Study on the effect of Koumiss on the intestinal flora of mice infected with Toxoplasma gondii

Toxoplasma gondii is a worldwide food-borne parasite that can infect almost all warm-blooded animals, including humans. To date, there are no effective drugs to prevent or eradicate T. gondii infection. Recent studies have shown that probiotics could influence the relationship between the microbiota and parasites in the host. Koumiss has been used to treat many diseases based on its probiotic diversity. Therefore, we explored the effect of koumiss on T. gondii infection via its effect on the host intestinal flora. BALB/c mice were infected with T. gondii and treated with PBS, koumiss and mares’ milk. Brain cysts were counted, and long-term changes in the microbiota and the effect of koumiss on gut flora were investigated with high-throughput sequencing technology. The results suggested that koumiss treatment significantly decreased the cyst counts in the brain (P < 0.05). Moreover, T. gondii infection changed the microbiota composition, and koumiss treatment increased the relative abundance of Lachnospiraceae and Akkermansia muciniphila, which were associated with preventing T. gondii infection. Moreover, koumiss could inhibit or ameliorate T. gondii infection by increasing the abundance of certain bacteria that control unique metabolic pathways. The study not only established a close interaction among the host, intracellular pathogens and intestinal flora but also provided a novel focus for drug development to prevent and eradicate T. gondii infection.

Protective Immunity Induced by TgMIC5 and TgMIC16 DNA Vaccines Against Toxoplasmosis

Toxoplasma gondii is an obligate intracellular parasite, which is responsible for a widely distributed zoonosis. Effective vaccines against toxoplasmosis are necessary to protect the public health. The aim of this study is to evaluate the immune efficacy of DNA vaccines encoding TgMIC5 and TgMIC16 genes against T. gondii infection. The recombinant plasmid pVAX-MIC5 and pVAX-MIC16 were constructed and injected intramuscularly in mice. The specific immune responses and protection against challenge with T. gondii RH tachyzoites were evaluated by measuring the cytokine levels, serum antibody concentrations, lymphocyte proliferation, lymphocyte populations, and the survival time. The protection against challenge with the T. gondii RH tchyzoites and PRU cysts was examined by evaluation of the reduction in the brain cyst burden. The results indicated that immunized mice showed significantly increased levels of IgG, IFN-γ, IL-2, IL-12p70, and IL-12p40 and percentages of CD4+ and CD8+ T cells. Additionally, vaccination prolonged the mouse survival time and reduced brain cysts compared with controls. Mouse groups immunized with a two-gene cocktail of pVAX-MIC5 + pVAX-MIC16 were more protected than mouse groups immunized with a single gene of pVAX-MIC5 or pVAX-MIC16. These results demonstrate that TgMIC5 and TgMIC16 induce effective immunity against toxoplasmosis and may serve as a good vaccine candidate against T. gondii infection.

Valproic acid inhibits chronic Toxoplasma infection and associated brain inflammation in mice

Individuals infected with Toxoplasma gondii ( T. gondii ) are prone to psycho-behavioral disorders, most notably schizophrenia and bipolar. Valproic acid reportedly inhibited the proliferation of T. gondii tachyzoites in vitro. However, animals treated with the drug neither lived longer during acute infection nor had fewer brain cysts upon chronic infection. In this study, a quantitative real-time PCR (qPCR) method was applied to quantify copy numbers of BAG1 (a bradyzoite-specific protein), REP529 DNA (a repetitive DNA fragment of the parasite), and SAG1 (a highly expressed tachyzoite-specific surface protein) in brains of chronically infected mice treated by valproic acid. The treatment inhibited the infection and decreased BAG1, SAG1, and REP529 copy numbers in mice brains ( P < 0.0001 ), comparable to Trimethoprim/Sulfamethoxazole (TMP/SMZ), the common medication for Toxoplasmosis treatment. Moreover, valproic acid decreased brain TNF-α expression ( P < 0.0001 ), comparable to TMP/SMZ. Histological examination of mice brains showed a marked reduction in cyst establishment, perivascular infiltration of lymphocytes, and glial nodules to the same level as the TMP/SMZ group. Our results provide direct evidence for the efficacy of valproic acid, a mood-stabilizing and antipsychotic drug against chronic Toxoplasma infection. These results might help modulate therapeutic regimens for neuropsychiatric patients and design more effective anti- Toxoplasma drugs.

Toxoplasma TgATG9 is critical for autophagy and long-term persistence in tissue cysts

Many of the world's warm-blooded species are chronically infected with Toxoplasma gondii tissue cysts, including an estimated one third of the global human population. The cellular processes that permit long-term persistence within the cyst are largely unknown for T. gondii and related coccidian parasites that impact human and animal health. Herein we show that genetic ablation of TgATG9 substantially reduces canonical autophagy and compromises bradyzoite viability. Transmission electron microscopy revealed numerous structural abnormalities occurring in ∆atg9 bradyzoites. Intriguingly, abnormal mitochondrial networks were observed in TgATG9-deficient bradyzoites, some of which contained numerous different cytoplasmic components and organelles. ∆atg9 bradyzoite fitness was drastically compromised in vitro and in mice, with very few brain cysts identified in mice 5 weeks post-infection. Taken together, our data suggests that TgATG9, and by extension autophagy, is critical for cellular homeostasis in bradyzoites and is necessary for long-term persistence within the cyst of this coccidian parasite.

Epidermoid brain cysts: a case report and a literature review

The article is devoted to rare brain pathology, i.e. epidermoid cysts. The histological picture of the tumor is described; the classification is given. On the basis of our own clinical observations and a literature review, the features of the clinical manifestations of cholesteatoma of the cerebellopontine angle, modern approaches to the diagnosis and tactics of complex treatment are presented.

Imiquimod Targets Toxoplasmosis Through Modulating Host Toll-Like Receptor-MyD88 Signaling

Toxoplasma gondii is a prevalent parasite of medical and veterinary importance. Tachyzoïtes and bradyzoïtes are responsible for acute and chronic toxoplasmosis (AT and CT), respectively. In immunocompetent hosts, AT evolves into a persistent CT, which can reactivate in immunocompromised patients with dire consequences. Imiquimod is an efficient immunomodulatory drug against certain viral and parasitic infections. In vivo, treatment with Imiquimod, throughout AT, reduces the number of brain cysts while rendering the remaining cysts un-infectious. Post-establishment of CT, Imiquimod significantly reduces the number of brain cysts, leading to a delay or abortion of reactivation. At the molecular level, Imiquimod upregulates the expression of Toll-like receptors 7, 11, and 12, following interconversion from bradyzoïtes to tachyzoïtes. Consequently, MyD88 pathway is activated, resulting in the induction of the immune response to control reactivated Toxoplasma foci. This study positions Imiquimod as a potent drug against toxoplasmosis and elucidates its mechanism of action particularly against chronic toxoplasmosis, which is the most prevalent form of the disease.

Molecular characterization of Toxoplasma gondii isolates from free-range chickens reveals new genotypes in Goiânia, Goiás, Brazil

The aim of this study was to evaluate the genotypic characteristics of Toxoplasma gondii isolated from free-range chickens in the metropolitan area of Goiânia, Goiás, in Brazil’s central-west region. The seroprevalence rate was found to be 96%, according to an indirect hemagglutination assay. Brain and heart samples were processed by peptic digestion for a mice bioassay. The tissues were homogenized and the resulting samples were subjected to polymerase chain reaction (PCR), which revealed that 64% of them contained the parasite's DNA. The mice bioassay revealed 15 isolates, 8 of them tachyzoites isolates from the peritoneal lavage and 7 from brain cysts. T. gondii genotypes were determined through PCR-RFLP, using the following markers: SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, alt. SAG2, Apico and CS3. Three genotypes were identified, inclued ToxoDB #65, and the other two are not yet described in the literature. Hence, we conclude that the isolates obtained from the metropolitan area of Goiânia showed relatively low genetic diversity.

Computational image analysis reveals the structural complexity of Toxoplasma gondii tissue cysts

Toxoplasma gondii is an obligate intracellular parasite infecting up to one third of the human population. The central event in the pathogenesis of toxoplasmosis is the conversion of tachyzoites into encysted bradyzoites. A novel approach to analyze the structure of in vivo-derived tissue cysts may be the increasingly used computational image analysis. The objective of this study was to quantify the geometrical complexity of T. gondii cysts by morphological, particle, and fractal analysis, as well as to determine if and how it is impacted by parasite strain, cyst age, and host factors. Analyses were performed on 31 images of T. gondii brain cysts of four type-2 strains (the reference Me49 strain and three local isolates, named BGD1, BGD14, and BGD26) using ImageJ software package. The parameters of interest included diameter, circularity, relative particle count (RPC), fractal dimension (FD), lacunarity, and packing density (PD). Although cyst diameter varied widely, its negative correlation with RPC was observed. Circularity was remarkably close to 1, indicating that the shape of the brain cysts was a perfect circle. RPC, FD, and PD did not vary among cysts of different strains, age, and derived from mice of different genetic background. Conversely, lacunarity, which is a measure of heterogeneity, was significantly lower for BGD1 strain vs. all other strains, and higher for Me49 vs. BGD14 and BGD26, but did not differ among Me49 cysts of different age, and derived from genetically different mice. This study is the first application of fractal analysis in describing the structural complexity of T. gondii cysts. Despite all the differences among the analyzed cysts, most parameters remained conserved. Fractal analysis is a novel and widely accessible approach, which along with particle analysis may be applied to gain further insight into T. gondii cyst morphology.

Toxoplasma TgATG9 is critical for autophagy and long-term persistence in tissue cysts

ABSTRACTMany of the world’s warm-blooded species are chronically infected with Toxoplasma gondii tissue cysts, including up to an estimated one third of the global human population. The cellular processes that permit long-term parasite persistence within the cyst are largely unknown, not only for T. gondii but also for related coccidian parasites that impact human and animal health. A previous study revealed an accumulation of autophagic material in the lysosome-like Vacuolar Compartment (VAC) of chronic stage bradyzoites lacking functional cathepsin L protease (TgCPL) activity. Furthermore, it was shown that TgCPL knockout bradyzoites have compromised viability, indicating the turnover of autophagic material could be necessary for bradyzoite survival. However, the extent to which autophagy itself contributes to bradyzoite development and fitness remained unknown. Herein we show that genetic ablation of TgATG9 substantially reduces canonical autophagy and compromises bradyzoite viability. Transmission electron microscopy revealed structural abnormalities occurring in Δatg9 bradyzoites, including disorganization of the inner membrane complex and plasma membrane, the occurrence of multiple nuclei within a single bradyzoite cell, as well as various anomalies associated with the VAC. TgATG9-deficient bradyzoites accumulated significantly less undigested material in the VAC upon inhibition of TgCPL activity, suggesting that autophagy contributes material to the VAC for degradation. Intriguingly, abnormal mitochondria networks were observed in TgATG9-deficient bradyzoites. They were thin and elongated and often adopted a horseshoe conformation. Some abnormal mitochondrial structures were found to contain numerous different cytoplasmic components and organelles. Bradyzoite fitness was found to be drastically compromised, both in vitro and in mice, with very few brain cysts identified in mice 5 weeks post-infection. Taken together, our data suggests that TgATG9, and by extension autophagy, is critical for cellular homeostasis in bradyzoites and is necessary for long-term persistence within the cyst of this coccidian parasite.