Invariant Valpha14(+) NKT cells are a specialized CD1-reactive T cell subset implicated in innate and adaptive immunity. We assessed whether Valpha14(+) NKT cells participated in the immune response against enteric Listeria monocytogenes infection in vivo. Using CD1d tetramers loaded with the synthetic lipid alpha-galactosylceramide (CD1d/alphaGC), we found that splenic and hepatic Valpha14(+) NKT cells in C57BL/6 mice were early producers of IFN-gamma (but not IL-4) after L. monocytogenes infection. Adoptive transfer of Valpha14(+) NKT cells derived from TCRalpha degrees Valpha14-Jalpha18 transgenic (TCRalpha degrees Valpha14Tg) mice into alymphoid Rag(null) gamma(c)(null) mice demonstrated that Valpha14(+) NKT cells were capable of providing early protection against enteric L. monocytogenes infection with systemic production of IFN-gamma and reduction of the bacterial burden in the liver and spleen. Rechallenge experiments demonstrated that previously immunized wild-type and Jalpha18null mice, but not TCRalpha(null) or TCRalpha(null) Valpha14Tg mice, were able to mount adaptive responses to L. monocytogenes. These data demonstrate that Valpha14(+) NKT cells are able to participate in the early response against enteric L. monocytogenes through amplification of IFN-gamma production, but are not essential for, nor capable of, mediating memory responses required to sterilize the host.