Acute Toxoplasma Gondii Infection Research Articles

Functional analysis of antigen presentation by enteric glial cells during intestinal inflammation.

The Enteric Nervous System is composed of a vastly interconnected network of neurons and glial cells that coordinate to regulate homeostatic gut function including intestinal motility, nutrient sensing, and mucosal barrier immunity. Enteric Glial Cells (EGCs) are a heterogeneous cell population located throughout the gastrointestinal tract and have well described roles in regulating intestinal immune responses. Enteric Glial Cells have been suggested to act as nonconventional antigen presenting cells via the Major Histocompatibility Complex II (MHC II), though this has not been confirmed functionally. Here, we investigate the capability of EGCs to present antigen on MHC I and MHC II using in vitro antigen presentation assays performed with primary murine EGC cultures. We found that EGCs are capable of functional antigen presentation on MHC I, including antigen cross-presentation, but are not capable of functional antigen presentation on MHC II. We also determined EGC cell surface MHC I and MHC II expression levels by flow cytometry during intestinal inflammation during Dextran Sodium Sulfate-induced colitis or acute Toxoplasma gondii infection. We found that EGCs upregulate MHC I during acute T. gondii infection and induce low-level MHC II expression. These findings suggest that EGCs may be important in the regulation of CD8+ T cell responses via MHC I mediated antigen (cross) presentation but may not be relevant for MHC II-mediated antigen presentation.

Isatin-1,2,3-triazole derivatives: synthesis, molecular docking and evaluation against acute experimental toxoplasmosis

Toxoplasmosis remains a challenge for both public health and animal husbandry which created a constant demand to develop novel compounds using innovative methods. To join this relentless quest for an ideal chemotherapeutic agent, herein, we developed newly synthesized isatin-1,2,3-triazole derivatives. Three compounds (5a, 5b and 5c) were synthesized, characterized, and loaded on chitosan nanoparticles (CNPs) and then evaluated accordingly. Initially, a molecular docking study was carried out which revealed the effective interaction with the target enzymes; purine nucleoside phosphorylase (PNPase) and T. gondii calcium-dependent protein kinase-1 (TgCDPK1). This was further substantiated by in vivo evaluation of the three compounds (5a-c) and their nanoformulae (5a-CNPs, 5b-CNPs, and 5c-CNPs) against acute Toxoplasma gondii infection in murine model. It is worthy of note that all tested compounds and their nanoformulae produced a significant reduction of parasite burden in both peritoneal fluid and liver impression smear and profound ultrastructural alterations, detected by scanning electron microscopy, compared to the infected non-treated control. The nanoformula 5c-CNPs yielded the most outstanding results with the highest tachyzoite reduction percentage in both peritoneal fluid (98.1%) and liver impression smear (95.3%). Furthermore, the serum levels of liver enzymes (aspartate transaminase (AST) and alanine transaminase (ALT), and renal function tests (urea and creatinine) in mice were within normal limits which makes them more appealing candidates with proven safety. To the best of our knowledge, the present work is the first in silico and in vivo study proving the anti-Toxoplasma effect of isatin-1,2,3- triazoles which paves the way for further development of isatin and triazole-based leads for the treatment of toxoplasmosis.

Caspase-1 in Cx3cr1-expressing cells drives an IL-18-dependent T cell response that promotes parasite control during acute Toxoplasma gondii infection.

Inflammasome activation is a robust innate immune mechanism that promotes inflammatory responses through the release of alarmins and leaderless cytokines, including IL-1α, IL-1β, and IL-18. Various stimuli, including infectious agents and cellular stress, cause inflammasomes to assemble and activate caspase-1. Then, caspase-1 cleaves targets that lead to pore formation and leaderless cytokine activation and release. Toxoplasma gondii has been shown to promote inflammasome formation, but the cell types utilizing caspase-1 and the downstream effects on immunological outcomes during acute in vivo infection have not been explored. Here, using knockout mice, we examine the role of caspase-1 responses during acute T. gondii infection globally and in Cx3cr1-positive populations. We provide in vivo evidence that caspase-1 expression is critical for, IL-18 release, optimal interferon-γ (IFN-γ) production, monocyte and neutrophil recruitment to the site of infection, and parasite control. Specifically, we find that caspase-1 expression in Cx3cr1-positive cells drives IL-18 release, which potentiates CD4+ T cell IFN-γ production and parasite control. Notably, our Cx3cr1-Casp1 knockouts exhibited a selective T cell defect, mirroring the phenotype observed in Il18 knockouts. In further support of this finding, treatment of Cx3cr1-Casp1 knockout mice with recombinant IL-18 restored CD4+ T cell IFN-γ responses and parasite control. Additionally, we show that neutrophil recruitment is dependent on IL-1 receptor accessory protein (IL-1RAP) signaling but is dispensable for parasite control. Overall, these experiments highlight the multifaceted role of caspase-1 in multiple cell populations contributing to specific pathways that collectively contribute to caspase-1 dependent immunity to T. gondii.

The Intestinal Barrier Protective Effect of Indole Aldehyde Derivatives on Acute Toxoplasma gondii Infection.

Toxoplasmosis, a zoonotic infection caused by Toxoplasma gondii (T. gondii), poses a significant risk to human health and public safety. Despite the availability of clinical treatments, none effectively mitigate the intestinal barrier damage, which is the primary defense against T. gondii invasion. This study introduced aldehyde groups into the indole scaffold of a peptide-like structure to investigate the protective effects of these indole aldehyde derivatives on the intestinal barrier in mice with acute T. gondii infection. This approach leveraged the propensity of peptides and aldehyde groups to form hydrogen bonds. We synthesized a range of indole derivatives using the Vilsmeier-Haack reaction and evaluated their intestinal barrier protective effects both in vitro and in vivo. Our findings revealed that indole derivatives A1 (1-Formyl-1H-indole-3-acetonitrile), A3 (Indole-3-carboxaldehyde), A5 (2-Chloro-1H-indole-3-carboxaldehyde), A8 (1-Methyl-indole-3-carboxaldehyde), and A9 (1-Methyl-2-phenyl-1H-indole-3-carboxaldehyde) demonstrated a higher selectivity index compared to the positive control, spiramycin. These derivatives enhanced gastrointestinal motility, increased glutathione (GSH) levels in the small intestine, and reduced malondialdehyde (MDA) and nitric oxide (NO) levels in the small intestine tissue and diamine oxidase (DAO) and NO levels in the serum of infected mice. Notably, A3 exhibited comparable anti-T. gondii tachyzoites activity in the peritoneal cavity. Molecular docking studies indicated that the aldehyde group on the indole scaffold not only formed a hydrogen bond with NTPase-II but also interacted with TgCDPK1 through hydrogen bonding. Among the derivatives, A3 showed promising intestinal barrier protective effects in mice with acute T. gondii infection. This research suggests that indole derivatives could serve as a potential therapeutic strategy for intestinal diseases induced by T. gondii, offering a novel direction for treating intestinal barrier damage and providing valuable insights for the chemical modification of drugs targeting T. gondii. Furthermore, it contributes to the advancement of therapeutic approaches for toxoplasmosis.

α2,6 sialylation distinguishes a novel active state in CD4+ and CD8+ cells during acute Toxoplasma gondii infection.

Toxoplasmosis is a worldwide parasitosis that is usually asymptomatic; cell-mediated immunity, particularly T cells, is a crucial mediator of the immune response against this parasite. Membrane protein expression has been studied for a long time in T lymphocytes, providing vital information to determine functional checkpoints. However, less is known about the role of post-translational modifications in T cell function. Glycosylation plays essential roles during maturation and function; particularly, sialic acid modulation is determinant for accurate T cell regulation of processes like adhesion, cell-cell communication, and apoptosis induction. Despite its importance, the role of T cell sialylation during infection remains unclear. Herein, we aimed to evaluate whether different membrane sialylation motifs are modified in T cells during acute Toxoplasma gondii infection using different lectins. To this end, BALB/c Foxp3EGFP mice were infected with T. gondii, and on days 3, 7, and 10 post-infection, splenocytes were obtained to analyze conventional (Foxp3-) CD4+ and CD8+ populations by flow cytometry. Among the different lectins used for analysis, only Sambucus nigra lectin, which detects sialic acid α2,6 linkages, revealed two distinctive populations (SNBright and SN-/Dim) after infection. Further characterization of CD4+ and CD8+ SN-/Dim lymphocytes showed that these are highly activated cells, with a TEf/EM or TCM phenotype that produce high IFN-γ levels, a previously undescribed cell state. This work demonstrates that glycan membrane analysis in T cells reveals previously overlooked functional states by evaluating only protein expression.

The opposing effect of acute and chronic Toxoplasma gondii infection on tumor development

BackgroundThe interplay between Toxoplasma gondii infection and tumor development is intriguing and not yet fully understood. Some studies showed that T. gondii reversed tumor immune suppression, while some reported the opposite, stating that T. gondii infection promoted tumor growth.MethodsWe created three mouse models to investigate the interplay between T. gondii and tumor. Model I aimed to study the effect of tumor growth on T. gondii infection by measuring cyst number and size. Models II and III were used to investigate the effect of different stages of T. gondii infection on tumor development via flow cytometry and bioluminescent imaging. Mouse strains (Kunming, BALB/c, and C57BL/6J) with varying susceptibilities to tumors were used in the study.ResultsThe size and number of brain cysts in the tumor-infected group were significantly higher, indicating that tumor presence promotes T. gondii growth in the brain. Acute T. gondii infection, before or after tumor cell introduction, decreased tumor growth manifested by reduced bioluminescent signal and tumor size and weight. In the tumor microenvironment, CD4+ and CD8+ T cell number, including their subpopulations (cytotoxic CD8+ T cells and Th1 cells) had a time-dependent increase in the group with acute T. gondii infection compared with the group without infection. However, in the peripheral blood, the increase of T cells, including cytotoxic CD8+ T cells and Th1 cells, persisted 25 days after Lewis lung carcinoma (LLC) cell injection in the group with acute T. gondii. Chronic T. gondii infection enhanced tumor growth as reflected by increase in tumor size and weight. The LLC group with chronic T. gondii infection exhibited decreased percentages of cytotoxic CD8+ T cells and Th1 cells 25 days post-LLC injection as compared with the LLC group without T. gondii infection. At week 4 post-LLC injection, chronic T. gondii infection increased tumor formation rate [odds ratio (OR) 1.71] in both KM and BALB/c mice.ConclusionsOur research elucidates the dynamics between T. gondii infection and tumorigenesis. Tumor-induced immune suppression promoted T. gondii replication in the brain. Acute and chronic T. gondii infection had opposing effects on tumor development.Graphical

Protective efficacy of Toxoplasma gondii GRA12 or GRA7 recombinant proteins encapsulated in PLGA nanoparticles against acute Toxoplasma gondii infection in mice.

Toxoplasma gondii is an apicomplexan parasite that affects the health of humans and livestock, and an effective vaccine is urgently required. Nanoparticles can modulate and improve cellular and humoral immune responses. In the current study, poly (D, L-lactic-co-glycolic acid) (PLGA) nanoparticles were used as a delivery system for the T. gondii dense granule antigens GRA12 and GRA7. BALB/c mice were injected with the vaccines and protective efficacy was evaluated. Mice immunized with PLGA+GRA12 exhibited significantly higher IgG, and a noticeable predominance of IgG2a over IgG1 was also observed. There was a 1.5-fold higher level of lymphocyte proliferation in PLGA+GRA12-injected mice compared to Alum+GRA12-immunized mice. Higher levels of IFN-g and IL-10 and a lower level of IL-4 were detected, indicating that Th1 and Th2 immune responses were induced but the predominant response was Th1. There were no significant differences between Alum+GRA7-immunized and PLGA+GRA7-immunized groups. Immunization with these four vaccines resulted in significantly reduced parasite loads, but they were lowest in PLGA+GRA12-immunized mice. The survival times of mice immunized with PLGA+GRA12 were also significantly longer than those of mice in the other vaccinated groups. The current study indicated that T. gondii GRA12 recombinant protein encapsulated in PLGA nanoparticles is a promising vaccine against acute toxoplasmosis, but PLGA is almost useless for enhancing the immune response induced by T. gondii GRA7 recombinant protein.

Efficacy of clofazimine against acute and chronic Toxoplasma gondii infection in mice

Toxoplasmosis is a zoonotic protozoal disease affecting approximately one-third of the world's population. The lack of current treatment options necessitates the development of drugs with good tolerance and effectiveness on the active and cystic stages of the parasite. The present study was established to investigate, for the first time, the potential potency of clofazimine (CFZ) against acute and chronic experimental toxoplasmosis. For this purpose, the type II T. gondii (Me49 strain) was used for induction acute (20 cysts in each mouse) and chronic (10 cysts in each mouse) experimental toxoplasmosis. The mice were treated with 20 mg/kg of CFZ intraperitoneally and orally. The histopathological changes, brain cyst count, total Antioxidant Capacity (TAC), malondialdehyde (MDA) assay, and the level of INF-γ were also evaluated. In the acute toxoplasmosis, both IP and oral administration of CFZ induced a significant reduction in brain parasite burden by 90.2 and 89%, respectively, and increased the survival rate to 100% compared with 60% in untreated controls. In the chronic infection, cyst burden decreased at 85.71 and 76.18% in CFZ-treated subgroups in comparison to infected untreated controls. In addition, 87.5% and 100% of CFZ-treated subgroups survived versus untreated control 62.5%. Moreover, CFZ significantly increased INF-γ levels in acute and chronic toxoplasmosis. Tissue inflammatory lesions were considerably reduced in the CFZ-treated chronic subgroups. CFZ treatment significantly reduced MDA levels and elevated TAC in both acute and chronic infections. In conclusion, CFZ showed a promising finding regarding the ability to reduce cyst burden in acute and chronic infection. Further studies are needed to investigate the therapeutic role of CFZ on toxoplasmosis using the long-term treatment and more advanced approaches. In addition, clofazimine may need to be accompanied by another drug to augment its effect and prevent the regrowth of parasites.

Diagnostic accuracy of adjusted low IgG avidity index to predict acute Toxoplasma gondii infection in the first trimester of pregnancy.

Congenital toxoplasmosis is reportable disease in Europe. To prevent it antibody serological tests were introduced in several European countries as a part of screening programmes. Immunoglobulin G (IgG) avidity index testing is one of these tests for diagnosing acute infection with Toxoplasma gondii (Nicolle et Manceaux, 1908) in pregnant women. However, a low or moderate IgG avidity index can give inconclusive results for predicting woman's status. From June 2012 until the end of 2014, 17,990 women were included in the national screening program to prevent congenital toxoplasmosis. One hundred and twenty-six women were consecutively included in the study because they had low or moderate IgG avidity. Every woman with possible acute toxoplasmosis was followed up every month till delivery. Fifty-eight of 126 (46%) women got infected in months before current pregnancy, 39 women (31%) were infected early in pregnancy. Twenty-nine pregnant women of 126 (23%) got infected in the second/third trimester of pregnancy. New cut off for IgG avidity index was 0.11. With this cut off, we were able to exclude T. gondii acute infection in the first trimester with very good diagnostic accuracy (area under the curve (AUC) = 0.95, 95% confidence Interval (CI) 0.91-0.99, sensitivity 0.95, specificity 0.86). If an IgG avidity index above 0.11 is measured in a woman's serum and she is in the first trimester of pregnancy, then a odds ratio (OR) for acute infection with T. gondii is below 1 (OR 0.11, 95% CI 0.05-0.25, P < 0.0001). If we measure IgG avidity index that is ≥ 0.11 in the first trimester of pregnancy, we can exclude infection with T. gondii with good diagnostic accuracy in our cohort of women. With a new cut off we could reduce number of invasive procedures such as amniocentesis and put less pregnant women in distress.

Effects of Notch signaling pathway inhibition by dibenzazepine in acute experimental toxoplasmosis

Notch signaling pathway plays a crucial role in cellular fate across species, being important for the differentiation and development of several cell types. The aim of this study was to evaluate the effect of Notch inhibition pathway by dibenzazepine (DBZ) in histological and inflammatory alterations and, tissue parasitism in acute Toxoplasma gondii infection. For this, C57BL/6 mice were treated with DBZ before infection with T. gondii, and the small intestine, lungs and liver were analyzed. The genes related to Notch signaling pathway were assayed through qPCR in the organs, and cytokine measurement was performed in serum samples. In the small intestine, T. gondii infection impaired the Hes1 and Math1 mRNA expressions, increased the inflammation and decreased goblet and Paneth cell numbers. The DBZ-treatment was able to partially preserve these cells, however, the parasitism and inflammation were not altered. In parallel, the high IL-2, IL-6, TNF and, IFN-γ levels induced by infection were not changed with the DBZ treatment, with the IFN-γ levels even higher. In contrast, in the liver and lungs, the DBZ-treatment diminished parasitism and inflammation. Our results highlight that Notch pathway inhibition in T.gondii infection results in different parasitological and inflammatory outcomes depending on the organ analyzed.

Global proteomic profiling of multiple organs of cats (Felis catus) and proteome-transcriptome correlation during acute Toxoplasma gondii infection

BackgroundToxoplasma gondii is a protozoan parasite which can infect almost all warm-blooded animals and humans. Understanding the differential expression of proteins and transcripts associated with T. gondii infection in its definitive host (cat) may improve our knowledge of how the parasite manipulates the molecular microenvironment of its definitive host. The aim of this study was to explore the global proteomic alterations in the major organs of cats during acute T. gondii infection.MethodsiTRAQ-based quantitative proteomic profiling was performed on six organs (brain, liver, lung, spleen, heart and small intestine) of cats on day 7 post-infection by cysts of T. gondii PRU strain (Genotype II). Mascot software was used to conduct the student’s t-test. Proteins with P values < 0.05 and fold change > 1.2 or < 0.83 were considered as differentially expressed proteins (DEPs).ResultsA total of 32,657 proteins were identified in the six organs, including 2556 DEPs; of which 1325 were up-regulated and 1231 were down-regulated. The brain, liver, lung, spleen, heart and small intestine exhibited 125 DEPs, 463 DEPs, 255 DEPs, 283 DEPs, 855 DEPs and 575 DEPs, respectively. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of all proteins and DEPs in all organs showed that many proteins were enriched in binding, cell part, cell growth and death, signal transduction, translation, sorting and degradation, extracellular matrix remodeling, tryptophan catabolism, and immune system. Correlations between differentially expressed proteins and transcripts were detected in the liver (n = 19), small intestine (n = 17), heart (n = 9), lung (n = 9) and spleen (n = 3).ConclusionsThe present study identified 2556 DEPs in six cat tissues on day 7 after infection by T. gondii PRU strain, and functional enrichment analyses showed that these DEPs were associated with various cellular and metabolic processes. These findings provide a solid base for further in-depth investigation of the complex proteotranscriptomic reprogramming that mediates the dynamic interplays between T. gondii and the different feline tissues.

Microfluidic model for in vitro acute Toxoplasma gondii infection and transendothelial migration

The protozoan parasite Toxoplasma gondii (T. gondii) causes one of the most common human zoonotic diseases and infects approximately one-third of the global population. T. gondii infects nearly every cell type and causes severe symptoms in susceptible populations. In previous laboratory animal studies, T. gondii movement and transmission were not analyzed in real time. In a three-dimensional (3D) microfluidic assay, we successfully supported the complex lytic cycle of T. gondii in situ by generating a stable microvasculature. The physiology of the T. gondii-infected microvasculature was monitored in order to investigate the growth, paracellular and transcellular migration, and transmission of T. gondii, as well as the efficacy of T. gondii drugs.

The correlation of TNF alpha levels with acute phase proteins in acute Toxoplasma gondii infection in mice

Acute Toxoplasma gondii infections can influence the liver as well as other organs. Many cytokines and proteins play a role in the acute response against infection. Tumor necrosis factor alpha (TNF alpha) is a cytokine that plays a key function in stimulating hepatocytes to produce acute phase proteins. In this study, we investigated TNF alpha levels associated with the levels of macroglobulin, haptoglobin, hemopexin, C-reactive protein (CRP), albumin, serum amyloid alpha protein (SAA), and clusterin, which are acute phase proteins, in serum of mice with T. gondii infection. In the experiment, a total of 24 mice were used; 6 mice constituted the control group and 18 mice were infected with the RH strain. On the 2nd, 4th, and 6th days following the infection, 6 animals were euthanized, and their serums were collected. Compared to the control group, we observed a statistically significant decrease in albumin concentration in the group with T. gondii infection on the 6th day. Also, this group displayed a statistically significant, gradual increase in clusterin levels on the 2nd and 6th days, C-reactive protein levels on the 4th day, haptoglobin levels on the 2nd and 4th days, hemopexin levels on the 2nd day, serum amyloid A levels on the 2nd, 4th, and 6th days, and TNF-α levels on the 2nd, 4th, and 6th days (p < 0.05). TNF-α was strongly positively correlated with CRP, SAA, and clusterin, moderately positively correlated with hemopexin, and strongly negatively correlated with albumin. The increase in CRP, SAA, clusterin, and hemopexin levels on the 2nd day after infection, in parallel with the increase in TNF-α levels, indicates that these proteins can be considered as major acute phase proteins in acute T. gondii infection in mice. The data obtained here may be helpful for the diagnosis of T. gondii infection and for monitoring treatments.

Cluster analysis of splenocyte microRNAs in the pig reveals key signal regulators of immunomodulation in the host during acute and chronic Toxoplasma gondii infection

BackgroundToxoplasma gondii is an obligate intracellular protozoan parasite that can cause a geographically widespread zoonosis. Our previous splenocyte microRNA profile analyses of pig infected with T. gondii revealed that the coordination of a large number of miRNAs regulates the host immune response during infection. However, the functions of other miRNAs involved in the immune regulation during T. gondii infection are not yet known.MethodsClustering analysis was performed by K-means, self-organizing map (SOM), and hierarchical clustering to obtain miRNA groups with the similar expression patterns. Then, the target genes of the miRNA group in each subcluster were further analyzed for functional enrichment by Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Reactome pathway to recognize the key signaling molecules and the regulatory signatures of the innate and adaptive immune responses of the host during T. gondii infection.ResultsA total of 252 miRNAs were successfully divided into 22 subclusters by K-means clustering (designated as K1–K22), 29 subclusters by SOM clustering (designated as SOM1–SOM29), and six subclusters by hierarchical clustering (designated as H1–H6) based on their dynamic expression levels in the different infection stages. A total of 634, 660, and 477 GO terms, 15, 26, and 14 KEGG pathways, and 16, 15, and 7 Reactome pathways were significantly enriched by K-means, SOM, and hierarchical clustering, respectively. Of note, up to 22 miRNAs mainly showing downregulated expression at 50 days post-infection (dpi) were grouped into one subcluster (namely subcluster H3-K17-SOM1) through the three algorithms. Functional analysis revealed that a large group of immunomodulatory signaling molecules were controlled by the different miRNA groups to regulate multiple immune processes, for instance, IL-1-mediated cellular response and Th1/Th2 cell differentiation partly depending on Notch signaling transduction for subclusters K1 and K2, innate immune response involved in neutrophil degranulation and TLR4 cascade signaling for subcluster K15, B cell activation for subclusters SOM17, SOM1, and SOM25, leukocyte migration, and chemokine activity for subcluster SOM9, cytokine–cytokine receptor interaction for subcluster H2, and interleukin production, chemotaxis of immune cells, chemokine signaling pathway, and C-type lectin receptor signaling pathway for subcluster H3-K17-SOM1.ConclusionsCluster analysis of splenocyte microRNAs in the pig revealed key regulatory properties of subcluster miRNA molecules and important features in the immune regulation induced by acute and chronic T. gondii infection. These results contribute new insight into the identification of physiological immune responses and maintenance of tolerance in pig spleen tissues during T. gondii infection.Graphical

Defining the Location of T-bet-expressing Myeloid Cells During Acute Intestinal Toxoplasma gondii Infection

Background/Objective:&#x0D; The protozoan parasite Toxoplasma gondii is the second leading cause of foodborne pathogen-related deaths in the United States. The transcription factor T-bet is indispensable for host immunity against T. gondii. The absence of T-bet results in rapid susceptibility during parasite infection. T-bet has been considered essential for T-cell-derived IFN-g during T. gondii infection; yet, recent research has shown that T-bet is not required for lymphocyte-derived IFN-g responses. Our preliminary research shows that T-bet-deficient mice succumb to parasite infection significantly quicker than mice lacking lymphocytes. This has led to our hypothesis that T-bet-dependent myeloid cells are critical for host resistance during acute intestinal T. gondii infection. The objective of this project was to define the location of the T-bet-expressing myeloid cells in the medial small intestines (MSI) of naïve and infected mice during acute mucosal parasite infection.&#x0D; Methods: &#x0D; We used immunofluorescence microscopy to determine the location of T-bet-expressing myeloid cells in the MSI of naïve and T. gondii infected mice. Mice were orally infected with 40 cysts of the ME49 strain of T. gondii. On days 0 and 5, one-inch MSI segments were harvested, fixed with 4% paraformaldehyde for at least one hour, and then frozen in OCT compound. Tissues were then cut into 8mm sections and placed onto slides for staining. Sections were stained for nuclei, CD11c, T-bet, and T. gondii.&#x0D; Results: &#x0D; Our results revealed T-bet-expressing CD11c+ cells in both the MSI and spleen on days 0 and 5 of T. gondii infection.&#x0D; Summary:&#x0D; These data indicate that T-bet-expressing myeloid cells are present in the MSI during T. gondii infection. Defining the position of these cells will allow us to determine T-bet’s role in mediating myeloid cell-dependent T. gondii clearance. Due to the limited treatment options for patients suffering from toxoplasmosis it is critical to define new mechanisms for eliminating T. gondii.

Seroprevalence and Associated Risk Factors of Toxoplasma gondii Among Pregnant Women in Southwest Iran

Background: Acute Toxoplasma gondii infection during pregnancy period can cause congenital toxoplasmosis. The aim of this study was to assess the seroprevalence rate of immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies against T. gondii infection during pregnancy and the associated risk factors in southwest Iran. Methods: This study was performed on 88 pregnant women from October to December 2019 in Khorramshahr County, Khuzestan province, Iran. Anti-T. gondii IgG and IgM antibodies were tested through enzyme-linked immunosorbent assay (ELISA) method. Results: Following serological assays, 38.63% (34/88) and 2.27% (2/88) serum samples were positive for IgG and IgM antibodies, respectively. Also, a statistically significant association was observed between IgG seroprevalence and drinking of unpurified water (P = 0.015). Conclusions: The serological evidence revealed that pregnant women of southwest Iran had moderate exposure to T. gondii parasite. Since the risk of acquiring acute toxoplasmosis in pregnant women is clinically important, we highly recommend regular screening tests for T. gondii infection during pregnancy period.

Detection of Acute and Chronic Toxoplasma gondii Infection among Women with History of Abortion in the Southwest of Iran.

Background Toxoplasma gondii (T. gondii) is one of the most common intracellular protozoan parasites, which can infect humans and a wide range of mammals and birds. The current study is aimed at investigating the occurrence of T. gondii infection in women with a history of abortion in Khuzestan, Iran. Materials and Methods A total of 480 women with an abortion history, as well as 200 pregnant women with a normal delivery, were examined in this study. The blood, placenta, and umbilical cord blood samples were assessed by the enzyme-linked immunosorbent assay (ELISA) and nested-polymerase chain reaction (PCR) assay. Results Based on the results of ELISA assay, the prevalence of toxoplasmosis was 30.83% in women with a history of abortion (25.62% with T. gondii IgG and 5.20% with T. gondii IgM). According to the IgG avidity test, 60.16% of IgG-positive samples showed high avidity, while 27.64% showed low avidity. On the other hand, the prevalence of toxoplasmosis in women with a normal delivery was 23% (21.5% with T. gondii IgG and 1.5% with T. gondii IgM). According to the IgG avidity test, 81.39% of these women showed high avidity, while only 4.65% showed low avidity. Based on the nested-PCR method, T. gondii DNA was detected in 14.18% of blood samples, 4.69% of placental samples, and 1.34% of umbilical cord samples, collected from 148 seropositive women with a history of abortion. Besides, using this method, the parasite DNA was identified in 4.34% of blood samples, collected from 46 seropositive women with a normal delivery, but not in any of the umbilical cord or placenta samples. Conclusion The present results showed that T. gondii infection contributes to abortion in Khuzestan Province, Iran. Therefore, it is essential to investigate toxoplasmosis in pregnant women, especially in those who are seronegative, using molecular and serological methods and inform them about their disease and the associated risks.

Analogs of Marinopyrrole A Show Enhancement to Observed In Vitro Potency against Acute Toxoplasma gondii Infection.

The apicomplexan parasite Toxoplasma gondii is the causative agent of toxoplasmosis, a globally distributed infection with severe clinical consequences for immunocompromised individuals and developing fetuses. There are few available treatments, and these are associated with potentially severe adverse effects. Marinopyrrole A, a compound discovered in a marine Streptomyces species, has previously been found to exhibit potent antimicrobial activity, prompting our interest in exploring efficacy against Toxoplasma gondii. We found that marinopyrrole A was a highly potent anti-Toxoplasma molecule, with an in vitro 50% maximal inhibitory concentration (IC50) of 0.31 μM, corresponding to a higher potency than that of the current standard of care (pyrimethamine); however, addition of 20% serum led to abrogation of potency, and toxicity to human cell lines was observed. Yet, application of marinopyrrole A to an in vivo lethal acute infection model facilitated significantly enhanced survival at doses of 5, 10, and 20 mg/kg. We then tested a series of marinopyrrole A analogs (RL002, RL003, and RL125) and demonstrated significantly increased potency in vitro, with IC50 values ranging from 0.09 to 0.17 μM (3.6- to 6.8-fold increase relative to pyrimethamine). No detectable cytotoxicity was observed up to 50 μM in human foreskin fibroblasts, with cytotoxicity in HepG2 cells ranging from ∼28 to 50 μM, corresponding to >200-fold selectivity for parasites over host cells. All analogs additionally showed reduced sensitivity to serum. Further, RL003 potently inhibited in vitro-generated bradyzoites at 0.245 μM. Taken together, these data support further development of marinopyrrole A analogs as promising anti-Toxoplasma molecules to further combat this prevalent infection.

Early Immune Initiation by Porcine Cells following Toxoplasma gondii Infection versus TLR Ligation.

Containment of acute Toxoplasma gondii infection is dependent on an efficient interferon gamma response. However, the earliest steps of immune response initiation immediately following exposure to the parasite have not been previously characterized in pigs. Murine and human myeloid cells produce large quantities of interleukin (IL)-12 during early T. gondii infection. We therefore examined IL-12 expression by porcine peripheral blood monocytes and dendritic cell (DC) subsets following toll-like receptor (TLR) ligation and controlled T. gondii tachyzoite infection. We detected IL-12p40 expression by porcine plasmacytoid DC, but not conventional or monocyte-derived DC following TLR ligation. Unexpectedly, we also observed considerable IL-12p40 production by porcine CD3– NKp46+ cells—a classical natural killer cell phenotype—following TLR ligation. However, in response to T. gondii exposure, no IL-12 production was observed by either DC or CD3– NKp46+ cells. Despite this, IL-18 production by DC-enriched peripheral blood mononuclear cells was detected following live T. gondii tachyzoite exposure. Only combined stimulation of porcine peripheral blood mononuclear cells with recombinant IL-12p70 and IL-18 induced innate interferon gamma production by natural killer cells, while T cells and myeloid cells did not respond. Therefore, porcine CD3– NKp46+ cells serve as important IL-12 producers following TLR ligation, while IL-18 likely plays a prominent role in early immune response initiation in the pig following T. gondii infection.

Seroprevalence and associated risk factors of Toxoplasma gondii infection among pregnant mother in Makassar, Indonesia.

The protozoan parasite, Toxoplasma gondii is estimated to infect one-third of the world's population. Infection in pregnant women can cause severe conditions for their babies. Until now, there is no data regarding Toxoplasma infection from Makassar pregnant mothers. This study aims to obtain information on Toxoplasma specific antibodies and to measure the risk factor associate with parasite infection. This cross-sectional study conducted in 9 of 47 primary health centres (Puskesmas) in Makassar. Blood samples and questionnaires were collected from 184 pregnant women aged 15-42 years old from September to October 2020. ELISA technique was used to examine the IgG and IgM antibodies. Univariable and multivariable analyses were carried out to measure factors that independently associate with Toxoplasma antibody positivity. Our result showed the range of Toxoplasma IgM and IgG are 0.06-1.01 and 0.09-3.01, respectively. While no one of our participants has an acute Toxoplasma gondii infection (IgM positive), we found 32,6% pregnant mothers are exposed to parasite (positive IgG). Contact with cats [OR(95%CI): 10.45(3.77-28.99)], consume chicken satay [OR(95%CI): 9.72(3.71-25.48)] and consume un-boiled water/ filtered water [OR(95%CI): 5.98(1.77-20.23)] are independently associate with positive Toxoplasma IgG antibody. Based on the result, we conclude that pregnant women in Makassar are exposed to T. gondii and the oocyst and tissue cyst of parasite contaminates food and water in Makassar.