We previously found that centaurein enhanced IFN-γ transcription in T cells. Here, we demonstrate that centaurein increased the IFN-γ expression in T and NK cells and the serum IFN-γ level in mice. Centaurein elevated the transcription of T-bet but not GATA-3, which is consistent with its effect on that of IFN-γ but not IL-4. Additionally, centaurein effectively protected mice against Listeria infection. Moreover, centaurein per se or in combination with antibiotics could treat Listeria infection. Our mechanistic studies suggest that centaurein augments IFN-γ expression via a transcriptional up-regulation of T-bet and that centaurein protects against or treats Listeria infection via a modulation of IFN-γ expression.