Major racial and gender differences have been documented in the natural history and treatment responses of chronic hepatitis C virus (HCV) infection; however, distinct mechanisms have remained enigmatic. We hypothesized that racial‐ and gender‐related differences in natural killer (NK) cell populations may explain altered natural history and treatment responses. Our study cohort consisted of 29 African‐American (AA; 55% male) and 29 Caucasian‐American (CA; 48% male) healthy uninfected control subjects. Multiparameter flow cytometric analysis was used to characterize levels, phenotype with respect to 14 NK receptors, and lymphokine‐activated killing (LAK) function. Gene expression was assessed by real‐time reverse‐transcriptase polymerase chain reaction after 6‐hour in vitro stimulation with Toll‐like receptor (TLR) ligands. The ability to control HCV infection was assessed in the Huh‐7.5/JFH‐1 coculture system. NK expression of natural cytotoxicity receptor NKp46 was strongly associated with CA race and female gender and correlated positively with LAK activity (P = 0.0054). NKp46^high NKs were more efficient at controlling HCV than their NKp46^low counterparts (P < 0.001). Similarly, ligation of NKp46 on isolated NK cells resulted in a significant reduction in the HCV copy number detected in Huh‐7.5/JFH‐1 coculture (multiplicity of infection: 0.01) at an effector:target ratio of 5:1 (P < 0.005). After TLR stimulation, genes involved in cytotoxicity, but not cytokine genes, were significantly up‐regulated in NKp46^high NKs. Cytokine stimulation (interleukin [IL]‐12 and IL‐15) demonstrated that NKp46^high NK cells have significantly higher interferon‐gamma production than NKp46^low cells. TLR stimulation significantly induced degranulation as well as tumor necrosis factor alpha (TNF‐α)‐related apoptosis‐inducing ligand, Fas, and TNF‐α protein expression in NKp46^high NKs. NKp46 ligand was induced on HCV‐infected hepatocytes. Conclusions: NKp46 expression may contribute to differential HCV responses. NKp46 expression correlates with anti‐HCV activity in vitro and thus may prove to be a useful therapeutic target. (HEPATOLOGY 2012)