CD161⁺⁺/MAIT cells are not preferentially infected in vitro. PBMCs from healthy subjects (n = 9) were activated with PHA, cultured in rhIL2 and rhIL7, and infected with a CCR5-tropic virus (JR-CSF) or CXCR4 tropic virus (MN) at an MOI of 10. FACS analysis was performed on days 6 and 9 after infection as described in “Methods.” The gating strategy is shown in supplemental Figure 9B through D. (A) CD8⁺ T cells were infected at a low frequency by both viruses, with the greatest frequency of p24⁺ cells observed on day 6 after infection with JR-CSF. (B) Representative FACS plots of CD3⁺ lymphocytes from day 6 after infection with JR-CSF. (C) When gating on the total p24⁺CD8⁺ T cells, the majority of the infected CD8⁺ T cells were found in the non-MAIT CD161⁻CD8⁺ T-cell population on both day 6 and day 9 after infection with JR-CSF virus. (D) Representative FACS plots of p24 staining of CD8⁺ T cells mock infected or infected with JR-CSF. (E) Comparison of the frequency of infection within the CD161⁺⁺/MAIT, and non-MAIT CD161⁺ and CD161⁻CD8⁺ T-cell populations. CD161⁺⁺/MAIT cells were infected, but not more frequently than either the non-MAIT CD161⁻ or CD161⁺CD8⁺ T-cell populations on either day 6 or day 9 after infection with JR-CSF virus. (F) There was no significant difference in the survival of the cells in the HIV infected cultures compared with the uninfected cultures. Cell survival ratio was calculated by normalizing the frequency of CD161⁺⁺/MAIT cells in the infected culture to that of the uninfected culture.