Qualitative analysis of Fasciola gigantica excretory and secretory products coimmunoprecipitated with buffalo secondary infection sera shows dissimilar components from primary infection sera

•Fasciola gigantica excretory and secretory products (FgESPs) coimmunoprecipitated from buffalo secondary infection sera contain different components than primary infection sera.•324 proteins were coimmunoprecipitated in FgESP by buffalo secondary infection sera.•Protein pathway analysis indicated that 324 proteins were mainly involved in the cellular processes and metabolism of F. gigantica.•14–3–3θ was consistently identified in secondary infection and may be involved in various cellular processes and innate immune signaling pathways.•Members of the HSP family were identified in both primary and secondary infection, indicating it may function in both primary and secondary infection processes. Buffaloes cannot mount a robust adaptive immune response to secondary infection by Fasciola gigantica. Even if excretory and secretory products (ESPs) exhibit potent immunoregulatory effects during primary infection, research on ESPs in secondary infection is lacking, even though the ESP components that are excreted/secreted during secondary infection are unknown. Therefore, qualitative analysis of ESP during secondary infection was performed and compared with that of primary infection to deepen the recognition of secondary infection and facilitate immunoregulatory molecules screening. Buffaloes were divided into three groups: A (n = 3, noninfected), B (n = 3, primary infection) and C (n = 3, secondary infection). Buffaloes in the primary (0 weeks post infection; wpi) and secondary (-4 and 0 wpi) infection groups were infected with 250 metacercariae by oral administration. Then, sera were collected from groups at different wpi, and interacting proteins were precipitated by coimmunoprecipitation (Co-IP), qualitatively analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and annotated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses to infer their potential functions. In group C, 324 proteins were identified, of which 76 proteins were consistently identified across 7 time points (1, 3, 6, 8, 10, 13, and 16 wpi). Compared with 87 proteins consistently identified in group B, 22 proteins were identified in group C. Meanwhile, 34 proteins were only identified in group C compared to 200 proteins identified in group B. Protein pathway analysis indicated that these proteins were mainly involved in the cellular processes and metabolism of F. gigantica. Among them, 14–3–3θ was consistently identified in group C and may be invo