Several molecular systems for direct detection of Mycobacterium tuberculosis complex (MTBC) have recently been developed. The GenoQuick MTB assay (GQ-MTB) used in this study detected 82 of the 96 (85.4%) samples with MTBC, including 50 of 64 (78.1%) samples with negative acid-fast bacillus smears. Fifteen samples containing nontuberculous mycobacteria were also studied: 13 were GQ-MTB negative, one was positive, and one was indeterminate. GQ-MTB showed good effectiveness for the direct detection of MTBC from clinical samples.