| Hae-Ji Kang | 8 Articles |
Toxoplasma gondii is an intracellular protozoan parasite capable of causing chronic infection by forming persistent cysts in the brain. Despite its global burden, no approved vaccine exists. Virus-like particle vaccines expressing microneme protein 8 (MIC8) or apical membrane antigen 1 (AMA1) of T. gondii have previously shown efficacy. In this study, we generated recombinant vaccinia viruses (rVVs) expressing MIC8 and AMA1 and evaluated their efficacy against T. gondii ME49 infection. BALB/c mice were intramuscularly immunized with a combination of MIC8 and AMA1 rVVs and challenged orally with T. gondii ME49. Immunization with MIC8+AMA1 rVVs produced a significant increase in T. gondii-specific IgG. Splenocyte analysis revealed robust activation of CD4+ and CD8+ T cells, as well as expansion of memory B cells. The immunized group exhibited an 89.6% reduction in brain cyst count, with significantly improved survival compared to the control group. These findings demonstrate that combining the antigens MIC8 and AMA1 using a vaccinia virus platform can effectively promote both humoral and cellular immunity, supporting its potential as a vaccine strategy against T. gondii ME49.
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Malaria is a global disease affecting a large portion of the world’s population. Although vaccines have recently become available, their efficacies are suboptimal. We generated virus-like particles (VLPs) that expressed either apical membrane antigen 1 (AMA1) or microneme-associated antigen (MIC) of Plasmodium berghei and compared their efficacy in BALB/c mice. We found that immune sera acquired from AMA1 VLP- or MIC VLP-immunized mice specifically interacted with the antigen of choice and the whole P. berghei lysate antigen, indicating that the antibodies were highly parasite-specific. Both VLP vaccines significantly enhanced germinal center B cell frequencies in the inguinal lymph nodes of mice compared with the control, but only the mice that received MIC VLPs showed significantly enhanced CD4+ T cell responses in the blood following P. berghei challenge infection. AMA1 and MIC VLPs significantly suppressed TNF-α and interleukin-10 production but had a negligible effect on interferon-γ. Both VLPs prevented excessive parasitemia buildup in immunized mice, although parasite burden reduction induced by MIC VLPs was slightly more effective than that induced by AMA1. Both VLPs were equally effective at preventing body weight loss. Our findings demonstrated that the MIC VLP was an effective inducer of protection against murine experimental malaria and should be the focus of further development.
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Toxoplasma gondii infections are primarily diagnosed by serological assays, whereas molecular and fluorescence-based techniques are garnering attention for their high sensitivity in detecting these infections. Nevertheless, each detection method has its limitations. The toxoplasmosis detection capabilities of most of the currently available methods have not been evaluated under identical experimental conditions. This study aimed to assess the diagnostic potential of enzyme-linked immunosorbent assay (ELISA), real-time polymerase chain reaction (RT-PCR), immunohistochemistry (IHC), and immunofluorescence (IF) in BALB/c mice experimentally infected with various doses of T. gondii ME49. The detection of toxoplasmosis from sera and brain tissues was markedly enhanced in mice subjected to high infection doses (200 and 300 cysts) compared to those subjected to lower doses (10 and 50 cysts) for all the detection methods. Additionally, increased B1 gene expression levels and cyst sizes were observed in the brain tissues of the mice. Importantly, IHC, IF, and ELISA, but not RT-PCR, successfully detected T. gondii infections at the lowest infection dose (10 cysts) in the brain. These findings may prove beneficial while designing experimental methodologies for detecting T. gondii infections in mice.
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Toxoplasma gondii ME49 infections are typically diagnosed by serological tests. However, serological diagnosis of RH strain-induced toxoplasmosis remains unknown. In order to develop seradiagnosis of above 2 kinds of infections, we generated recombinant virus-like particles (VLPs) displaying the T. gondii rhoptry protein 4 (ROP4) and evaluated their potential in T. gondii ME49 or RH strain infection diagnostics. Mice were orally infected with either the tachyzoites of T. gondii (RH) or cysts of T. gondii (ME49) at various dosages, and sera were collected at regular intervals. ELISA-based serological tests were performed to assess IgG, IgM, and IgA antibody responses against ROP4 VLP antigen and tissue lysate antigen (TLA). Compared to TLA, IgG, IgM, and IgA levels to ROP4 VLP antigen were significantly higher in the sera of T. gondii RH-infected mice 1 and 2 week post-infection (PI). T. gondii-specific IgG antibody was detected at 1, 2, 4, and 8 week PI in the T. gondii ME49-infected mice with infection dose-dependent manner. These results indicated that the ROP4 VLP antigen was highly sensitive antigens detecting T. gondii RH and ME49 antibodies at an early stage.
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Toxoplasma gondii can infect humans worldwide, causing serious diseases in pregnant women and immunocompromised individuals. T. gondii rhoptry protein 13 (ROP13) is known as one of the key proteins involved in host cell invasion. In this study, we generated virus-like particles (VLPs) vaccine expressing T. gondii rhoptry ROP13 and investigated VLPs vaccine efficacy in mice. Mice immunized with ROP13 VLPs vaccine elicited significantly higher levels of T. gondii-specific IgG, IgG1, IgG2a, and IgA antibody responses following boost immunization and challenge infection, whereas antibody inductions were insignificant upon prime immunization. Differing immunization routes resulted in differing antibody induction, as intranasal immunization (IN) induced greater antibody responses than intramuscular immunization (IM) after boost and challenge infection. IN immunization induced significantly higher levels of IgG and IgA antibody responses from feces, antibody-secreting cells (ASCs), CD4+ T, CD8+ T cells and germinal center B cell responses in the spleen compared to IM immunization. Compared to IM immunization, IN immunization resulted in significantly reduced cyst counts in the brain as well as lesser body weight loss, which contributed to better protection. All of the mice immunized through either route survived, whereas all na?ve control mice perished. These results indicate that the ROP13 VLPs vaccine could be a potential vaccine candidate against T. gondii infection.
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Human infections due to the monkey malaria parasite Plasmodium knowlesi is increasingly being reported from most Southeast Asian countries specifically Malaysia. The parasite causes severe and fatal malaria thus there is a need for urgent measures for its control. In this study, the level of polymorphisms, haplotypes and natural selection of full-length pkmsp8 in 37 clinical samples from Malaysian Borneo along with 6 lab-adapted strains were investigated. Low levels of polymorphism were observed across the full-length gene, the double epidermal growth factor (EGF) domains were mostly conserved, and non-synonymous substitutions were absent. Evidence of strong negative selection pressure in the non-EGF regions were found indicating functional constrains acting at different domains. Phylogenetic haplotype network analysis identified shared haplotypes and indicated geographical clustering of samples originating from Peninsular Malaysia and Malaysian Borneo. This is the first study to genetically characterize the full-length msp8 gene from clinical isolates of P. knowlesi from Malaysia; however, further functional characterization would be useful for future rational vaccine design.
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Both Plasmodium spp. and Toxoplasma gondii are important apicomplexan parasites, which infect humans worldwide. Genetic analyses have revealed that 33% of amino acid sequences of inner membrane complex from the malaria parasite Plasmodium berghei is similar to that of Toxoplasma gondii. Inner membrane complex is known to be involved in cell invasion and replication. In this study, we investigated the resistance against T. gondii (ME49) infection induced by previously infected P. berghei (ANKA) in mice. Levels of T. gondii-specific IgG, IgG1, IgG2a, and IgG2b antibody responses, CD4+ and CD8+ T cell populations were found higher in the mice infected with P. berghei (ANKA) and challenged with T. gondii (ME49) compared to that in control mice infected with T. gondii alone (ME49). P. berghei (ANKA) + T. gondii (ME49) group showed significantly reduced the number and size of T. gondii (ME49) cysts in the brains of mice, resulting in lower body weight loss compared to ME49 control group. These results indicate that previous exposure to P. berghei (ANKA) induce resistance to subsequent T. gondii (ME49) infection.
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Toxoplasma gondii is a ubiquitous protozoan parasite responsible for causing toxoplasmosis. Preventive measures for toxoplasmosis are currently lacking and as such, development of novel vaccines are of urgent need. In this study, we generated 2 virus-like particles (VLPs) vaccines expressing T. gondii rhoptry protein 4 (ROP4) or rhoptry protein 18 (ROP18) using influenza matrix protein (M1) as a core protein. Mice were intranasally immunized with VLPs vaccines and after the last immunization, mice were challenged with ME49 cysts. Protective efficacy was assessed and compared by determining serum antibody responses, body weight changes and the reduction of cyst counts in the brain. ROP18 VLPs-immunized mice induced greater levels of IgG and IgA antibody responses than those immunized with ROP4 VLPs. ROP18 VLPs immunization significantly reduced body weight loss and the number of brain cysts in mice compared to ROP4 VLPs post-challenge. These results indicate that T. gondii ROP18 VLPs elicited better protective efficacy than ROP4 VLPs, providing important insight into vaccine design strategy.
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