Although Mycobacterium marinum is closely related to Mycobacterium tuberculosis H37Rv genomically, the clinical outcome in humans is quite different for M. marinum and M. tuberculosis infections. We investigated possible factors in the host macrophages for determining differential pathological responses to M. tuberculosis and M. marinum using an in vitro model of mycobacterial infection. Using suppression-subtractive hybridization, we identified 12 differentially expressed genes in the human monocytic cell line U937 infected with M. tuberculosis and M. marinum. Of those genes, the most frequently recovered transcript encoded interleukin-8 (IL-8). Northern hybridization revealed that IL-8 mRNA was highly upregulated in M. tuberculosis-infected U937 cells compared with M. marinum-infected cells. In addition, enzyme-linked immunosorbent assay showed that IL-8 protein secretion was significantly elevated in M. tuberculosis-infected U937 cells, human primary monocytes, and monocyte-derived macrophages compared with that in M. marinum-infected cells. The depressed IL-8 expression was unique in M. marinum-infected cells compared with cells infected with other strains of mycobacteria, including M. tuberculosis H37Ra, Mycobacterium bovis BCG, or Mycobacterium smegmatis. When the expression of NF-kappaB was assessed in mycobacterium-infected U937 cells, IkappaBalpha proteins were significantly degraded in M. tuberculosis-infected cells compared with M. marinum-infected cells. Collectively, these results suggest that differential IL-8 expression in human macrophages infected with M. tuberculosis and M. marinum may be critically associated with distinct host responses in tuberculosis. Additionally, our data indicate that differential signal transduction pathways may underlie the distinct patterns of IL-8 secretion in cells infected by the two mycobacteria.