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

BACKGROUND: Toxoplasma 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. METHODS: iTRAQ-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). RESULTS: A 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). CONCLUSIONS: The 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.

Urine proteomics for profiling of mouse toxoplasmosis using liquid chromatography tandem mass spectrometry analysis.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that causes toxoplasmosis. Urine is an easily obtained clinical sample that has been widely applied for diagnostic purposes. However, changes in the urinary proteome during T. gondii infection have never been investigated. METHODS: Twenty four-hour urine samples were obtained from BALB/c mice with acute infection [11 days post infection (DPI)], mice with chronic infection (35 DPI) and healthy controls, and were analyzed using a label-free liquid chromatography tandem mass spectrometry analysis. RESULTS: We identified a total of 13,414 peptides on 1802 proteins, of which 169 and 47 proteins were significantly differentially expressed at acute and chronic infection phases, respectively. Clustering analysis revealed obvious differences in proteome profiles among all groups. Gene ontology analysis showed that a large number of differentially expressed proteins (DEPs) detected in acute infection were associated with biological binding activity and single-organism processes. KEGG pathway enrichment analysis showed that the majority of these DEPs were involved in disease-related and metabolic pathways. CONCLUSIONS: Our findings revealed global reprogramming of the urine proteome following T. gondii infection, and data obtained in this study will enhance our understanding of the host responses to T. gondii infection and lead to the identification of new diagnostic biomarkers.

Molecular and biological analysis revealed genetic diversity and high virulence strain of Toxoplasma gondii in Japan.

Toxoplasma gondii is classified into 16 haplogroups based on a worldwide genotyping study of the parasite. However, only a few isolates from Japan were included in this analysis. To conduct more precise genotyping of T. gondii, we examined the genotypes of Japanese isolates in this study. DNA sequences of 6 loci were determined in 17 Japanese isolates and compared with those of strains of 16 haplogroups. As a result, Japanese isolates were classified into four groups. We investigated the virulence of some Japanese isolates and found a highly virulent strain in mice, comparable to that of RH strain, although this Japanese isolate was sister to strains of haplogroup 2, which show moderate virulence in mice. We further investigated whether this high virulence isolate had different virulence mechanism and strategy to adapt to Japanese host from other strains by comparing the virulence-related genes, ROP5, 18 and the immunomodulatory gene, ROP16 of the isolate with those of archetypical strains (GT1, ME49 and VEG). This analysis indicated the high virulence of the isolate in mice was partly explained by gene sequences of ROP5 and ROP16. These findings lead to the elucidation of biodiversity of T. gondii and have potential to optimize the diagnostic protocol.

Toxoplasma Infection Induces Sustained Up-Regulation of Complement Factor B and C5a Receptor in the Mouse Brain via Microglial Activation: Implication for the Alternative Complement Pathway Activation and Anaphylatoxin Signaling in Cerebral Toxoplasmosis.

Toxoplasma gondii is a neurotropic protozoan parasite, which is linked to neurological manifestations in immunocompromised individuals as well as severe neurodevelopmental sequelae in congenital toxoplasmosis. While the complement system is the first line of host defense that plays a significant role in the prevention of parasite dissemination, Toxoplasma artfully evades complement-mediated clearance via recruiting complement regulatory proteins to their surface. On the other hand, the details of Toxoplasma and the complement system interaction in the brain parenchyma remain elusive. In this study, infection-induced changes in the mRNA levels of complement components were analyzed by quantitative PCR using a murine Toxoplasma infection model in vivo and primary glial cells in vitro. In addition to the core components C3 and C1q, anaphylatoxin C3a and C5a receptors (C3aR and C5aR1), as well as alternative complement pathway components properdin (CFP) and factor B (CFB), were significantly upregulated 2 weeks after inoculation. Two months post-infection, CFB, C3, C3aR, and C5aR1 expression remained higher than in controls, while CFP upregulation was transient. Furthermore, Toxoplasma infection induced significant increase in CFP, CFB, C3, and C5aR1 in mixed glial culture, which was abrogated when microglial activation was inhibited by pre-treatment with minocycline. This study sheds new light on the roles for the complement system in the brain parenchyma during Toxoplasma infection, which may lead to the development of novel therapeutic approaches to Toxoplasma infection-induced neurological disorders.

Toxoplasma gondii induces prolonged host epidermal growth factor receptor signalling to prevent parasite elimination by autophagy: Perspectives for in vivo control of the parasite.

Toxoplasma gondii causes retinitis and encephalitis. Avoiding targeting by autophagosomes is key for its survival because T. gondii cannot withstand lysosomal degradation. During invasion of host cells, T. gondii triggers epidermal growth factor receptor (EGFR) signalling enabling the parasite to avoid initial autophagic targeting. However, autophagy is a constitutive process indicating that the parasite may also use a strategy operative beyond invasion to maintain blockade of autophagic targeting. Finding that such a strategy exists would be important because it could lead to inhibition of host cell signalling as a novel approach to kill the parasite in previously infected cells and treat toxoplasmosis. We report that T. gondii induced prolonged EGFR autophosphorylation. This effect was mediated by PKCα/PKCß âž” Src because T. gondii caused prolonged activation of these molecules and their knockdown or incubation with inhibitors of PKCα/PKCß or Src after host cell invasion impaired sustained EGFR autophosphorylation. Addition of EGFR tyrosine kinase inhibitor (TKI) to previously infected cells led to parasite entrapment by LC3 and LAMP-1 and pathogen killing dependent on the autophagy proteins ULK1 and Beclin 1 as well as lysosomal enzymes. Administration of gefitinib (EGFR TKI) to mice with ocular and cerebral toxoplasmosis resulted in disease control that was dependent on Beclin 1. Thus, T. gondii promotes its survival through sustained EGFR signalling driven by PKCα/ß âž” Src, and inhibition of EGFR controls pre-established toxoplasmosis.

TLR11-independent inflammasome activation is critical for CD4+ T cell-derived IFN-γ production and host resistance to Toxoplasma gondii.

Innate recognition of invading intracellular pathogens is essential for regulating robust and rapid CD4+ T cell effector function, which is critical for host-mediated immunity. The intracellular apicomplexan parasite, Toxoplasma gondii, is capable of infecting almost any nucleated cell of warm-blooded animals, including humans, and establishing tissue cysts that persist throughout the lifetime of the host. Recognition of T. gondii by TLRs is essential for robust IL-12 and IFN-γ production, two major cytokines involved in host resistance to the parasite. In the murine model of infection, robust IL-12 and IFN-γ production have been largely attributed to T. gondii profilin recognition by the TLR11 and TLR12 heterodimer complex, resulting in Myd88-dependent IL-12 production. However, TLR11 or TLR12 deficiency failed to recapitulate the acute susceptibility to T. gondii infection seen in Myd88-/- mice. T. gondii triggers inflammasome activation in a caspase-1-dependent manner resulting in cytokine release; however, it remains undetermined if parasite-mediated inflammasome activation impacts IFN-γ production and host resistance to the parasite. Using mice which lack different inflammasome components, we observed that the inflammasome played a limited role in host resistance when TLR11 remained functional. Strikingly, in the absence of TLR11, caspase-1 and -11 played a significant role for robust CD4+ TH1-derived IFN-γ responses and host survival. Moreover, we demonstrated that in the absence of TLR11, production of the caspase-1-dependent cytokine IL-18 was sufficient and necessary for CD4+ T cell-derived IFN-γ responses. Mechanistically, we established that T. gondii-mediated activation of the inflammasome and IL-18 were critical to maintain robust CD4+ TH1 IFN-γ responses during parasite infection in the absence of TLR11.

The lectin-specific activity of Toxoplasma gondii microneme proteins 1 and 4 binds Toll-like receptor 2 and 4 N-glycans to regulate innate immune priming.

Infection of host cells by Toxoplasma gondii is an active process, which is regulated by secretion of microneme (MICs) and rhoptry proteins (ROPs and RONs) from specialized organelles in the apical pole of the parasite. MIC1, MIC4 and MIC6 assemble into an adhesin complex secreted on the parasite surface that functions to promote infection competency. MIC1 and MIC4 are known to bind terminal sialic acid residues and galactose residues, respectively and to induce IL-12 production from splenocytes. Here we show that rMIC1- and rMIC4-stimulated dendritic cells and macrophages produce proinflammatory cytokines, and they do so by engaging TLR2 and TLR4. This process depends on sugar recognition, since point mutations in the carbohydrate-recognition domains (CRD) of rMIC1 and rMIC4 inhibit innate immune cells activation. HEK cells transfected with TLR2 glycomutants were selectively unresponsive to MICs. Following in vitro infection, parasites lacking MIC1 or MIC4, as well as expressing MIC proteins with point mutations in their CRD, failed to induce wild-type (WT) levels of IL-12 secretion by innate immune cells. However, only MIC1 was shown to impact systemic levels of IL-12 and IFN-γ in vivo. Together, our data show that MIC1 and MIC4 interact physically with TLR2 and TLR4 N-glycans to trigger IL-12 responses, and MIC1 is playing a significant role in vivo by altering T. gondii infection competency and murine pathogenesis.

Comparison of splenocyte microRNA expression profiles of pigs during acute and chronic toxoplasmosis.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that infects humans and other warm-blooded animals. Previous quantitative proteomic analyses of infected host cells revealed that the expression of many host proteins is modulated by T. gondii infection. However, at present limited data are available on the differentially expressed miRNAs (DEMs) associated with the pathology and host immune responses induced by acute and chronic infection with T. gondii in pigs in vivo. In this study, high-throughput sequencing was used to investigate expression profiles of spleen miRNAs at 10, 25 and 50 days post-infection (DPI) in pigs infected with Chinese I genotype strain T. gondii isolated from a dead pig. RESULTS: When compared to the control group, 34, 6 and 86 DEMs were found in spleens of infected pigs at 10, 25 and 50 DPI, respectively. Gene Ontology (GO) enrichment analysis of the target genes of DEMs showed that no GO terms were enriched at 25 DPI, whereas 28 and 241 GO terms, of which two and 215 were sample-specific, were significantly enriched at 10 and 50 DPI, respectively. The top 20 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of the target genes of DEMs included signal transduction, immune system, metabolism and diseases. miRNA-gene network analysis revealed that the DEMs played important roles in the host immune response to T. gondii infection by modulating expression levels of cellular immunity-related cytokines and immune-related C-type lectins. CONCLUSION: Our results not only showed that host miRNA expression is altered by T. gondii but also revealed differences in the regulation of key biological processes and pathways involved in host responses to acute versus chronic T. gondii infection. This will aid future research into miRNA-target interactions during T. gondii infection in pigs and the development of novel therapies against T. gondii.

Expression profile of microRNAs in porcine alveolar macrophages after Toxoplasma gondii infection.

BACKGROUND: Toxoplasma gondii is an apicomplexan protozoan parasite that can cause serious clinical illnesses in both humans and animals. microRNAs (miRNAs) are non-protein-coding RNAs that can regulate the expression of target genes. A previous study found that many miRNAs were differentially expressed after T. gondii infection and exert significant effects and revealed that both host survival and the virulence of different strains can be regulated by different miRNAs. Macrophages play an important role in T. gondii infection, but few studies have investigated the relationship between miRNAs and porcine alveolar macrophages infected with T. gondii. METHODS: Porcine alveolar macrophages (3D4-21) were infected with the RH (Type I) and Me49 (Type II) strains of T. gondii for 12 h and 24 h and then harvested. miRNA libraries were generated using the NEBNext® Multiplex Small RNA Library Prep Set for Illumina® (NEB, USA), and the miRNA expression levels were estimated based on transcripts per million reads (TPM). RESULTS: Our study generated six miRNA expression profiles from macrophages infected with RH and Me49 compared with the control groups. The comparison of the T. gondii-infected and uninfected samples identified 81 differentially expressed miRNAs, including 36 novel miRNAs and 45 mature miRNAs. The target genes of these differentially expressed miRNAs were predicted using miRanda software, and ssc-miR-127 and ssc-miR-143-3p were predicted to regulate nitric oxide synthase 1 (NOS1) and nitric oxide synthase 3 (NOS3), respectively, which play essential roles in synthesizing nitric oxide (NO) by oxidizing L-arginine. These genes were differentially expressed in both the RH- and Me49-infected groups. A KEGG enrichment analysis indicated that the predicted target genes were involved in multiple signaling pathways, including FcγR-mediated phagocytosis, the AMPK signaling pathway, the mTOR signaling pathway, and the FcγRI signaling pathway, all of which are indispensable for the normal functioning of porcine alveolar macrophages. CONCLUSIONS: Our results provide data on the miRNA profile of porcine alveolar macrophages infected with T. gondii. To our knowledge, this study provides the first demonstration of the relationship between miRNA and macrophages of swine origin. Understanding the functions of these regulated miRNAs will aid the investigation of T. gondii infectious diseases, and the differentially expressed miRNAs might be candidate drug targets for T. gondii infection in pigs.

A Double-Edged Sword: Complement Component 3 in Toxoplasma gondii Infection.

Sprague Dawley rats and Kunming (KM) mice are artificially infected with type II Toxoplasma gondii strain Prugniaud (Pru) to generate toxoplasmosis, which is a fatal disease mediated by T. gondii invasion of the central nervous system (CNS) by unknown mechanisms. The aim is to explore the mechanism of differential susceptibility of mice and rats to T. gondii infection. Therefore, a strategy of isobaric tags for relative and absolute quantitation (iTRAQ) is established to identify differentially expressed proteins (DEPs) in the rats' and the mice's brains compared to the healthy groups. In KM mice, which is susceptible to T. gondii infection, complement component 3 (C3) is upregulated and the tight junction (TJ) pathway shows a disorder. It is presumed that T. gondii-stimulated C3 disrupts the TJ of the blood-brain barrier in the CNS. This effect allows more T. gondii passing to the brain through the intercellular space.

Acute Toxoplasma Gondii Infection in Cats Induced Tissue-Specific Transcriptional Response Dominated by Immune Signatures.

RNA-sequencing was used to detect transcriptional changes in six tissues of cats, seven days after T. gondii infection. A total of 737 genes were differentially expressed (DEGs), of which 410 were up-regulated and 327 were down-regulated. The liver exhibited 151 DEGs, lung (149 DEGs), small intestine (130 DEGs), heart (123 DEGs), brain (104 DEGs), and spleen (80 DEGs)-suggesting tissue-specific transcriptional patterns. Gene ontology and KEGG analyses identified DEGs enriched in immune pathways, such as cytokine-cytokine receptor interaction, Jak-STAT signaling pathway, NOD-like receptor signaling pathway, NF-kappa B signaling pathway, MAPK signaling pathway, T cell receptor signaling pathway, and the cytosolic DNA sensing pathway. C-X-C motif chemokine 10 (CXCL10) was involved in most of the immune-related pathways. PI3K/Akt expression was down-regulated in all tissues, except the spleen. The genes for phosphatase, indoleamine 2,3-dioxygenase, Hes Family BHLH Transcription Factor 1, and guanylate-binding protein 5, playing various roles in immune defense, were co-expressed across various feline tissues. Multivariate K-means clustering analysis produced seven gene clusters featuring similar gene expression patterns specific to individual tissues, with lung tissue cluster having the largest number of DEGs. These findings suggest the presence of a broad immune defense mechanism across various tissues in cats against acute T. gondii infection.

Resistance to Chronic Toxoplasma gondii Infection Induced by a DNA Vaccine Expressing GRA16.

Toxoplasma gondii can infect all warm-blooded animals including human beings. T. gondii dense granule protein 16 (TgGRA16) as a crucial virulence factor could modulate the host gene expression. Here, a DNA vaccine expressing TgGRA16 was constructed to explore the protective efficacy against T. gondii infection in Kunming mice. The immune responses induced by pVAX-GRA16 were also evaluated. Mice immunized with pVAX-GRA16 could elicit higher levels of specific IgG antibody and strong cellular response compared to those in controls. The DNA vaccination significantly increased the levels of cytokines (IFN-γ, IL-2, IL-4, and IL-10) and the percentages of CD4+ and CD8+ T cells in mice. After lethal challenge, mice immunized with pVAX-GRA16 (8.4 ± 0.78 days) did not show a significant longer survival time than that in controls (7.1 ± 0.30 days) (p > 0.05). However, in chronic toxoplasmosis model (administration of 10 brain cysts of PRU strain orally), numbers of tissue cysts in mice immunized with pVAX-GRA16 were significantly reduced compared to those in controls (p < 0.05) and the rate of reduction could reach 43.89%. The results indicated that the TgGRA16 would be a promising vaccine candidate for further development of effective epitope-based vaccines against chronic T. gondii infection in mice.

Evidence for host genetic regulation of altered lipid metabolism in experimental toxoplasmosis supported with gene data mining results.

Toxoplasma gondii is one of the most successful parasites on Earth, infecting a wide array of mammals including one third of the global human population. The obligate intracellular protozoon is not capable of synthesizing cholesterol (Chl), and thus depends on uptake of host Chl for its own development. To explore the genetic regulation of previously observed lipid metabolism alterations during acute murine T. gondii infection, we here assessed total Chl and its fractions in serum and selected tissues at the pathophysiological and molecular level, and integrated the observed gene expression of selected molecules relevant for Chl metabolism, including its biosynthetic and export KEGG pathways, with the results of published transcriptomes obtained in similar murine models of T. gondii infection. The serum lipid status as well as the transcript levels of relevant genes in the brain and the liver were assessed in experimental models of acute and chronic toxoplasmosis in wild-type mice. The results showed that acute infection was associated with a decrease in Chl content in both the liver and periphery (brain, peripheral lymphocytes), and a decrease in Chl reverse transport. In contrast, in chronic infection, a return to normal levels of Chl metabolism has been noted. These changes corresponded to the brain and liver gene expression results as well as to data obtained via mining. We propose that the observed changes in Chl metabolism are part of the host defense response. Further insight into the lipid metabolism in T. gondii infection may provide novel targets for therapeutic agents.

Global miRNA expression profiling of domestic cat livers following acute Toxoplasma gondii infection.

Although microRNAs (miRNAs) play an important role in liver homeostasis, the extent to which they can be altered by Toxoplasma gondii infection is unknown. Here, we utilized small RNA sequencing and bioinformatic analyses to characterize miRNA expression profiles in the liver of domestic cats at 7 days after oral infection with T. gondii (Type II) strain. A total of 384 miRNAs were identified and 82 were differentially expressed, of which 33 were up-regulated and 49 down-regulated. Also, 5690 predicted host gene targets for the differentially expressed miRNAs were identified using the bioinformatic algorithm miRanda. Gene ontology analysis revealed that the predicted gene targets of the dysregulated miRNAs were significantly enriched in apoptosis. Kyoto Encyclopedia of Genes and Genomes analysis showed that the predicted gene targets were involved in several pathways, including acute myeloid leukemia, central carbon metabolism in cancer, choline metabolism in cancer, estrogen signaling pathway, fatty acid degradation, lysosome, nucleotide excision repair, progesterone-mediated oocyte maturation, and VEGF signaling pathway. The expression level of 6 upregulated miRNAs (mmu-miR-21a-5p, mmu-miR-20a-5p, mmu-miR-17-5p, mmu-miR-30e-3p, mmu-miR-142a-3p, and mmu-miR-106b-3p) was confirmed by stem-loop quantitative reverse transcription PCR, which yielded results consistent with the sequencing data. These findings expand our understanding of the regulatory mechanisms of miRNAs underlying T. gondii pathogenesis and contribute new database information on cat miRNAs, opening a new perspective on the prevention and treatment of T. gondii infection.

Detection of Natural Toxoplasma gondii Infection in Chicken in Thika Region of Kenya Using Nested Polymerase Chain Reaction.

The detection of Toxoplasma gondii in free-range chickens is a good indicator of possible risk to human beings. The aim of this study was to investigate the occurrence of T. gondii in free-range chicken using polymerase chain reaction (PCR). Brain samples from 105 free-range chickens from three administrative areas in Thika region, Kenya, were collected, DNA-extracted, and analyzed using PCR to detect presence of T. gondii. The overall prevalence of T. gondii in all the three areas was 79.0% (95% CI: 70.0-86.4%) and the prevalence across the three areas was not significantly different (P = 0.5088; χ2 = 1.354). Female chickens had higher (79.4%) prevalence than males (78.6%), although the difference was not significant (P = 0.922, χ2 = 0.01). However, chickens that were more than 2 years old had significantly (P = 0.003; χ2 = 11.87) higher prevalence compared to younger ones. The study indicates that there was a high occurrence of T. gondii infection in free-range chickens from Thika region and that the infection rate is age dependent. Further studies should be carried out to determine the possible role of roaming chickens in the epidemiology of the disease among humans in the area.

Increased Cerebral Tff1 Expression in Two Murine Models of Neuroinflammation.

BACKGROUND/AIMS: The trefoil factor family (TFF) peptide TFF1 is a typical secretory product of the gastric mucosa and a very low level of expression occurs in nearly all regions of the murine brain. TFF1 possesses a lectin activity and binding to a plethora of transmembrane glycoproteins could explain the diverse biological effects of TFF1 (e.g., anti-apoptotic effect). It was the aim to test whether TFF expression is changed during neuroinflammation. METHODS: Expression profiling was performed using semi-quantitative RT-PCR analyses in two murine models of neuroinflammation, i.e. Toxoplasma gondii-induced encephalitis and experimental autoimmune encephalomyelitis (EAE), the latter being the most common animal model of multiple sclerosis. Tff1 expression was also localized using RNA in situ hybridization histochemistry. RESULTS: We report for the first time on a significant transcriptional induction in cerebral Tff1 expression in both T. gondii-induced encephalitis and EAE. In contrast, Tff2 and Tff3 expression were not altered. Tff1 transcripts were predominantly localized in the internal granular layer of the cerebellum indicating neuronal expression. Furthermore, also glial cells are expected to express Tff1. Characterization of both experimental models by expression profiling (e.g., inflammasome sensors, inflammatory cytokines, microglial marker Iba1, ependymin related protein 1) revealed differences concerning the expression of the inflammasome sensor Nlrp1 and interleukin 17a. CONCLUSION: The up-regulated expression of Tff1 is probably the result of a complex inflammatory process as its expression is induced by tumor necrosis factor α as well as interleukins 1ß and 17. However on the transcript level, Tff1KO mice did not show any significant signs of an altered immune response after infection with T. gondii in comparison with the wild type animals.

Transcriptomic analysis of mouse liver reveals a potential hepato-enteric pathogenic mechanism in acute Toxoplasma gondii infection.

BACKGROUND: Toxoplasma gondii is a worldwide spread pathogen which can infect all tissues of its host. The transcriptomic responses of infected brain and spleen have been reported. However, our knowledge of the global transcriptomic change in infected liver is limited. Additionally, T. gondii infection represents a highly dynamic process involving complex biological responses of the host at many levels. Herein, we describe such processes at a global level by discovering gene expression changes in mouse livers after acute infection with T. gondii ToxoDB#9 strain. RESULTS: Global transcriptomic analysis identified 2,758 differentially expressed transcripts in infected liver, of which 1,356 were significantly downregulated and 1,402 upregulated. GO and KEGG database analyses showed that host immune responses were upregulated, while the metabolic-related processes/pathways were downregulated, especially xenobiotic metabolism, fatty acid metabolism, energy metabolism, and bile biosynthesis and secretion. The metabolism of more than 800 chemical compounds including anti-Toxoplasma prescribed medicines were predicted to be modulated during acute T. gondii infection due to the downregulation of enzymes involved in xenobiotic metabolism. CONCLUSIONS: To the best of our knowledge, this is the first global transcriptomic analysis of mouse liver infected by T. gondii. The present data indicate that during the early stage of liver infection, T. gondii can induce changes in liver xenobiotic metabolism, upregulating inflammatory response and downregulating hepatocellular PPAR signaling pathway, altering host bile biosynthesis and secretion pathway; these changes could enhance host intestinal dysbacteriosis and thus contribute to the pathological changes of both liver and intestine of infected mice. These findings describe the biological changes in infected liver, providing a potential mechanistic pathway that links hepatic and intestinal pathologies to T. gondii infection.

Seroprevalence and genetic characterization of Toxoplasma gondii in masked palm civet (Paguma larvata) in Hainan province, tropical China.

In the present study, the seroprevalence, risk factors and genotyping of Toxoplasma gondii in masked palm civet were investigated in tropical China. A total of 500 serum were collected from five administrative farms in tropical China, and assayed for T. gondii antibodies by modified agglutination test (MAT). The brain samples of 20 aborted fetuses were examined by semi-nested-PCR, and positive aborted fetuses (50%) were necropsied to collect the brain tissue for molecular and bioassay examinations. Genomic DNA was extracted from the 29 brain tissues of infected mice and T. gondii B1 gene was amplified using multilocus PCR-RFLP. Overall, 27.6% (95% CI: 23.682-31.518) of the animals was positive for T. gondii antibodies. Ages of masked palm civet was considered as a main risk factor associated with T. gondii infection. 4 DNA samples (13.8%) were positive for the T. gondii B1 gene. Three samples belong to ToxoDB#9, and one belongs to genotype the type II variant (ToxoDB genotype#3). Our results indicated that ToxoDB Genotype#9 has a distribution in masked palm civet that could be potential reservoirs for T. gondii transmission, which may pose a threat to human health.

Proteomic Profiling of Mouse Liver following Acute Toxoplasma gondii Infection.

Toxoplasma gondii remains a global public health problem. However, its pathophysiology is still not-completely understood particularly the impact of infection on host liver metabolism. We performed iTRAQ-based proteomic analysis to evaluate early liver protein responses in BALB/c mice following infection with T. gondii PYS strain (genotype ToxoDB#9) infection. Our data revealed modification of protein expression in key metabolic pathways, as indicated by the upregulation of immune response and downregulation of mitochondrial respiratory chain, and the metabolism of fatty acids, lipids and xenobiotics. T. gondii seems to hijack host PPAR signaling pathway to downregulate the metabolism of fatty acids, lipids and energy in the liver. The metabolism of over 400 substances was affected by the downregulation of genes involved in xenobiotic metabolism. The top 10 transcription factors used by upregulated genes were Stat2, Stat1, Irf2, Irf1, Sp2, Egr1, Stat3, Klf4, Elf1 and Gabpa, while the top 10 transcription factors of downregulated genes were Hnf4A, Ewsr1, Fli1, Hnf4g, Nr2f1, Pparg, Rxra, Hnf1A, Foxa1 and Foxo1. These findings indicate global reprogramming of the metabolism of the mouse liver after acute T. gondii infection. Functional characterization of the altered proteins may enhance understanding of the host responses to T. gondii infection and lead to the identification of new therapeutic targets.