti ate M. tuberculosis from M. bovis. In this case, DNAs from all 121 M. tuberculosis isolates had the expected base (guanine) at this position . In addition, all 116 M. bovis isolates, including those hom goats, s howed the identical polymorphism (adenine). The oxyR allele-specific a mplification method can differentiate M. bovis from M. tuberculosis, i s rapid (results can be obtained in less than 3 h), and is easy to per form.