Previous studies have suggested that yS T cells can be increased in HIV-1 -seropositive individuals, although characterization of y6 T cell subtypes and correlation with clinical status of these patients have not been performed. We investigated groups of adult HIV-seropositive persons to determine the prevalence of elevated levels of y6 T cells and whether any y6 T cell subtypes were preferentially expressed. Since a large proportion of human yS T cells appear to be reactive to proteins encoded by mycobacteria, we also examined our patients for the incidence of mycobacterial infection. Our results show that a significant number of HIV-positive patients have an elevated number of yS T cells in their peripheral blood as compared to normal controls. HIV-seropositive patients with clinical or laboratory evidence of mycobacterial infection had statistically significant increases in the percentage and total numbers of y6 T cells over the HIV-positive persons without mycobacterial infection. An examination of the subtypes of yS T cells revealed that certain subtypes such as Vy9+ and VS2+ T cells were preferentially elevated in the mycobacteria-positive patients. These results suggest that an increased number of y6 T cells in HIV-positive patients is most often seen in the setting of an opportunistic mycobacterial infection and that specific yS T cell subtypes are stimulated under these conditions. The role of these increased numbers of yS T cells in HIV-associated disease is unclear but is likely a component of the response and degree of host resistance to this organism.