quil-a and Fascioliasis

Fluke burdens, faecal egg output and hepatic damage were assessed in goats immunised with recombinant cathepsin L1 (rCL1) plus Quil A (n=7) or Quil A alone (control; n=7) and challenged with Fasciola hepatica. There were no significant differences in fluke burdens (56+/-26 vs. 92+/-53), average fluke length (20.9+/-3.0 vs. 21.0+/-3.4mm) or faecal egg output (525+/-533 vs. 758+/-677.8 eggs per gram at 17 weeks post-infection) between vaccinated and infected control groups, respectively, but high individual variability was evident. Morphometric and histopathological studies showed reduced hepatic damage in the vaccinated group compared to the infected control, but high individual variability was also recorded. Further vaccine trials in goats should be carried out using a larger number of animals to evaluate rCL1 as a vaccine for fasciolosis in goats.

Topics: Adjuvants, Immunologic; Animals; Cathepsins; Fasciola hepatica; Fascioliasis; Feces; Female; Goat Diseases; Goats; Immunization; Liver; Parasite Egg Count; Quillaja Saponins; Recombinant Proteins; Saponins

Glutathione S-transferase (GST) from the liver fluke Fasciola hepatica was assessed as a vaccine immunogen in cattle in a number of immunological adjuvants. Significant reductions in fluke burdens (49-69%) were only observed in cattle vaccinated with GST in Quil Alsqualene Montanide (SM) and PLG microspheres in SM but there was no correlation between anti-GST IgG titres and protection. In separate experiments, animals vaccinated with GST in Quil AlSM were still significantly protected (48%, P < 0.05) 6 months after boosting and no significant differences in protection were seen when the metacercarial challenge was given over 1 month instead of as a single bolus. Inhibition of GST enzyme activity in vitro by cattle antisera did not correlate with reduced fluke burdens.

Topics: Adjuvants, Immunologic; Animals; Antibodies; Body Burden; Cattle; Cattle Diseases; Enzyme-Linked Immunosorbent Assay; Fasciola hepatica; Fascioliasis; Glutathione Transferase; Male; Quillaja Saponins; Saponins; Titrimetry; Trematoda; Vaccination

Leucine aminopeptidase (FhLAP) and cathepsin L1 (FhCL1) of Fasciola hepatica play a critical role in parasite feeding, migration through host tissue, and immune evasion. These antigens have been tested for immune protection as single components with variable degrees of success. The chimeric-protein approach could improve protection levels against fasciolosis. Previously, we reported the design and construction of a chimeric protein composed of antigenic sequences of FhLAP and FhCL1 of F. hepatica. The goal of the present study was to express and evaluate the immune-protective capacity of this chimeric protein (rFhLAP-CL1) in sheep. Animals were randomly allocated into five groups with five animals in each group. Groups 1, 2 and 3 were immunized twice with 100 μg, 200 μg and 400 μg of rFhLAP-CL1 emulsified with Quil A adjuvant, whereas groups 4 and 5 were the adjuvant control and infection control groups, respectively. The animals were then challenged with 200 metacercariae two weeks after the rFhLAP-CL1 booster. The fluke burden was reduced by 25.5%, 30.7% (p < 0.05) and 46.5% (p < 0.01) in sheep immunized with 100 μg, 200 μg and 400 μg of chimeric protein, respectively, in comparison to the infection control group. There was a reduction of 22.7% (p < 0.05) and 24.4% (p < 0.01) in fecal egg count in groups 2 and 3, respectively, compared to the infection control group. Sheep immunized with chimeric protein produced F. hepatica excretion-secretion product-specific total IgG antibody, which were increased after challenge. Moreover, the levels of rFhLAP-CL1-specific IgG1 and IgG2 isotypes in immunized sheep increased rapidly two weeks after the first immunization and were significantly more elevated than those of the control groups, indicating a mixed Th1/Th2 response. This is a preliminary evaluation of the chimeric protein rFhLAP-CL1 as a possible immunogen against F. hepatica infection in sheep.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Helminth; Cathepsin L; Fasciola hepatica; Fascioliasis; Feces; Immunization, Secondary; Immunoglobulin G; Leucyl Aminopeptidase; Male; Parasite Egg Count; Quillaja Saponins; Recombinant Fusion Proteins; Sheep; Sheep Diseases; Th1 Cells; Th2 Cells

Thirty goats were randomly allocated in five groups of six animals each, for immunization with 1 × 10(14) phage particles of clones 11, 13, and 13 with Quil A adjuvant and wild-type M13KE phage at the beginning and 4 weeks later. The control group received phosphate-buffered saline. All groups were challenged with 200 metacercariae at week 6 and slaughtered 14 weeks later. The mean worm burdens after challenge were reduced by 46.91% and 79.53% in goats vaccinated with clones 13 and 13 with Quil A (P < 0.05), respectively; no effect was observed in animals immunized with clone 11 and M13KE phage. Animals receiving clones 11, 13, and 13 with Quil A showed a significant reduction in eggs output. Vaccinated animals produced parasite-specific total IgG antibody which were boosted after challenge with metacercariae of F. hepatica. Furthermore, levels of anti-phage total IgG increased rapidly within 2 weeks of the first vaccination and were always significantly higher in all vaccinated goats than in the infected control group. The fluke burden of goats immunized with clones 13 and 13 with Quil A was significantly correlated with IgG2 and total IgG. Goats vaccinated with phage clones produced significantly high titres of IgG1 and IgG2 antibodies indicating a mixed Th1/Th2 response. These data indicate that cathepsin L1 mimotopes has a potential as a vaccine candidate against Fasciola hepatica, whose efficacy will be evaluated in other host species, including those of veterinary importance.

Topics: Adjuvants, Immunologic; Amino Acid Sequence; Animals; Antibodies, Helminth; Cathepsin L; Fasciola hepatica; Fascioliasis; Goats; Immunity, Humoral; Immunoglobulin G; Molecular Sequence Data; Parasite Egg Count; Peptide Library; Quillaja Saponins; Saponins; Th1 Cells; Th2 Cells; Vaccines

Changes and local immune response were evaluated in the peritoneal cell populations, duodenal lamina propria and liver from goats immunized with recombinant glutathione transferase sigma class (rFhGST-S1) during early stages of infection with Fasciola hepatica. Group 1 (n=7) was unimmunized and uninfected; group 2 (n=10) was immunized with adjuvant Quil A and infected; group 3 (n=10) was immunised with rFhGST-S1 and infected. Three goats from each group were killed at 7-9 days post-infection (dpi) to evaluate early changes and immune response. The remaining goats were killed at 15 weeks post-infection (wpi). rFhGST-S1 vaccination induced variable response: three goats showed low fluke burden at 15 wpi and two goats showed low hepatic damage at early infection stages. This response was associated to a severe infiltrate of eosinophils in peritoneal fluid and hepatic necrotic foci, high iNOS expression in peritoneal cells and abundant infiltrate of eosinophils surrounding hepatic migrating flukes. T lymphocyte subsets were found in the vicinity of necrotic areas but they were absent in the vicinity of migrating larvae. No significant variation for T cell subsets, except for CD4 and γδ T lymphocytes, that were higher in the Quil A group compared to the rFhGST-S1 group. Expression of IL4 and IFN-γ in the hepatic inflammatory infiltrates was very occasional.

Topics: Adjuvants, Immunologic; Animals; Fasciola hepatica; Fascioliasis; Glutathione Transferase; Goat Diseases; Goats; Immunity, Cellular; Immunity, Humoral; Liver; Male; Peritoneum; Quillaja Saponins; Saponins; Vaccines; Vaccines, Synthetic

Protection against Fasciola hepatica in goats immunized with Peroxiredoxin (Prx) was assessed. The experimental trial consisted of three groups of seven animals; group 1 were unimmunized and uninfected, group 2 were immunized with adjuvant only and group 3 were immunized with recombinant Prx in adjuvant (immunized and infected). Immunization with Prx in Quil A adjuvant, group 3, induced a reduction in fluke burden of 33.04% when compared to adjuvant control, group 2, although this difference was not significant. The hepatic gross and microscopical morphometric study revealed lower damage in the Prx-immunized compared to group 2 (p<0.05). Furthermore, immunohistochemical studies revealed that the Prx-immunized group exhibited reduced infiltration of CD4(+), CD8(+), IFN-gamma(+) and TCR(+) (p<0.05); and CD2(+) and IL-4(+) (p<0.001) in hepatic lesions. Levels of anti-Prx serum IgG in group 3 showed a significant increase at the 4th week after challenge infection compared with group 2 (p<0.0001). This is the first report of ruminant immunization with recombinant Prx of F. hepatica. The study shows that this vaccine significantly reduces hepatic damage and encourages further studies to improve the vaccine efficacy.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Helminth; CD2 Antigens; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Fasciola hepatica; Fascioliasis; Goat Diseases; Goats; Immunoglobulin G; Immunohistochemistry; Interferon-gamma; Interleukin-4; Liver; Peroxiredoxins; Quillaja Saponins; Receptors, Antigen, T-Cell; Saponins; Vaccination; Vaccines

Fasciola hepatica infection causes significant clinical disease in ruminants. Current control methods, based on flukicidal drugs, are becoming less useful because of resistance in fluke populations. Vaccination would be a viable alternative, but as yet no vaccine to protect ruminants against liver fluke infection has been commercialised. Adjuvants can be used to enhance and promote protective immune responses by vaccines. In previous vaccination trials, we have observed a distinct adjuvant effect, or a degree of protection, in animals administered adjuvant alone in the absence of any specific F. hepatica antigen. Understanding this effect will be important for continuing efforts to develop vaccines effective against fasciolosis. This study investigated the effects of three adjuvants (Quil A, Freund's Incomplete and TiterMax Gold) on the course of experimental F. hepatica infection in 6-month-old sheep (n=33). At completion of the trial, all animals were necropsied to determine fluke burden and fluke weight. Quil A administration led to a significant reduction in faecal egg count (P<0.0001) and significantly higher parasite-specific serum antibody activity for all isotypes measured (P<0.01). This suggests that Quil A, which promotes a Th1 response, may be useful as an adjuvant in anti-Fasciola vaccines. Furthermore, it reinforces the results of our previous studies indicating that enhanced Th1 responsiveness to vaccine antigens is required to achieve protection against challenge by F. hepatica.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Helminth; Fasciola hepatica; Fascioliasis; Feces; Parasite Egg Count; Quillaja Saponins; Random Allocation; Saponins; Sheep; Sheep Diseases; Time Factors; Weight Gain