ABSTRACT The human polyomavirus BK virus (BKV) establishes a latent and asymptomatic infection in the majority of the population. In immunocompromised individuals, the virus frequently (re)activates and may cause severe disease such as interstitial nephritis and hemorrhagic cystitis. Currently, the therapeutic options are limited to reconstitution of the antiviral immune response. T cells are particularly important for controlling this virus, and T cell therapies may provide a highly specific and effective mode of treatment. However, little is known about the phenotype and function of BKV-specific T cells in healthy individuals. Using tetrameric BKV peptide-HLA-A02 complexes, we determined the presence, phenotype, and functional characteristics of circulating BKV VP1-specific CD8 ⁺ T cells in 5 healthy individuals. We show that these cells are present in low frequencies in the circulation and that they have a resting CD45RA ⁻ CD27 ⁺ memory and predominantly CCR7 ⁻ CD127 ⁺ KLRG1 ⁺ CD49d ^hi CXCR3 ^hi T-bet ^int Eomesodermin ^lo phenotype. Furthermore, their direct cytotoxic capacity seems to be limited, since they do not readily express granzyme B and express only little granzyme K. We compared these cells to circulating CD8 ⁺ T cells specific for cytomegalovirus (CMV), Epstein-Barr virus (EBV), and influenza virus (Flu) in the same donors and show that BKV-specific T cells have a phenotype that is distinct from that of CMV- and EBV-specific T cells. Lastly, we show that BKV-specific T cells are polyfunctional since they are able to rapidly express interleukin-2 (IL-2), gamma interferon (IFN-γ), tumor necrosis factor α, and also, to a much lower extent, MIP-1β and CD107a.