Like αβ T cells, human γδ T cells also have different subsets with distinct characteristics. Whether human Vγ9Vδ2 T cells have functionally different subsets in response to influenza A (fluA) viruses remains unknown. In this study, we show for the first time that both central (CD45RA(-)CD27(+)) and effector (CD45RA(-)CD27(-)) memory Vγ9Vδ2 T cells have similar levels of immediate interferon (IFN) γ and cytotoxic responses to human and avian fluA virus-infected cells. In contrast, CD56(+) Vγ9Vδ2 T cells have significantly higher cytotoxicity against fluA virus-infected cells compared with their CD56(-) counterparts, whereas both subsets have similar IFN-γ responses. We further demonstrate that the CD16-dependent degranulation pathway, but not antibody-dependent cell-mediated cytotoxicity, contribute to the superior cytotoxicity of CD56(+) Vγ9Vδ2 T cells. Our study provides further evidence for the phenotypic and functional characterization of human Vγ9Vδ2 T-cell subsets during fluA virus infection and may help improve the γδ T-cell-based immunotherapy for viral infection.