Serosurvey and Molecular Detection of Toxoplasma gondii in Dogs in Rural Areas of Kazeroun District, Fars Province, Southern Iran

Background. Toxoplasma gondii is an intracellular protozoan parasite responsible for systemic disease in a wide range of warm-blooded animals. The current study is aimed at evaluating the prevalence of Toxoplasma infection in dogs, using serological and molecular methods in rural areas in Kazeroun Township, Fars province, southern Iran. Methods. Blood samples were obtained from 60 clinically healthy dogs with an age range of 1 to 7 years in three rural areas of Fars province, southern Iran. Sera and buffy coats were used to assess the T. gondii infection using both modified agglutination test (MAT) and real-time PCR. Results. Antibodies against T. gondii were detected in 5 out of 60 (8.3%) dogs by the MAT method, and T. gondii DNA was detected in 17 out of 60 (28.3%) studied animals. There was no significant association between sex and seropositivity to Toxoplasma ( p > 0.05 ). Fair agreement ( kappa = 0.27 ) was seen between molecular and serological findings where three dogs with positive serological results had a positive molecular test. Conclusion. Findings of the present study show a relatively high prevalence of T. gondii infection in dogs in rural areas in Fars province, southern Iran. Finding the parasite genotype in dogs deserves further study.

Gut microbiota composition does not associate with Toxoplasma infection in rats

Toxoplasma infection in intermediate host species closely associates with inflammation. This association has led to suggestions that the behavioural changes associated with infection may be indirectly driven by the resulting sustained inflammation rather than a direct behavioural manipulation by the parasite. If this is correct, sustained inflammation in chronically infected rodents should present as widespread changes in the gastrointestinal microbiota due to the dependency between the composition of these microbiota and sustained inflammation. We conducted a randomized controlled experiment in rats that were assigned to a Toxoplasma-treatment, placebo-treatment or negative control group. We sacrificed rats during the chronic phase of infection, collected their cecal stool samples and sequenced the V3-V4 region of the 16S rRNA gene to characterise the bacterial community in these samples. Toxoplasma infection did not induce widespread changes in the bacterial community composition of the gastrointestinal tract of rats. Rather, we found sex differences in the bacterial community composition and only minor changes in Toxoplasma infected rats. We conclude that it is unlikely that sustained inflammation is the mechanism driving the highly specific behavioural changes observed in Toxoplasma-positive rats.

Dense granule protein, GRA64 interacts with host cell ESCRT proteins during Toxoplasma gondii infection

The intracellular parasite Toxoplasma gondii adapts to diverse host cell environments within a replicative compartment that is heavily decorated by secreted proteins. In attempts to identify novel parasite secreted proteins that influence host cell activity, we identified and characterized a trans-membrane dense granule protein dubbed GRA64 (TGME49_202620). We found that GRA64 is on the parasitophorous vacuolar membrane (PVM) and is partially exposed to the host cell cytoplasm in both tachyzoite and bradyzoite parasitophorous vacuoles. Using co-immunoprecipitation and proximity-based biotinylation approaches, we demonstrate that GRA64 appears to interact with certain components of the host Endosomal Sorting Complexes Required for Transport (ESCRT). Genetic disruption of GRA64 does not affect acute Toxoplasma virulence in mice nor encystation as observed via tissue cyst burdens in mice during chronic infection. However, ultrastructural analysis of Dgra64 tissue cysts using electron tomography revealed enlarged vesicular structures underneath the cyst membrane, suggesting a role for GRA64 in organizing the recruitment of ESCRT proteins and subsequent intracystic vesicle formation. This study uncovers a novel host-parasite interaction that contributes to an emerging paradigm in which specific host ESCRT proteins are recruited to the limiting membranes (PVMs) of tachyzoite and bradyzoite vacuoles formed during acute and chronic Toxoplasma infection.

Effect of 17β-estradiol, Progesterone, and Tamoxifen on Neurons Infected with Toxoplasma gondii In Vitro

Toxoplasma gondii (T. gondii) is the causal agent of toxoplasmosis, which produces damage in the central nervous system (CNS). Toxoplasma–CNS interaction is critical for the development of disease symptoms. T. gondii can form cysts in the CNS; however, neurons are more resistant to this infection than astrocytes. The probable mechanism for neuron resistance is a permanent state of neurons in the interface, avoiding the replication of intracellular parasites. Steroids regulate the formation of Toxoplasma cysts in mice brains. 17β-estradiol and progesterone also participate in the control of Toxoplasma infection in glial cells in vitro. The aim of this study was to evaluate the effects of 17β-estradiol, progesterone, and their specific agonists–antagonists on Toxoplasma infection in neurons in vitro. Neurons cultured were pretreated for 48 h with 17β-estradiol or progesterone at 10, 20, 40, 80, or 160 nM/mL or tamoxifen 1 μM/mL plus 17β-estradiol at 10, 20, 40, 80, and 160 nM/mL. In other conditions, the neurons were pretreated during 48 h with 4,4′,4″-(4-propyl-[1H] pyrozole-1,3,5-triyl) trisphenol or 23-bis(4-hydroxyphenyl) propionitrile at 1 nM/mL, and mifepristone 1 µM/mL plus progesterone at 10, 20, 40, 80, and 160 nM/mL. Neurons were infected with 5000 tachyzoites of the T. gondii strain RH. The effect of 17β estradiol, progesterone, their agonists, or antagonists on Toxoplasma infection in neurons was evaluated at 24 and 48 h by immunocytochemistry. T. gondii replication was measured with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay. 17β-Estradiol alone or plus tamoxifen reduced infected neurons (50%) compared to the control at 48 h. Progesterone plus estradiol decreased the number of intracellular parasites at 48 h of treatment compared to the control (p < 0.001). 4,4′,4″-(4-propyl-[1H] pyrozole-1,3,5-triyl) trisphenol and 23-bis(4-hydroxyphenyl) propionitrile reduced infected neurons at 48 h of treatment significantly compared to the control (p < 0.05 and p < 0.001, respectively). The Toxoplasma infection process was decreased by the effect of 17β-estradiol alone or combined with tamoxifen or progesterone in neurons in vitro. These results suggest the essential participation of progesterone and estradiol and their classical receptors in the regulation of T. gondii neuron infection.

Le Petit Machiavellian Prince: Effects of latent toxoplasmosis on political beliefs and values

Humans infected by Toxoplasma gondii express no specific symptoms but manifest higher incidence of many diseases, disorders and differences in personality and behaviour. The aim of this study was to compare the political beliefs and values of Toxoplasma-infected and Toxoplasma-free participants. We measured beliefs and values of 2315 responders via an online survey (477 Toxoplasma-infected) using the Political Beliefs and Values Inventory (PI34). This study showed Toxoplasma-infected and Toxoplasma-free participants of our cross-sectional study differed in three of four factors of PI34, scoring higher in Tribalism and lower in Cultural liberalism and Anti-Authoritarianism. We found sex differences in political beliefs associated with Toxoplasma infection. Infected women scored higher in tribalism and lower in cultural liberalism, compared with the Toxoplasma-free control group, while infected men scored higher in economic equity. These results fit with sexual differences in behaviour and attitude observed after toxoplasmosis infection. Controlling for the effect of worse physical health and mental health had little impact, suggesting that impaired health did not cause these changes. Rather than adaptation to prevalence of parasites, as suggested by parasite-stress theory, the differences might be side-effects of long-term mild inflammatory re-action, although definitive conclusions cannot be reached due to lack of relevant literature.

Co Expression of GMFβ, IL33, CCL2 and SDF1 Genes in the Acute Stage of Toxoplasmosis in Mice Model and Relation for Neuronal Impairment

Background: Toxoplasma gondii is an obligate intracellular parasite that migrates through macrophages or dendritic cells to neurons and nerve cells. Glia Maturation Factor (GMF) is a pre-inflammatory protein that is expressed in the central nervous system (CNS). GMFβ expression is related to IL33 and CCL2 and SDF1 in some neurodegenerative diseases. According to the importance of GMFβ in neurodegenerative diseases and its association with IL33, CCL2 and SDF1 genes, this study was designed to determine the level of expression of these genes in the brains of mice with acute toxoplasmosis. Methods: Tachyzoites of T. gondii RH strains were injected to 5 Swiss Albino mice. At the same time, healthy mice were inoculated with the Phosphate-buffered saline (PBS). Their brains were removed and kept at -70 oC in order to RNA extraction, cDNA syntheses and Real Time PCR performance. The level of gene expression was investigated with SYBR Green Quantitative Real-Time PCR. Results: GMFβ gene expression increased significantly (P=0.003) 3.26 fold in Toxoplasma infected mice in comparison to the control. GMFβ gene expression was associated with increased expression level of IL33, CCL2, and SDF1 genes. Conclusion: Considering the prominent role of GMFβ in CNS as well as the immune system, the elevation of GMFβ, IL33, CCL2 and SDF1 genes expression in the early stage of toxoplasmosis is associated with the occurrence of neuropathological alterations. Detection of these genes as an indication of brain damage in the early stages of Toxoplasma infection can prevent neurodegenerative disorders following acquired toxoplasmosis.

Tbet expression by Tregs is needed to protect against Th1-mediated immunopathology during Toxoplasma infection in mice

T. gondii infection has proven to be an ideal model to understand the delicate balance between protective immunity and immune-mediated pathology during infection. Lethal infection causes a collapse of Tregs mediated by loss of IL-2, and conversion of Tregs to IFNγ producing cells. Importantly, these Tregs highly express the Th1 transcription factor Tbet. To determine the role of Tbet in Tregs, we infected Tbx21f/f-Foxp3YFPCre and control Foxp3YFPCre mice with the type II strain of T. gondii, ME49. The majority of Tbx21f/f-Foxp3YFPCre mice succumb to a non-lethal acute infection. Notably, parasite burden is comparable between Tbx21f/f-Foxp3YFPCre and Foxp3YFPCre control mice. We found that Tbx21f/f-Foxp3YFPCre mice have significantly higher serum levels of proinflammatory cytokines IFNγ and TNFα, suggestive of a heightened immune response. To test if CD4+ T cells were driving immunopathology, we treated Tbx21f/f-Foxp3YFPCre mice with anti-CD4 depleting antibody and partially rescued these mice. Broad spectrum antibiotic treatment also improved survival, demonstrating a role for commensal flora in immunopathology in Tbx21f/f-Foxp3YFPCre mice. RNA-seq analysis reinforced that Tbet regulates several key cellular pathways, including chromosome segregation, cytokine receptor activity and cell cycle progression, that help to maintain fitness in Tregs during Th1 responses. Taken together, our data shows an important role for Tbet in Tregs in preventing lethal immunopathology during Toxoplasma gondii infection, further highlighting the protective role of Treg plasticity to self and microbiota.