In this study, several cytokine responses were investigated during Mycoplasma hyopneumoniae (Mhp) infection using a gnotobiotic infection model. We found that several inflammatory cytokines (IL-1beta, IL-8, IL-18, and TNF-alpha) and an anti-inflammatory cytokine IL-10 were induced from peripheral blood mononuclear cells (PBMC) of germ-free (GF) piglets stimulated with heat killed Mhp whole antigens, but no IFN-gamma and IL-4 were induced by Mhp. After the intranasal infection of Mhp, IL-1beta, IL-8, IL-18, and IFN-gamma were also detected in the broncho-alveolar lavage fluids (BALF). The antigen-specific IFN-gamma and IL-10 responses after infection of Mhp were gradually suppressed during Mhp infection as well as non-specific immune response to concanavalin A (ConA) and lipopolysacchalide (LPS) at early stage of infection. These results suggested that Mhp infection modulates the immune response of pigs by inducing several cytokines, and causes immuno-suppression of pigs in a gnotobiotic condition.