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IntroductionWidespread Fasciola gigantica infection in buffaloes has caused great economic losses in buffalo farming. Studies on F. gigantica excretory and secretory products (FgESP) have highlighted their importance in F. gigantica parasitism and their potential in vaccine development. Identifying FgESP components involved in F. gigantica-buffalo interactions during different periods is important for developing effective strategies against fasciolosis.MethodsBuffaloes were assigned to non-infection (n = 3, as control group) and infection (n = 3) groups. The infection group was orally administrated 250 metacercariae. Sera were collected at 3, 10, and 16 weeks post-infection (wpi) for the non-infection group and at 0 (pre-infection), 1, 3, 6, 8, 10, 13, and 16 wpi for the infection group. FgESP components interacting with sera from the non-infection and infection groups assay were pulled down by co-IP and identified using LC–MS/MS. Interacting FgESP components in infection group were subjected to Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathway and gene ontology (GO) functional annotation to infer their potential functions.Results and discussionProteins of FgESP components identified in the non-infection group at 3, 10, and 16 wpi accounted for 80.5%, 84.3%, and 82.1% of all proteins identified in these three time points, respectively, indicating surroundings did not affect buffalo immune response during maintenance. Four hundred and ninety proteins were identified in the infection group, of which 87 were consistently identified at 7 time points. Following GO analysis showed that most of these 87 proteins were in biological processes, while KEGG analysis showed they mainly functioned in metabolism and cellular processing, some of which were thought to functions throughout the infection process. The numbers of specific interactors identified for each week were 1 (n = 12), 3 (n = 5), 6 (n = 8), 8 (n = 15), 10 (n = 23), 13 (n = 22), and 16 (n = 14) wpi, some of which were thought to functions in specific infection process. This study screened the antigenic targets in FgESP during a dense time course over a long period. These findings may enhance the understanding of molecular F. gigantica-buffalo interactions and help identify new potential vaccine and drug target candidates.

BackgroundFasciola gigantica, a tropical liver fluke, infects buffalo in Asian and African countries, causing significant economic losses and posing public health threats. The diagnostic of buffalo fascioliasis caused by F. gigantica is vital in fascioliasis control and preventation. The 22nd gel filtration chromatography fraction of F. gigantica Excretory-Secretory Products (FgESP), namely Fasciola 22 (F22), which was used as a diagnostic antigen in indirect ELISA, has demonstrated great potential for fascioliasis diagnosing. In the absence of rapid diagnostic methods, the use of a colloidal gold immunochromatographic strip based on F22 was applied to detect F. gigantica infection in buffalo.MethodsIn the present study, the 22nd gel filtration chromatography fraction of FgESP (F22) was used as an antigen to establish the colloidal gold-based immunochromatographic strip (ICS). The nitrocellulose membrane was incubated with F22 at the test line (T line) and goat anti-mouse secondary antibody at the control line (C line). The mouse anti-buffalo secondary antibody 2G7 conjugated to colloidal gold particles was used as the detection system for line visualization. The strip was assembled and developed by optimizing reaction conditions. The sensitivity, specificity, stability, and early diagnostic value of the strip were evaluated employing buffalo-derived sera.ResultsAn immunochromatographic strip for the rapid detection of antibodies against F. gigantica-FgICS was developed. The strip demonstrated high sensitivity and specificity. Sensitivity tests confirmed positive results even when the positive reference serum was diluted 4,096 times. Except for one Schistosoma japonicum-positive serum that tested positive via FgICS, specificity tests confirmed no cross-reactivity with other positive sera of Schistosoma japonicum and Babesia bovis. The strip remained stable after storage at 4°C for up to 3 months. In infected buffalo, antibodies could be detected as early as 14–21 days post-infection. The detection of 17 positive sera yielded an 82.4% positive rate via FgICS vs. a 100.0% positive rate via ELISA based on FgESP. For FgICS, the 95% confidence interval of sensitivity was 84.8–95.4%, while specificity was 4.2–14.7%.ConclusionThe immunochromatographic strip FgICS developed in this study provides a simple and rapid method of F. gigantica antibody detection and infected buffalo monitoring in the field.

Additional file 1: Table S1. Sequence information of primers used in qPCR validation. Table S2. Summary of statistics for the quality control of sequencing data from adult male and female worms of ivermectin resistant and susceptible strains of Haemonchus contortus. Table S3. Summary of total reads in the transcriptome data from RNA sequencing and those mapped to the Haemonchus contortus genome. Table S4. Information on the fold changes for 114 differentially expressed genes identified in both RM_vs_SM and RF_vs_SF comparisons of Haemonchus contortus. Table S5. Top 10 up-regulated (up) and down-regulated (down) genes based on fold changes in the ivermectin (IVM)-resistant male (RM) versus IVM-susceptible male (SM) and IVM-resistant female (RF) versus IVM-susceptible female (SF) worms of Haemonchus contortus. Table S6. Information on differentially expressed genes in the ivermectin-resistant male and female worms of Haemonchus contortus encoding receptors, transporters, and detoxification enzymes. Table S7. Information on differentially expressed genes in the ivermectin-resistant male and female worms of Haemonchus contortus encoding molecules involved in lipid metabolism and cuticle collagen formation.

Additional file 1: Table S1. PCR Primers used in the present study. Table S2. Sequences of DAF-5 homologues used for alignment and phylogenetic analyses. Table S3. Sequences of Hc-daf-5-specific siRNA and control siRNA used in RNAi.

Additional file 1: Table S1. Primers used for PCR-amplification of target gene and for real-time PCR analysis. Table S2. Sequences of SMAD4 homologues from all species used for alignment and phylogenetic analysis. Table S3. Sequences of Hc-daf-3-specific siRNA and control siRNA used for RNA interference. Table S4. Sequence identities of Hc-DAF-3 and its MH1 domain and MH2 domain relative to homologues from selected metazoan species.