Toxoplasmosis is an opportunistic infection caused by the protozoan parasite Toxoplasma gondii. T. gondii is widespread globally and causes severe diseases in individuals with impaired immune defences as well as congenitally infected infants. The high prevalence rate in some parts of the world such as South America and Africa, coupled with the current drug treatments that trigger hypersensitivity reactions, makes the development of immunotherapeutics intervention a highly important research priority. Immunotherapeutics strategies could either be a vaccine which would confer a pre-emptive immunity to infection, or passive immunization in cases of disease recrudescence or recurrent clinical diseases. As the severity of clinical manifestations is often greater in developing nations, the development of well-tolerated and safe immunotherapeutics becomes not only a scientific pursuit, but a humanitarian enterprise. In the last few years, much progress has been made in vaccine research with new antigens, novel adjuvants, and innovative vaccine delivery such as nanoparticles and antigen encapsulations. A literature search over the past 5 years showed that most experimental studies were focused on DNA vaccination at 52%, followed by protein vaccination which formed 36% of the studies, live attenuated vaccinations at 9%, and heterologous vaccination at 3%; while there were few on passive immunization. Recent progress in studies on vaccination, passive immunization, as well as insights gained from these immunotherapeutics is highlighted in this review.
Previously, we have identified a gene encoding thrombospondin-related anonymous protein of Babesia gibsoni (BgTRAP), and have shown that the antisera raised against recombinant BgTRAP expressed in Escherichia coli inhibited the growth of parasites. In the present study, a recombinant vaccinia virus expressing the BgTRAP (VV/BgTRAP) was constructed. A specific band with a molecular mass of 80 kDa, which is similar to that of native BgTRAP on the merozoites of B. gibsoni, was detected in the supernatant of VV/ BgTRAP-infected RK13 cells. Mice inoculated with VV/BgTRAP produced a specific antiBgTRAP response. The antiserum against VV/BgTRAP showed reactivity against the native BgTRAP on parasites. These results indicated that the recombinant vaccinia virus expressing BgTRAP might be a vaccine candidate against canine B. gibsoni infection.
Toxoplasmosis, a parasitic disease in human and animals, is caused by Toxoplasma gondii. Our previous study has led to the discovery of a novel RAP domain binding protein antigen (TgRA15), an apparent in-vivo induced antigen recognised by antibodies in acutely infected individuals. This study is aimed to evaluate the humoral response and cytokine release elicited by recombinant TgRA15 protein in C57BL/6 mice, demonstrating its potential as a candidate vaccine for Toxoplasma gondii infection. In this study, the recombinant TgRA15 protein was expressed in Escherichia coli, purified and refolded into soluble form. C57BL/6 mice were immunised intradermally with the antigen and CASAC (Combined Adjuvant for Synergistic Activation of Cellular immunity). Antigen-specific humoral and cell-mediated responses were evaluated using Western blot and ELISA. The total IgG, IgG1 and IgG2a antibodies specific to the antigen were significantly increased in treatment group compare to control group. A higher level of interferon gamma (IFN-γ) secretion was demonstrated in the mice group receiving booster doses of rTgRA15 protein, suggesting a potential Th1-mediated response. In conclusion, the rTgRA15 protein has the potential to generate specific antibody response and elicit cellular response, thus potentially serve as a vaccine candidate against T. gondii infection.
Toxoplasmosis in humans and other animals is caused by the protozoan parasite Toxoplasma gondii. During the process of host cell invasion and parasitophorous vacuole formation by the tachyzoites, the parasite secretes Rhoptry protein 8 (ROP8), an apical secretory organelle. Thus, ROP8 is an important protein for the pathogenesis of T. gondii. The ROP8 DNA was constructed into a pVAX-1 vaccine vector and used for immunizing BALB/c mice. Immunized mice developed immune response characterized by significant antibody responses, antigen-specific proliferation of spleen cells, and production of high levels of IFN-γ (816 ± 26.3 pg/mL). Challenge experiments showed significant levels of increase in the survival period (29 days compared with 9 days in control) in ROP8 DNA vaccinated mice after a lethal challenge with T. gondii. Results presented in this study suggest that ROP8 DNA is a promising and potential vaccine candidate against toxoplasmosis.
In this study, we successfully expressed a chimerical surface antigen 1 and 2 (SAG1/2) of Toxoplasma gondii in Pichia pastoris. Eighty human serum samples, including 60 from confirmed cases of toxoplasmosis, were tested against the purified recombinant SAG1/2 in Western blots. Results of Western blots targeted at Toxoplasma IgG and IgM showed that the recombinant SAG1/2 reacted with all sera from the toxoplasmosis cases but none with the Toxoplasma-negative serum samples. These results showed that the P. pastoris-derived recombinant SAG1/2 was sensitive and specific and suitable for use as antigen for detecting anti-Toxoplasma antibodies. To further investigate the immunological characteristic of the recombinant protein, the recombinant SAG1/2 was injected subcutaneously into BALB/c mice, and their serum was tested against total protein lysate of T. gondii. Mice immunized with the recombinant SAG1/2 reacted specifically with the native SAG1 and SAG2 of T. gondii. Significant proliferation of splenocytes stimulated with tachyzoite total protein lysate was observed in vaccinated BALB/c mice but not in those from negative control mice. Specific production of IFN-γ, the Th1-type cytokines, was also found in stimulated splenocytes from vaccinated mice. These results show that the chimeric protein recombinant SAG1/2 can elicit a Th1-associated protection against T. gondii infections in mice. Finally, vaccinated mice were significantly protected against lethal challenge with live T. gondii RH strain tachyzoites (P
The phylum Apicomplexa comprises obligate intracellular parasites that infect vertebrates. All invasive forms of Apicomplexa possess an apical complex, a unique assembly of organelles localized to the anterior end of the cell and involved in host cell invasion. Previously, we generated a chicken monoclonal antibody (mAb), 6D-12-G10, with specificity for an antigen located in the apical cytoskeleton of Eimeria acervulina sporozoites. This antigen was highly conserved among Apicomplexan parasites, including other Eimeria spp., Toxoplasma, Neospora, and Cryptosporidium. In the present study, we identified the apical cytoskeletal antigen of Cryptosporidium parvum (C. parvum) and further characterized this antigen in C. parvum to assess its potential as a target molecule against cryptosporidiosis. Indirect immunofluorescence demonstrated that the reactivity of 6D-12-G10 with C. parvum sporozoites was similar to those of anti-β- and anti-γ-tubulins antibodies. Immunoelectron microscopy with the 6D-12-G10 mAb detected the antigen both on the sporozoite surface and underneath the inner membrane at the apical region of zoites. The 6D-12-G10 mAb significantly inhibited in vitro host cell invasion by C. parvum. MALDI-TOF/MS and LC-MS/MS analysis of tryptic peptides revealed that the mAb 6D-12-G10 target antigen was elongation factor-1α (EF-1α). These results indicate that C. parvum EF-1α plays an essential role in mediating host cell entry by the parasite and, as such, could be a candidate vaccine antigen against cryptosporidiosis.
AbstractToxoplasma gondii infects a broad range of warm-blooded hosts, including humans. Important clinical manifestations include encephalitis in immunocompromised patients as well as miscarriage and fetal damage during early pregnancy. Toxoplasma gondii dense granule antigen 2 and 5 (GRA2 and GRA5) are essential for parasitophorous vacuole development of the parasite. To evaluate the potential of GRA2 and GRA5 as recombinant DNA vaccine candidates, these antigens were cloned into eukaryotic expression vector (pcDNA 3.1C) and evaluated in vaccination experiments. Recombinant DNA vaccines constructed with genes encoding GRAs were validated in Chinese hamster ovary cells before evaluation using lethal challenge of the virulent T. gondii RH strain in BALB/c mice. The DNA vaccines of pcGRA2 and pcGRA5 elicited cellular-mediated immune response with significantly higher levels of interferon-gamma, interleukin-2 (IL-2), IL-4, and IL-10 (P < 0.05) compared with controls. A mixed T-helper cell 1 (Th1)/Th2 response was associated with slightly prolonged survival. These findings provide evidence that DNA vaccination with GRA2 and GRA5 is associated with Th1-like cell-mediated immune responses. It will be worthwhile to construct recombinant multiantigen combining full-length GRA2 or/and GRA5 with various antigenic proteins such as the surface antigens and rhoptry antigens to improve vaccination efficacy.