An investigation into Toxoplasma gondii at the human-livestock-wildlife interface, South Africa.

Interface areas shared by humans, domestic and wild animals may serve as high transmission contexts for Toxoplasma gondii. However, knowledge about the epidemiology of T. gondii in such areas is currently limited. The present study assessed the seroprevalence of T. gondii in different hosts from Mpumalanga, South Africa. Furthermore, we investigated the local knowledge and related practices about T. gondii by conducting a questionnaire study in the community. Blood samples were obtained and analysed for T. gondii antibodies using a commercial multispecies latex agglutination kit. The seroprevalence detected in humans (n = 160; patients showing signs of acute febrile illness), cats (n = 9), chickens (n = 336) and goats (n = 358) was 8.8%, 0.0%, 4.2% and 11.2%, respectively. Seroprevalence in impalas (n = 97), kudus (n = 55), wild dogs (n = 54), wildebeests (n = 43), warthogs (n = 97) and zebras (n = 68) was calculated at 5.2%, 7.3%, 100.0%, 20.9%, 13.4% and 9.1%, respectively. The questionnaire revealed that 63.0% of household owners were subsistence farmers, and 35.9% were pet owners. A high level of female participation was found (75.3%) when compared to male participation (24.7%). The results show a low circulation of T. gondii in the domestic cycle and suggest the presence of possible bridges between the wildlife cycle and the surrounding domestic cycle.Contribution: The study contributes to identifying transmission patterns and risk factors of T. gondii within human and animal populations. This topic fits within the scope of the journal presenting original research in veterinary science, with the focus on wild and domestic populations on the African continent on a topic of universal importance.

Observing astrocyte polarization in brains from mouse chronically infected with Toxoplasma gondii.

Toxoplasma gondii (T. gondii) is a protozoan parasite that infects approximately one-third of the global human population, often leading to chronic infection. While acute T. gondii infection can cause neural damage in the central nervous system and result in toxoplasmic encephalitis, the consequences of T. gondii chronic infection (TCI) are generally asymptomatic. However, emerging evidence suggests that TCI may be linked to behavioral changes or mental disorders in hosts. Astrocyte polarization, particularly the A1 subtype associated with neuronal apoptosis, has been identified in various neurodegenerative diseases. Nevertheless, the role of astrocyte polarization in TCI still needs to be better understood. This study aimed to establish a mouse model of chronic TCI and examine the transcription and expression levels of glial fibrillary acidic protein (GFAP), C3, C1q, IL-1α, and TNF-α in the brain tissues of the mice. Quantitative real-time PCR (qRT-PCR), enzyme-linked immunosorbent assay, and Western blotting were employed to assess these levels. Additionally, the expression level of the A1 astrocyte-specific marker C3 was evaluated using indirect fluorescent assay (IFA). In mice with TCI, the transcriptional and expression levels of the inflammatory factors C1q, IL-1α, and TNF-α followed an up-down-up pattern, although they remained elevated compared to the control group. These findings suggest a potential association between astrocyte polarization towards the A1 subtype and synchronized changes in these three inflammatory mediators. Furthermore, immunofluorescence assay (IFA) revealed a significant increase in the A1 astrocytes (GFAP+C3+) proportion in TCI mice. This study provides evidence that TCI can induce astrocyte polarization, a biological process that may be influenced by changes in the levels of three inflammatory factors: C1q, IL-1α, and TNF-α. Additionally, the release of neurotoxic substances by A1 astrocytes may be associated with the development of TCI.

Improved ELISPOT protocol for monitoring Th1/Th17 T-cell response following T.gondii infection.

In the monitoring of human Toxoplasma gondii infection, it is crucial to confirm the development of a specific Th1/Th17 immune response memory. The use of a simple, specific, and sensitive assay to follow the T-cell activation is thus required. Current protocols are not always specific as stimulation with peptides is Human Leukocyte Antigen (HLA)-dependent, while stimulation with total-lysis antigens tends to stimulate seronegative donors resulting to false positives. Here, an improved ELISPOT protocol is reported, using peripheral blood mononuclear cells (PBMC) of T.gondii-infected donors, incubated with the inactivated parasite. The results showed that, contrary to standard protocols, a pre-incubation step at high cell density in presence of the inactivated parasite allowed a specific Th1/Th17 response with the secretion of IFN-γ, IL-2, IL-12 and IL-17 cytokines. This protocol allows to evaluate precisely the immune response after a T.gondii infection.

Seroprevalence of Toxoplasma gondii, risk factors and knowledge about toxoplasmosis in undergraduate students from Lima, Peru.

Toxoplasma gondii is the causative agent of toxoplasmosis, a zoonotic disease of worldwide distribution. The aim of this study was to assess the seroprevalence of T. gondii in undergraduate students from Lima, Peru, and to identify the risk factors linked to the infection. For this, serum samples of 100 undergraduate students of the Faculty of Biological Sciences were tested for T. gondii antibodies with a commercially available ELISA. The seroprevalence of T. gondii in these subjects was 7 %. Only the age of students showed a statistical association with T. gondii seropositivity. The level of awareness regarding toxoplasmosis was also investigated. In the sample, 71 % of the students are aware of toxoplasmosis and 64 % that a parasite is the cause of the infection. Most know it is transmitted through undercooked meat (57 %), but are unaware of contaminated vegetables (40 %), organ transplants (17 %), blood transfusions (32 %), and soil contact (39 %). In the epidemiological context it will be valuable to verify toxoplasmosis awareness in other population groups and other regions in Peru.

Far-East Asian Toxoplasma isolates share ancestry with North and South/Central American recombinant lineages.

Toxoplasma gondii is a global protozoan pathogen. Clonal lineages predominate in Europe, North America, Africa, and China, whereas highly recombinant parasites are endemic in South/Central America. Far East Asian T. gondii isolates are not included in current global population genetic structure analyses at WGS resolution. Here we report a genome-wide population study that compared eight Japanese and two Chinese isolates against representative worldwide T. gondii genomes using POPSICLE, a novel population structure analyzing software. Also included were 7 genomes resurrected from non-viable isolates by target enrichment sequencing. Visualization of the genome structure by POPSICLE shows a mixture of Chinese haplogroup (HG) 13 haploblocks introgressed within the genomes of Japanese HG2 and North American HG12. Furthermore, two ancestral lineages were identified in the Japanese strains; one lineage shares a common ancestor with HG11 found in both Japanese strains and North American HG12. The other ancestral lineage, found in T. gondii isolates from a small island in Japan, is admixed with genetically diversified South/Central American strains. Taken together, this study suggests multiple ancestral links between Far East Asian and American T. gondii strains and provides insight into the transmission history of this cosmopolitan organism.

The role of microRNA-142a in Toxoplasma gondii infection-induced downregulation of Foxp3: implications for adverse pregnancy outcomes.

BACKGROUND: Toxoplasma gondii (T. gondii) is capable of infecting nearly all warm-blooded animals and approximately 30% of the global population. Though most infections are subclinical in immunocompetent individuals, congenital contraction can lead to severe consequences such as spontaneous abortion, stillbirth, and a range of cranio-cerebral and/or ocular abnormalities. Previous studies reported that T. gondii-infected pregnancy mice unveiled a deficit in both the amount and suppressive functions of regulatory T (Treg) cells, accompanied with reduced levels of forkhead box p3 (Foxp3). Recently, accumulative studies have demonstrated that microRNAs (miRNAs) are, to some extent, relevant to T. gondii infection. However, the link between alterations in miRNAs and downregulation of Foxp3 triggered by T. gondii has been only sporadically studied. METHODS: Quantitative reverse transcription polymerase chain reaction (RT-qPCR), protein blotting and immunofluorescence were employed to evaluate the impact of T. gondii infection and antigens on miRNA transcription and Foxp3 expression. Dual-luciferase reporter gene assays were performed to examine the fluorescence activity in EL4 cells, which were transfected with recombinant plasmids containing full-length/truncated/mutant microRNA-142a-3p (miR-142a) promoter sequence or wild type/mutant of Foxp3 3' untranslated region (3' UTR). RESULTS: We found a pronounced increase in miR-142a transcription, concurrent with a decrease in Foxp3 expression in T. gondii-infected mouse placental tissue. Similarly, comparable findings have been experimentally confirmed through the treatment of EL4 cells with T. gondii antigens (TgAg) in vitro. Simultaneously, miR-142a mimics attenuated Foxp3 expression, whereas its inhibitors markedly augmented Foxp3 expression. miR-142a promoter activity was elevated upon the stimulation of T. gondii antigens, which mitigated co-transfection of mutant miR-142a promoter lacking P53 target sites. miR-142a mimics deceased the fluorescence activity of Foxp3 3' untranslated region (3' UTR), but it did not affect the fluorescence activity upon the co-transfection of mutant Foxp3 3' UTR lacking miR-142a target site. CONCLUSION: In both in vivo and in vitro studies, a negative correlation was discovered between Foxp3 expression and miR-142a transcription. TgAg enhanced miR-142a promoter activity to facilitate miR-142a transcription through a P53-dependent mechanism. Furthermore, miR-142a directly targeted Foxp3 3' UTR, resulting in the downregulation of Foxp3 expression. Therefore, harnessing miR-142a may be a possible therapeutic approach for adverse pregnancy caused by immune imbalances, particularly those induced by T. gondii infection.

Fam96a is essential for the host control of Toxoplasma gondii infection by fine-tuning macrophage polarization via an iron-dependent mechanism.

BACKGROUND: Toxoplasmosis affects a quarter of the world's population. Toxoplasma gondii (T.gondii) is an intracellular parasitic protozoa. Macrophages are necessary for proliferation and spread of T.gondii by regulating immunity and metabolism. Family with sequence similarity 96A (Fam96a; formally named Ciao2a) is an evolutionarily conserved protein that is highly expressed in macrophages, but whether it play a role in control of T. gondii infection is unknown. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we utilized myeloid cell-specific knockout mice to test its role in anti-T. gondii immunity. The results showed that myeloid cell-specific deletion of Fam96a led to exacerbate both acute and chronic toxoplasmosis after exposure to T. gondii. This was related to a defectively reprogrammed polarization in Fam96a-deficient macrophages inhibited the induction of immune effector molecules, including iNOS, by suppressing interferon/STAT1 signaling. Fam96a regulated macrophage polarization process was in part dependent on its ability to fine-tuning intracellular iron (Fe) homeostasis in response to inflammatory stimuli. In addition, Fam96a regulated the mitochondrial oxidative phosphorylation or related events that involved in control of T. gondii. CONCLUSIONS/SIGNIFICANCE: All these findings suggest that Fam96a ablation in macrophages disrupts iron homeostasis and inhibits immune effector molecules, which may aggravate both acute and chronic toxoplasmosis. It highlights that Fam96a may autonomously act as a critical gatekeeper of T. gondii control in macrophages.

Acute response to pathogens in the early human placenta at single-cell resolution.

The placenta is a selective maternal-fetal barrier that provides nourishment and protection from infections. However, certain pathogens can attach to and even cross the placenta, causing pregnancy complications with potential lifelong impacts on the child's health. Here, we profiled at the single-cell level the placental responses to three pathogens associated with intrauterine complications-Plasmodium falciparum, Listeria monocytogenes, and Toxoplasma gondii. We found that upon exposure to the pathogens, all placental lineages trigger inflammatory responses that may compromise placental function. Additionally, we characterized the responses of fetal macrophages known as Hofbauer cells (HBCs) to each pathogen and propose that they are the probable niche for T. gondii. Finally, we revealed how P. falciparum adapts to the placental microenvironment by modulating protein export into the host erythrocyte and nutrient uptake pathways. Altogether, we have defined the cellular networks and signaling pathways mediating acute placental inflammatory responses that could contribute to pregnancy complications.

A CTCF-binding site in the Mdm1-Il22-Ifng locus shapes cytokine expression profiles and plays a critical role in early Th1 cell fate specification.

Cytokine expression during T cell differentiation is a highly regulated process that involves long-range promoter-enhancer and CTCF-CTCF contacts at cytokine loci. Here, we investigated the impact of dynamic chromatin loop formation within the topologically associating domain (TAD) in regulating the expression of interferon gamma (IFN-γ) and interleukin-22 (IL-22); these cytokine loci are closely located in the genome and are associated with complex enhancer landscapes, which are selectively active in type 1 and type 3 lymphocytes. In situ Hi-C analyses revealed inducible TADs that insulated Ifng and Il22 enhancers during Th1 cell differentiation. Targeted deletion of a 17 bp boundary motif of these TADs imbalanced Th1- and Th17-associated immunity, both in vitro and in vivo, upon Toxoplasma gondii infection. In contrast, this boundary element was dispensable for cytokine regulation in natural killer cells. Our findings suggest that precise cytokine regulation relies on lineage- and developmental stage-specific interactions of 3D chromatin architectures and enhancer landscapes.

Translation initiation factor eIF1.2 promotes Toxoplasma stage conversion by regulating levels of key differentiation factors.

The parasite Toxoplasma gondii persists in its hosts by converting from replicating tachyzoites to latent bradyzoites housed in tissue cysts. The molecular mechanisms that mediate T. gondii differentiation remain poorly understood. Through a mutagenesis screen, we identified translation initiation factor eIF1.2 as a critical factor for T. gondii differentiation. A F97L mutation in eIF1.2 or the genetic ablation of eIF1.2 (∆eif1.2) markedly impeded bradyzoite cyst formation in vitro and in vivo. We demonstrated, at single-molecule level, that the eIF1.2 F97L mutation impacts the scanning process of the ribosome preinitiation complex on a model mRNA. RNA sequencing and ribosome profiling experiments unveiled that ∆eif1.2 parasites are defective in upregulating bradyzoite induction factors BFD1 and BFD2 during stress-induced differentiation. Forced expression of BFD1 or BFD2 significantly restored differentiation in ∆eif1.2 parasites. Together, our findings suggest that eIF1.2 functions by regulating the translation of key differentiation factors necessary to establish chronic toxoplasmosis.

Seroprevalence of Toxoplasma gondii and Borrelia burgdorferi infections in patients with multiple sclerosis in Poland.

Multiple sclerosis (MS) is a chronic, demyelinating disease of the central nervous system that affects mainly young people. It is believed that the autoimmune process observed in the pathogenesis of MS is influenced by a complex interaction between genetic and environmental factors, including infectious agents. The results of this study suggest the protective role of Toxoplasma gondii infections in MS. Interestingly, high Toxoplasma IgM seropositivity in MS patients receiving immunomodulatory drugs (IMDs) was identified. On the other hand, Borrelia infections seem to be positively associated with MS. Although the interpretation of our results is limited by the retrospective nature of the studies, the results strongly indicate that further experimental and clinical studies are needed to explain the role of infectious agents in the development and pathophysiological mechanisms of MS.

Single-cell transcriptomic analysis of decidual immune cell landscape in the occurrence of adverse pregnancy outcomes induced by Toxoplasma gondii infection.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that can lead to adverse pregnancy outcomes, particularly in early pregnancy. Previous studies have illustrated the landscape of decidual immune cells. However, the landscape of decidual immune cells in the maternal-fetal microenvironment during T. gondii infection remains unknown. METHODS: In this study, we employed single-cell RNA sequencing to analyze the changes in human decidual immune cells following T. gondii infection. The results of scRNA-seq were further validated with flow cytometry, reverse transcription-polymerase chain reaction, western blot, and immunofluorescence staining. RESULTS: Our results showed that the proportion of 17 decidual immune cell clusters and the expression levels of 21 genes were changed after T. gondii infection. Differential gene analysis demonstrated that T. gondii infection induced the differential expression of 279, 312, and 380 genes in decidual NK cells (dNK), decidual macrophages (dMφ), and decidual T cells (dT), respectively. Our results revealed for the first time that several previously unknown molecules in decidual immune cells changed following infection. This result revealed that the function of maternal-fetal immune tolerance declined, whereas the killing ability of decidual immune cells enhanced, eventually contributing to the occurrence of adverse pregnancy outcomes. CONCLUSIONS: This study provides valuable resource for uncovering several novel molecules that play an important role in the occurrence of abnormal pregnancy outcomes induced by T. gondii infection.

Human T-cell activation with Toxoplasma gondii antigens loaded in maltodextrin nanoparticles.

Toxoplasmosis is the most prevalent parasitic zoonosis worldwide, causing ocular and neurological diseases. No vaccine has been approved for human use. We evaluated the response of peripheral blood mononuclear cells (PBMCs) to a novel construct of Toxoplasma gondii total antigen in maltodextrin nanoparticles (NP/TE) in individuals with varying infectious statuses (uninfected, chronic asymptomatic, or ocular toxoplasmosis). We analyzed the concentration of IFN-γ after NP/TE ex vivo stimulation using ELISA and the immunophenotypes of CD4+ and CD8+ cell populations using flow cytometry. In addition, serotyping of individuals with toxoplasmosis was performed by ELISA using GRA6-derived polypeptides. Low doses of NP/TE stimulation (0.9 µg NP/0.3 µg TE) achieved IFN-γ-specific production in previously exposed human PBMCs without significant differences in the infecting serotype. Increased IFN-γ expression in CD4+ effector memory cell subsets was found in patients with ocular toxoplasmosis with NP/TE but not with TE alone. This is the first study to show how T-cell subsets respond to ex vivo stimulation with a vaccine candidate for human toxoplasmosis, providing crucial insights for future clinical trials.

Seroprevalence and sociodemographic characteristics of Toxoplasma gondii infection in patients with psychiatric disorders in Malaysia.

Toxoplasma gondii is a neurotropic protozoan parasite that affects neuronal processing in the brain. This study aimed to investigate the prevalence of T. gondii infection in psychiatric disorder patients. We also investigated the potential association between sociodemographic, clinical manifestation, and behavior of Toxoplasma-seropositive patients with psychiatric disorders. Commercial ELISAs (IgG, IgM, and IgG avidity) using serum and PCR using buffy coat were performed on samples from 54 individuals in each of the following groups: patients diagnosed with depressive disorder, bipolar disorder, and schizophrenia, as well as psychiatrically healthy subjects (control group). They were recruited from the Hospital Universiti Sains Malaysia in Kelantan, Malaysia. Of 54 patients with depressive disorder, 24/54 (44.4 %) were seropositive for IgG, and four (16.7 %) were IgG+/IgM+. Among the latter, a high avidity index indicating a past infection was observed in half of the samples (50.0 %), and the other half (50.0 %) showed a low avidity index, indicating a possible recent infection. Meanwhile, 30/54 (55.6 %) patients with bipolar disorder were seropositive for IgG+, five (16.7 %) were IgG+/IgM+, and four of them had a high avidity index, and one had a low avidity index. Patients with schizophrenia showed 29/54 (53.7 %) seropositive for IgG, two of them (6.9 %) were IgG+/IgM+; one of latter had a high avidity index, and one had a low avidity index. Of 54 people in the control group, 37.0 % (20/54) were seropositive for T. gondii IgG antibodies. However, no significant difference was observed in seroprevalence between the control group and each patient group. No PCR-positive results were documented. A Chi-Square and multiple logistic regression showed that age (p = 0.031), close contact with cats/pets (p = 0.033) and contact with soil (p = 0.012) were significantly associated with Toxoplasma seropositivity in patients with psychiatric disorders. Additional research is needed to elucidate the causal relationships and underlying mechanisms.

AP2XII-1 and AP2XI-2 Suppress Schizogony Gene Expression in Toxoplasma gondii.

Toxoplasma gondii is an intracellular parasite that is important in medicine and veterinary science and undergoes distinct developmental transitions in its intermediate and definitive hosts. The switch between stages of T. gondii is meticulously regulated by a variety of factors. Previous studies have explored the role of the microrchidia (MORC) protein complex as a transcriptional suppressor of sexual commitment. By utilizing immunoprecipitation and mass spectrometry, constituents of this protein complex have been identified, including MORC, Histone Deacetylase 3 (HDAC3), and several ApiAP2 transcription factors. Conditional knockout of MORC or inhibition of HDAC3 results in upregulation of a set of genes associated with schizogony and sexual stages in T. gondii tachyzoites. Here, our focus extends to two primary ApiAP2s (AP2XII-1 and AP2XI-2), demonstrating their significant impact on the fitness of asexual tachyzoites and their target genes. Notably, the targeted disruption of AP2XII-1 and AP2XI-2 resulted in a profound alteration in merozoite-specific genes targeted by the MORC-HDAC3 complex. Additionally, considerable overlap was observed in downstream gene profiles between AP2XII-1 and AP2XI-2, with AP2XII-1 specifically binding to a subset of ApiAP2 transcription factors, including AP2XI-2. These findings reveal an intricate cascade of ApiAP2 regulatory networks involved in T. gondii schizogony development, orchestrated by AP2XII-1 and AP2XI-2. This study provides valuable insights into the transcriptional regulation of T. gondii growth and development, shedding light on the intricate life cycle of this parasitic pathogen.

Systematic Review on the Relationship between Toxoplasmosis and Mental Disorders.

BACKGROUND: Toxoplasmosis is a worldwide parasitic zoonosis caused by the protozoan Toxoplasma gondii. In cases of vertical infection, and in immunosuppressed people by the human immunodeficiency virus (HIV) serious clinical conditions may appear, while immunocompetent people do not present symptoms. However, T. gondii infection has been linked to several mental disorders for decades. OBJECTIVE: To substantiate the possible relationship between T. gondii and mental disorders and suggest control and prevention strategies. MATERIAL AND METHODS: A systematic review has been carried out to analyze the relationship between T. gondii exposure (presence of IgG) and the onset of mental disorders in minors and adults. The etiopathogenic mechanisms described by the authors have also been included and the systems of surveillance, prevention and control of infection have been evaluated. RESULTS: Several processes linked to the presence of cysts and the reactivation of the parasite in certain situations produce an immune and inflammatory response. Also, direct and indirect actions on different neurotransmitters. These mechanisms, together with other environmental and genetic factors, would predispose to different psychiatric pathologies. CONCLUSIONS: Due to the limits of the study, no conclusions can be drawn in childhood and adolescence. However, the results of this systematic review show a possible association of schizophrenia, bipolar disorder and compulsive disorder with T. gondii infection in adults. There is a need to improve control, integrated surveillance and extend prevention measures to the entire population.

Effect of concurrent infection of Helicobacter pylori with Toxoplasma gondii infection on gastric pathology.

BACKGROUND: Toxoplasma gondii (T. gondii) and Helicobacter pylori (H. pylori) are among the most prevalent foodborne parasitic and bacterial infections worldwide. However, the concurrent impact of coinfection on gastric pathology has yet to be studied in depth. The effect of coinfection generally either adds a synergetic or antagonistic impact; we aimed in the current work to assess the impact of T. gondii coinfection on the progression of H. pylori-associated gastric pathology and reporting H. pylori virulent strains. The study was conducted on 82 patients complaining of persistent gastrointestinal symptoms with failed treatment response and prone to endoscopy. They were subjected to stool examination to detect H. pylori antigen, serological screening for latent toxoplasmosis, endoscopy, histopathological examination, and molecular detection of H. pylori virulence strains in gastric biopsies. Out of the 82 patients, 62 patients were positive for H. pylori antigen in stool and 55 patients confirmed positivity by histopathology; out of them, 37 patients had isolated Vac As1 variants, 11 patients had combined Vac As1 and Cag A variants, and 7 patients had combined Vac As1, Cag A and VacAs2 variants. Patients with the combined two or three variances showed significantly deteriorated histopathological features than patients with a single Vac As1 variant (P < 0.05). Latent toxoplasmosis was positive among 35/82 patients. Combined H. pylori and Toxoplasma gondii infection had significantly marked inflammation than patients with isolated infection (P < 0.05). CONCLUSION: Screening for toxoplasmosis among H. pylori-infected patients is recommended as it is considered a potential risk factor for gastric inflammation severity. H. pylori gastric inflammation may be heightened by Toxoplasma coinfection.

IL-12 Mediates T-bet-Expressing Myeloid Cell-Dependent Host Resistance against Toxoplasma gondii.

To defend against intracellular pathogens such as Toxoplasma gondii, the host generates a robust type 1 immune response. Specifically, host defense against T. gondii is defined by an IL-12-dependent IFN-γ response that is critical for host resistance. Previously, we demonstrated that host resistance is mediated by T-bet-dependent ILC-derived IFN-γ by maintaining IRF8+ conventional type 1 dendritic cells during parasitic infection. Therefore, we hypothesized that innate lymphoid cells are indispensable for host survival. Surprisingly, we observed that T-bet-deficient mice succumb to infection quicker than do mice lacking lymphocytes, suggesting an unknown T-bet-dependent-mediated host defense pathway. Analysis of parasite-mediated inflammatory myeloid cells revealed a novel subpopulation of T-bet+ myeloid cells (TMCs). Our results reveal that TMCs have the largest intracellular parasite burden compared with other professional phagocytes, suggesting they are associated with active killing of T. gondii. Mechanistically, we established that IL-12 is necessary for the induction of inflammatory TMCs during infection and these cells are linked to a role in host survival.

Inhibition of Foxp3 expression in the placenta of mice infected intraperitoneally by toxoplasma gondii tachyzoites: insights into the PPARγ/miR-7b-5p/Sp1 signaling pathway.

BACKGROUND: Toxoplasma gondii, an obligate intracellular parasitic protozoa, infects approximately 30% of the global population. Contracting T. gondii at the primary infection of the mother can result in neonatal microcephaly, chorioretinitis, hydrocephalus, or mortality. Our previous study indicated that pregnant mice infected with T. gondii displayed a decrease in both the number and the suppressive ability of regulatory T cells, accompanied by the reduced Forkhead box P3 (Foxp3). Numerous studies have proved that microRNAs (miRNAs) are implicated in T. gondii infection, but there is meager evidence on the relationship between alterations of miRNAs and downregulation of Foxp3 induced by T. gondii. METHODS: Quantitative reverse transcription polymerase chain reaction was utilized to detect the transcriptions of miRNAs and Foxp3. Protein blotting and immunofluorescence were used to detect the expressions of Foxp3 and related transcription factors. The structure of mouse placenta was observed by hematoxylin and eosin (HE) staining. To examine the activity of miR-7b promoter and whether miR-7b-5p targets Sp1 to suppress Foxp3 expression, we constructed recombinant plasmids containing the full-length/truncated/mutant miR-7b promoter sequence or wildtype/mutant of Sp1 3' untranslated region (3' UTR) to detect the fluorescence activity in EL4 cells. RESULTS: In T. gondii-infected mice, miR-7b transcription was significantly elevated, while Foxp3 expression was decreased in the placenta. In vitro, miR-7b mimics downregulated Foxp3 expression, whereas its inhibitors significantly upregulated Foxp3 expression. miR-7b promoter activity was elevated upon the stimulation of T. gondii antigens, which was mitigated by co-transfection of mutant miR-7b promoter lacking peroxisome proliferator-activated receptor γ (PPARγ) target sites. Additionally, miR-7b mimics diminished Sp1 expression, while miR-7b inhibitors elevated its expression. miR-7b mimics deceased the fluorescence activity of Sp1 3' untranslated region (3' UTR), but it failed to impact the fluorescence activity upon the co-transfection of mutant Sp1 3' UTR lacking miR-7b target site. CONCLUSIONS: T. gondii infection and antigens promote miR-7b transcription but inhibit Foxp3 protein and gene levels. T. gondii antigens promote miR-7b promoter activity by a PPARγ-dependent mechanism. miR-7b directly binds to Sp1 3' UTR to repress Sp1 expression. Understanding the regulatory functions by which T. gondii-induced miR-7b suppresses Foxp3 expression can provide new perspectives for the possible therapeutic avenue of T. gondii-induced adverse pregnancy outcomes.