The ability of two different Lactobacillus strains (Lactobacillus salivarius CECT5713 and Lactobacillus fermentum CECT5716), isolated from human breast milk, to modulate the immune response was examined. In rodent bone-marrow-derived macrophages (BMDM), the presence of Lact. fermentum CECT5716 induced pro-inflammatory cytokines, in contrast to the activation of IL-10 induced by Lact. salivarius CECT5713. Although both strains reduced the lipopolysaccharide (LPS)-induced inflammatory response in BMDM, the effect of Lact. salivarius CECT5713 was more efficient, probably because of the production of higher amounts of IL-10 cytokine. In vivo assays in mice showed similar results; the consumption of Lact. fermentum CECT5716 enhanced the production of Th1 cytokines by spleen cells and increased the IgA concentration in faeces. However, the consumption of Lact. salivarius CECT5713 induced IL-10 production by spleen cells. Therefore, in general, the effect of Lact. fermentum CECT5716 is immunostimulatory in contrast to the anti-inflammatory effect of Lact. salivarius CECT5713. The results of this study show that two Lactobacillus strains isolated from breast milk can exert different and even opposing effects on immune response demonstrating the specificity of each strain.