Abstract: In order to explore the dynamic changes of peripheral blood mononuclear cell（PBMC） cytokines after primary and secondary infection with Fasciola gigantica, in the experiment, six 8-10 months old hybrid healthy water buffalo were selected and divided into primary infection group and secondary infection group. The primary infection group was orally infected with 250 metacercariae at week 4, and the secondary infection group was orally infected with 250 metacercariae at week 0 and 4. ELISA method was used to detect the success of infection. Jugular vein blood was collected and PBMC was isolated from buffalo in the primary infection group at 4（pre-infection）-10, 15 and 20 weeks and in the secondary infection group at 0（pre-infection）-10, 15 and 20 weeks. The RNA was extracted and amplified by real-time quantitative PCR. The relative expression levels of PBMC marker immune-related cytokines--Th1 γ interferon（IFN-γ）, Th2interleukin-4（IL-4）, Treg interleukin-10（IL-10）, transforming growth factor-β（TGF-β） were analyzed in different weeks. The results showed that the buffalo in the primary infection group and the secondary infection group were confirmed to be infected at the 6th and 3rd week, respectively, and the animal model was successfully constructed. Compared with pre-infection, the relative expression level of IFN-γ gene in the primary infection group was significantly decreased at the 6th week（P＜0.05）, significantly increased at the 15th week（P＜0.01）, and significantly increased at the 20th week（P＜0.05）, while there was no significant difference in the secondary infection group during the whole experiment period（P＞0.05）. The relative expression of IL-4 gene was significantly increased at weeks 15 and 20（P＜0.01） in the primary infection group, and at week 15（P＜0.01） and at week 20（P＜0.05）. The relative expression level of IL-10 gene in primary infection group was significantly increased at the 5th week（P＜0.05）. The secondary infection group was significantly increased at the 15th week（P＜0.05）. The relative expression level of TGF-β gene in the primary infection group was significantly increased at the 5th week（P＜0.01）, and that in the secondary infection group was significantly increased at the 5th week（P＜0.05）. Compared with the secondary infection group, the relative expression of IFN-γ, IL-4 and TGF-β genes was not significantly different（P＞0.05）, the relative expression of IL-10 gene was significantly different in week 15（P＜0.05）, and the relative expression of IFN-γ gene showed different trends in week 5. These results indicated that the immune responses of buffalo were similar to those of buffalo infected with buffalo once and twice. It was suggested that buffalo were susceptible to Fasciola gigantica infected with Fasciola gigantica once because Fasciola gigantica were not able to induce an effective immune response against Fasciola gigantica.