Fig 1.
Expression of SjTH in parasites at different developmental stages.
(A) Relative mRNA expression of SjTH in the five developmental stages of S. japonicum, cercariae (C), eggs (E), adult females (F), adult males (M, 42 days post-infection) and hepatic schistosomula (S, 14 days post-infection), was detected by qRT-PCR. Data represents the mean + standard deviation. *, #, Δ, and Φ indicate S group was compared with the E, C, M and F groups, respectively. *p < 0.01, ##, ΔΔ, ΦΦp < 0.01 (Mann-Whitney test). (B) Expression of SjTH and Actin in the five developmental stages of S. japonicum detected by western blot.
Fig 2.
Localization of SjTH in schistosome parasites.
Cryosections of anterior (A) of male adults, and mid-section of male adults (B, E), female adults (C), and hepatic schistosomula (D) were incubated with rabbit anti-SjTH polyclonal antibodies (A, B, C) or an IgG control (D), followed by Alexa Fluor 555 donkey anti-rabbit IgG (green fluorescence). The samples were counterstained with DAPI (in blue). Positive staining is indicated by red arrows. Results are representative of parasites from three independent experiments, and at least 3 worms of each experiment were analyzed for each stage. Abbreviations: F, female adult; M, male adult; S, hepatic schistosomula; I, intestine; VS, ventral sucker.
Fig 3.
Levels of SjTH-specific antibodies in host serum.
(A, B) BABL/c mice and New Zealand white rabbits were percutaneously infected with cercariae (40 ± 2 parasites per mouse and 1,000 ± 100 parasites per rabbit). Sera from infected animals were collected on days 0, 7, 14, 21, 28, 35, 42, and 56 post infection (dpi). Antibody titers against SjTH in these sera were determined by ELISA. The linear charts show the detailed dynamics of antibodies against SjTH. *, #, and Δ indicate comparisons with day 0, 28, and 42 dpi, respectively. ***, ###, ΔΔΔ indicate p < 0.0001. (C, D) SjTH-specific antibodies (C) and SjSP-13-specific antibodies (D) in human serum samples were determined by ELISA. Sera from patients with confirmed schistosomiasis japonica (Sj, n = 20), from the same patients 3 months after praziquantel treatment (Sj-3M, n = 20), from patients with confirmed echinococcosis (Es, n = 15), and from healthy individuals (healthy, n = 20) were included in the assay. The cutoff value for positive results was set at ≥ 2.1-times the mean optical density (OD) value of healthy individuals (dotted lines). Red and black dots indicate positive and negative individuals.
Fig 4.
Evaluation of the protective effect of SjTH in S. japonicum-infected mice.
To assess the protective role of SjTH as a vaccine candidate against schistosome infection, mice (n = 10 per group) were immunized with His-tagged recombinant SjTH, SjSP-13 (a novel diagnostic marker for schistosomiasis), or phosphate-buffered saline (PBS) as control. After immunization, the mice were challenged with cercariae (40 ± 2 per mouse). The blank group was immunized with PBS and received no cercariae. Results are representative of two independent experiments. The weight of the bodies (A), spleens (B) and livers (C) were determined and compared between the groups at 42 days post infection. The number of perfused adult worms (D) and liver eggs (E) was determined by microscopy at 42 days post infection. (F) Granuloma formation in livers was detected by haematoxylin-eosin staining. The proportion of total area of granulomas was compared between the groups. The results are representative of two independent experiments. Data are presented as mean±SD.* indicates p < 0.05, ** p < 0.01, and *** p < 0.0001 as compared with the control (PBS) group.
Fig 5.
SjTH immunization reduces cytokines of both Th1 and Th2 response in the sera of S. japonicum-infected mice.
Mice (n = 10 per group) were immunized with His-tagged recombinant SjTH, SjSP-13, or PBS as control, and were then challenged with cercariae (40 ± 2 per mouse). The blank group was immunized with PBS and received no cercariae. Results are representative of two independent experiments. The levels of interferon (IFN)-γ (A), tumor necrosis factor (TNF)-α (B) interleukin (IL)-4 (C) and IL-10 (D) in the sera 42 days post infection were determined by ELISA and compared between the groups. # and * indicate comparisons with the blank and control (PBS) groups, respectively. * indicates p < 0.05, **, ## p < 0.01, and *** p < 0.0001.