Forms of mutation never before described in the rpoB gene are reported for a sample of 20 rifampicin-resistant Mycobacterium avium Complex (MAC) strains isolated from AIDS patients in Thailand. All strains were analyzed by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and polymerase chain reaction-DNA sequencing (PCR-DNA sequencing). Sequence analysis of these strains revealed that only one strain (5%) has missense mutation at Lys-626 (Thr) and the rest of the strains had 15 different silent mutations within a 542 bp region of the rpoB gene. Five strains (25%) had a silent mutation at only one position, 7 (35%) at 2 positions, 7 (35%) at 3 positions, and 1 (5%) at 7 positions. The silent mutation at the Ala-630 codon occurred in the largest proportion of the strains (15 strains, 75%), followed by the Val-581 in 8 strains (40%), Tyr-578 and Thr-600 in 4 strains (20%), and Gly-597 in 3 strains (15%). This investigation demonstrates that mutation in the rpoB gene of MAC strains from Thailand are more varied than previously reported for RIF MAC strains. PCR-SSCP screening clearly separated RIFr strains from rifampicin-susceptible (RIFs) strains.