Friday, January 20, 2017

Surface attachment, promoted by the actomyosin system of Toxoplasma gondii is important for efficient gliding motility and invasion

2017 Jan 18;15(1):1. doi: 10.1186/s12915-016-0343-5.

BACKGROUND:

Apicomplexan parasites employ a unique form of movement, termed gliding motility, in order to invade the host cell. This movement depends on the parasite's actomyosin system, which is thought to generate the force during gliding. However, recent evidence questions the exact molecular role of this system, since mutants for core components of the gliding machinery, such as parasite actin or subunits of the MyoA-motor complex (the glideosome), remain motile and invasive, albeit at significantly reduced efficiencies. While compensatory mechanisms and unusual polymerisation kinetics of parasite actin have been evoked to explain these findings, the actomyosin system could also play a role distinct from force production during parasite movement.

RESULTS:

In this study, we compared the phenotypes of different mutants for core components of the actomyosin system in Toxoplasma gondii to decipher their exact role during gliding motility and invasion. We found that, while some phenotypes (apicoplast segregation, host cell egress, dense granule motility) appeared early after induction of the act1 knockout and went to completion, a small percentage of the parasites remained capable of motility and invasion well past the point at which actin levels were undetectable. Those act1 conditional knockout (cKO) and mlc1 cKO that continue to move in 3D do so at speeds similar to wildtype parasites. However, these mutants are virtually unable to attach to a collagen-coated substrate under flow conditions, indicating an important role for the actomyosin system of T. gondii in the formation of attachment sites.

CONCLUSION:

We demonstrate that parasite actin is essential during the lytic cycle and cannot be compensated by other molecules. Our data suggest a conventional polymerisation mechanism in vivo that depends on a critical concentration of G-actin. Importantly, we demonstrate that the actomyosin system of the parasite functions in attachment to the surface substrate, and not necessarily as force generator.

KEYWORDS:

Actin; Apicomplexa; Attachment; Host cell invasion; Membrane flow; Motility; Myosin; Toxoplasma
PMID:
28100223
DOI:
10.1186/s12915-016-0343-5

Thursday, January 19, 2017

The Effect of Fluphenazine and Thioridazine on Toxoplasma gondii In Vivo

 2016 Apr-Jun;11(2):226-231.

Abstract

BACKGROUND:

Toxoplasma gondii is the most common parasite causing latent cerebral infections in human. It has been shown that some anti-psychotic drugs are able to inhibit the proliferation of the parasite in in vitro study. There is very limited data regarding the inhibitory effect of anti-psychotics on Toxoplasma in in vivo. In this study, we evaluated anti-Toxoplasma activity of fluphenazine and thioridazine drugs on T. gondii in mice.

METHODS:

Mice were divided into six groups: Control, sesame as vehicle, thioridazine 10 mg/kg, thioridazine 20 mg/kg, fluphenazine 0.06 mg/kg and fluphenazine 0.6 mg/kg. They were inoculated intraperitoneally with brain suspension containing tissue cysts of T. gondii Tehran strain. Two months after inoculation, the number of cysts in crushed smears of mice brain were counted microscopically and considered as an indicator of anti-Toxoplasma activity. This work has conducted in Qazvin, central Iran, 2014.

RESULTS:

Our study showed that fluphenazine and thioridazine could not significantly inhibit the brain cystogenesis of T. gondii in mice. However, the number of brain cysts was less at higher dose compared to lower doses for both drugs.

CONCLUSION:

Further studies need to clear the mechanism of different structure of anti-psychotic drugs on activity of Toxoplasma.

KEYWORDS:

Brain cysts; Fluphenazine; In vivo; Thioridazine; Toxoplama gondii
PMID:
 
28096857

Acetylcholinesterase Inhibitor Improves Learning and Memory Impairment Induced by Toxoplasma gondii Infection

 2016 Apr-Jun;11(2):177-185.

Abstract

BACKGROUND:

Here, we established the mouse models of chronic toxoplasmosis by T. gondii Tehran strain to provide a good understanding about defining the possible association between T. gondii exposure and learning and memory impairments. Moreover, as secondary objective of the present study, we hypothesized whether administration of an acetylcholinesterase (AChE) inhibitor could reduce learning and memory impairments induced by T. gondii infection.

METHODS:

Twenty-four male BALB/c mice were used to establishment of latent toxoplasmosis. The animal model of Toxoplasma infection was established by the intraperitoneal inoculation of 20-25 tissue cysts from Tehran strain of T. gondii. Donepezil (2 mg/kg) an AChE inhibitor to treat Alzheimer disease was injected intraperitoneally once a day for two weeks starting from post-infection day 90. Morris water maze (MWM) task was used to assay spatial learning and short term spatial memory in all groups. One-way ANOVA with Tukey's post-hoc test was used to assess differences between experimental groups. P0.05 was considered statistically significant.

RESULTS:

Toxoplasma infection impaired spatial leaning and short term spatial memory of the infected BALB/c mice, whereas donepezil, an AChE inhibitor, improved impairments induced by Toxoplasma infection.

CONCLUSION:

T. gondii infection through increasing AChE reduces the level of Acetylcholine (Ach) and consequently affects learning and memory activity in infected hosts, whereas, donepezil as an AChE inhibitor improves these impairments by restoring ACh levels at synapses of neurons in brain.

KEYWORDS:

Donepezil; Maze learning; Memory; Spatial learning; Toxoplasma gondii; Toxoplasmosis
PMID:
 
28096851

Assessment of the Effects of Extremely Low Frequency Electromagnetic Fields on Toxoplasma gondii

 2016 Apr-Jun;11(2):159-167.

Abstract

BACKGROUND:

The effects of extremely low frequency electromagnetic fields (ELF-EMF) on Toxoplasma gondii have not been explained yet. The aim of this study was to assess the possible effects of ELF-EMF on growth, survival time and viability of Toxoplasma gondii. In addition, the life span of Toxoplasma infected animals was investigated.

METHODS:

Sixty adult male BALB/c mice were used for in vivo and in vivo experiments in Laboratory of Biopyhsics and Parasitology of Medical Faculty, Adnan Menderes University, Turkey, in 2010. During in vivo experiments, pulsed and continuous EMFs were applied for 5 d to the infected mice. During in vivo experiments, pulsed and continuous EMF was applied to the tachyzoites within peritoneal exudates for 8 h/d at 4 °C and the tachyzoites were then injected to mice. In both experiments, the number of T. gondii in peritoneal exudates was counted and T. gondii protein bands patterns were investigated with polyacrylamide gel electrophoresis and Western Blotting.

RESULTS:

Pulsed and continuous EMF exposure reduced the number of T. gondii tachyzoites in comparison to controls. However, no statistically significant differences were observed at the patterns of protein bands among the samples.

CONCLUSION:

EMF exposure induces a decrease in the number of T. gondii. Further studies are required to understand the mechanism of EMF on intracellular parasites.

KEYWORDS:

Electromagnetic field; Mice; Polyacrylamide gel electrophoresis; Toxoplasma gondii; Western Blotting
PMID:
 
28096849

Production of IL-1β and Inflammasome with Up-Regulated Expressions of NOD-Like Receptor Related Genes in Toxoplasma gondii-Infected THP-1 Macrophages

 2016 Dec;54(6):711-717. doi: 10.3347/kjp.2016.54.6.711. Epub 2016 Dec 31.

Abstract

Toxoplasma gondii is an obligate intracellular parasite that stimulates production of high levels of proinflammatory cytokines, which are important for innate immunity. NLRs, i.e., nucleotide-binding oligomerization domain (NOD)-like receptors, play a crucial role as innate immune sensors and form multiprotein complexes called inflammasomes, which mediate caspase-1-dependent processing of pro-IL-1β. To elucidate the role of inflammasome components in T. gondii-infected THP-1 macrophages, we examined inflammasome-related gene expression and mechanisms of inflammasome-regulated cytokine IL-1β secretion. The results revealed a significant upregulation of IL-1β after T. gondii infection. T. gondii infection also upregulated the expression of inflammasome sensors, including NLRP1, NLRP3, NLRC4, NLRP6, NLRP8, NLRP13, AIM2, and NAIP, in a time-dependent manner. The infection also upregulated inflammasome adaptor protein ASC and caspase-1 mRNA levels. From this study, we newly found that T. gondii infection regulates NLRC4, NLRP6, NLRP8, NLRP13, AIM2, and neuronal apoptosis inhibitor protein (NAIP) gene expressions in THP-1 macrophages and that the role of the inflammasome-related genes may be critical for mediating the innate immune responses to T. gondii infection.

KEYWORDS:

THP-1 macrophage; Toxoplasma gondii; inflammasome; nucleotide-binding oligomerization domain (NOD)-like receptor (NLR); qRT-PCR
PMID:
 
28095655
 
DOI:
 
10.3347/kjp.2016.54.6.711

Tuesday, January 17, 2017

CCAAT/Enhancer-Binding Protein β Mediates the Killing of Toxoplasma gondii by Inducing Autophagy in Nonhematopoietic Cells


2017 Jan 16. doi: 10.1089/dna.2016.3434. [Epub ahead of print]


Autophagy is a main defense strategy by which infected host cells can virtually induce the killing of parasite, including Toxoplasma gondii. However, the regulatory mechanisms of autophagy in T. gondii-infected nonhematopoietic cells are still unknown. Emerging evidence indicates that CCAAT/enhancer-binding protein β (C/EBP β) is associated with the regulation of autophagy. Herein, we hypothesized that C/EBP β plays roles in inducing autophagy in nonhematopoietic cells. Expression of C/EBP β was aberrantly regulated in endothelial cells and retinal pigment epithelial cells challenged by T. gondii. Inhibition of C/EBP β reduced the killing of T. gondii in nonhematopoietic cells, whereas C/EBP β overexpression resulted in the enhancement of killing of T. gondii as well as the increase in autophagy in infected cells. Furthermore, the mammalian target of rapamycin (mTOR) activation was found to be reduced by C/EBP β overexpression, but increased by C/EBP β inhibition. The increase in T. gondii killing induced by C/EBP β overexpression was blocked by the mTOR activator phosphatidic acid and was increased by the inhibitor AZD8055. In conclusion, we demonstrate that C/EBP β expression is increased in nonhematopoietic cells infected by T. gondii, resulting in the activation of autophagy in host cells by inhibiting mTOR pathway.

KEYWORDS:

C/EBP β; T. gondii; autophagy; killing; mTOR; nonhematopoietic cells
PMID:
28092463
DOI:
10.1089/dna.2016.3434

Monday, January 16, 2017

Transcriptional repression by ApiAP2 factors is central to chronic toxoplasmosis

Joshua B Radke, Danielle Worth, Dong-Pyo Hong, Sherri Huang, William J Sullivan Jr, Emma H Wilson, Michael White

doi: https://doi.org/10.1101/100628

Abstract

Bradyzoite differentiation is marked by major changes in gene expression resulting in a parasite that expresses a new repertoire of surface antigens hidden inside a modified parasitophorous vacuole called the tissue cyst. The factors that control this important life cycle transition are not well understood. Here we describe an important Toxoplasma repressor mechanism controlling bradyzoite differentiation that operates exclusively in the tachyzoite stage. The ApiAP2 factor, AP2IV-4, is a nuclear factor dynamically expressed in late S phase through mitosis/cytokinesis of the tachyzoite cell cycle and absent from the bradyzoite. Remarkably, deletion of the AP2IV-4 locus resulted in the increased expression of bradyzoite mRNAs in replicating tachyzoites and in two different genetic lineages, we confirmed the misexpression of tissue cyst wall components (e.g. BPK1, MCP4, CST1) and the bradyzoite surface antigen SRS9 in the tachyzoite stage. In the murine animal model, the loss of AP2IV-4 had profound biological consequences. Prugniaud strain parasites lacking AP2IV-4 were unable to form tissue cysts in brain tissue and the absence of this factor also recruited a potent immune response characterized by increases inflammatory monocytes, IFN-γ and higher numbers of both CD8+ and CD4+ T-cells. Altogether these results indicate that suppression of bradyzoite antigens by AP2IV-4 during acute infection is required for Toxoplasma to establish a chronic infection in the immune-competent host.

A human proteome array approach to identifying key host proteins targeted by Toxoplasma kinase ROP18

 2017 Jan 13. pii: mcp.M116.063602. doi: 10.1074/mcp.M116.063602. [Epub ahead of print]

Yang Z1Hou Y1Hao T1Rho HS2Wan J2Luan Y1Gao X1Yao J1Pan A1Xie Z1Qian J2Liao W1Zhu H2Zhou X3.

Abstract

Toxoplasma kinase ROP18 is a key molecule responsible for the virulence of Toxoplasma gondii; however, the mechanisms by which ROP18 exerts parasite virulence via interaction with host proteins remain limited to a small number of identified substrates. To identify a broader array of ROP18 substrates we successfully purified bioactive mature ROP18 and used it to probe a human proteome array. Sixty-eight new putative host targets were identified. Functional annotation analysis suggested that these proteins have a variety of functions including metabolic process, kinase activity and phosphorylation, cell growth, apoptosis and cell death, and immunity, indicating a pleiotropic role of ROP18 kinase. Among these proteins, four candidates, p53, p38, UBE2N, and Smad1 were further validated. We demonstrated that ROP18 targets p53, p38, UBE2N, and Smad1 for degradation. Importantly, we demonstrated that ROP18 phosphorylates Smad1 Ser187 to trigger its proteasome-dependent degradation. Further functional characterization of the substrates of ROP18 may enhance understanding of the pathogenesis of Toxoplasma infection and providing new therapeutic targets. Similar strategies could be used to identify novel host targets for other microbial kinases functioning at the pathogen-host interface.

KEYWORDS: 

Kinases*; Microbiology; Pathogens; Protein array; Protein-Protein Interactions*; host-pathogen interaction; proteome array
PMID:
 
28087594
 
DOI:
 
10.1074/mcp.M116.063602

Toxoplasmosis-associated IRIS involving the CNS: a case report with longitudinal analysis of T cell subsets

 2017 Jan 13;17(1):66. doi: 10.1186/s12879-016-2159-x.

Abstract

BACKGROUND: 

HIV-infected patients may present an unforeseen clinical worsening after initiating antiretroviral therapy known as immune reconstitution inflammatory syndrome (IRIS). This syndrome is characterized by a heightened inflammatory response toward infectious or non-infectious triggers, and it may affect different organs. Diagnosis of IRIS involving the central nervous system (CNS-IRIS) is challenging due to heterogeneous manifestations, absence of biomarkers to identify this condition, risk of long-term sequelae and high mortality. Hence, a deeper knowledge of CNS-IRIS pathogenesis is needed.

CASE PRESENTATION: 

A 37-year-old man was diagnosed with AIDS and cerebral toxoplasmosis. Anti-toxoplasma treatment was initiated immediately, followed by active antiretroviral therapy (HAART) 1 month later. At 2 months of HAART, he presented with progressive hyposensitivity of the right lower limb associated with brain and dorsal spinal cord lesions, compatible with paradoxical toxoplasmosis-associated CNS-IRIS, a condition with very few reported cases. A stereotactic biopsy was planned but was postponed based on its inherent risks. Patient showed clinical improvement with no requirement of corticosteroid therapy. Routine laboratorial analysis was complemented with longitudinal evaluation of blood T cell subsets at 0, 1, 2, 3 and 6 months upon HAART initiation. A control group composed by 9 HIV-infected patients from the same hospital but with no IRIS was analysed for comparison. The CNS-IRIS patient showed lower percentage of memory CD4+ T cells and higher percentage of activated CD4+ T cells at HAART initiation. The percentage of memory CD4+ T cells drastically increased at 1 month after HAART initiation and became higher in comparison to the control group until clinical recovery onset; the percentage of memory CD8+ T cells was consistently lower throughout follow-up. Interestingly, the percentage of regulatory T cells (Treg) on the CNS-IRIS patient reached a minimum around 1 month before symptoms onset.

CONCLUSION: 

Although both stereotactic biopsies and steroid therapy might be of use in CNS-IRIS cases and should be considered for these patients, they might be unnecessary to achieve clinical improvement as shown in this case. Immunological characterization of more CNS-IRIS cases is essential to shed some light on the pathogenesis of this condition.

KEYWORDS: 

Human immunodeficiency virus; Immune reconstitution inflammatory syndrome; Regulatory T cells; T cell subsets; Toxoplasmosis
PMID:
 
28086758
 
DOI:
 
10.1186/s12879-016-2159-x

A critical assessment of the association between postnatal toxoplasmosis and epilepsy in immune-competent patients

 2017 Jan 12. doi: 10.1007/s10096-016-2897-0. [Epub ahead of print]

Abstract

While postnatal toxoplasmosis in immune-competent patients is generally considered a self-limiting and mild illness, it has been associated with a variety of more severe clinical manifestations. The causal relation with some manifestations, e.g. myocarditis, has been microbiologically proven, but this is not unequivocally so for other reported associations, such as with epilepsy. We aimed to systematically assess causality between postnatal toxoplasmosis and epilepsy in immune-competent patients. A literature search was performed. The Bradford Hill criteria for causality were used to score selected articles for each component of causality. Using an arbitrary but defined scoring system, the maximal score was 15 points (13 for case reports). Of 704 articles, five case reports or series and five case-control studies were selected. The strongest evidence for a causal relation was provided by two case reports and one case-control study, with a maximal causality score of, respectively, 9/13, 10/13 and 10/15. The remaining studies had a median causality score of 7 (range 5-9). No selection bias was identified, but 6/10 studies contained potential confounders (it was unsure whether the infection was pre- or postnatal acquired, or immunodeficiency was not specifically excluded). Based on the evaluation of the available literature, although scanty and of limited quality, a causal relationship between postnatal toxoplasmosis and epilepsy seems possible. More definite proof requires further research, e.g. by performing Toxoplasma serology in all de novo epilepsy cases.
PMID:
 
28083719
 
DOI:
 
10.1007/s10096-016-2897-0

Friday, January 13, 2017

Bromodomains in Protozoan Parasites: Evolution, Function, and Opportunities for Drug Development

2017 Jan 11;81(1). pii: e00047-16. doi: 10.1128/MMBR.00047-16. Print 2017 Mar.


Parasitic infections remain one of the most pressing global health concerns of our day, affecting billions of people and producing unsustainable economic burdens. The rise of drug-resistant parasites has created an urgent need to study their biology in hopes of uncovering new potential drug targets. It has been established that disrupting gene expression by interfering with lysine acetylation is detrimental to survival of apicomplexan (Toxoplasma gondii and Plasmodium spp.) and kinetoplastid (Leishmania spp. and Trypanosoma spp.) parasites. As "readers" of lysine acetylation, bromodomain proteins have emerged as key gene expression regulators and a promising new class of drug target. Here we review recent studies that demonstrate the essential roles played by bromodomain-containing proteins in parasite viability, invasion, and stage switching and present work showing the efficacy of bromodomain inhibitors as novel antiparasitic agents. In addition, we performed a phylogenetic analysis of bromodomain proteins in representative pathogens, some of which possess unique features that may be specific to parasite processes and useful in future drug development.

KEYWORDS:

Leishmania; Plasmodium; Toxoplasma; epigenetics; malaria; trypanosomes
PMID:
28077462
DOI:
10.1128/MMBR.00047-16

Modulation of host cell SUMOylation facilitates efficient development of Plasmodium berghei and Toxoplasma gondii

2017 Jan 12. doi: 10.1111/cmi.12723. [Epub ahead of print]


SUMOylation is a reversible post translational modification of proteins that regulates protein stabilization, nucleocytoplasmic transport and protein-protein interactions. Several viruses and bacteria modulate host SUMOylation machinery for efficient infection. Plasmodium sporozoites are infective forms of malaria parasite that infect hepatocytes and transforms into exoerythrocytic forms (EEFs). Here we show that, during EEF development, the distribution of SUMOylated proteins in host cell nuclei was significantly reduced and expression of the SUMOylation enzymes was downregulated. Plasmodium EEFs destabilized the host cytoplasmic protein SMAD4 by inhibiting its SUMOylation. SUMO1 overexpression was detrimental to EEF growth and insufficiency of the only conjugating enzyme Ubc9/E2 promoted EEF growth. The expression of genes involved in suppression of host cell defense pathways during infection was reversed during SUMO1 overexpression, as revealed by transcriptomic analysis. The inhibition of host cell SUMOylation was also observed in Toxoplasma. We provide a hitherto unknown mechanism of regulating host gene expression through altering host SUMOylation by Apicomplexan parasites.
PMID:
28078755
DOI:
10.1111/cmi.12723

Thursday, January 12, 2017

Is Toxoplasma gondii a Trigger of Bipolar Disorder?

 2017 Jan 10;6(1). pii: E3. doi: 10.3390/pathogens6010003.

Abstract

Toxoplasma gondii, a ubiquitous intracellular parasite, has a strong tropism for the brain tissue, where it forms intracellular cysts within the neurons and glial cells, establishing a chronic infection. Although latent toxoplasmosis is generally assumed to be asymptomatic in immunocompetent individuals, it is now clear that it can induce behavioral manipulations in mice and infected humans. Moreover, a strong relation has emerged in recent years between toxoplasmosis and psychiatric disorders. The link between T. gondii and schizophrenia has been the most widely documented; however, a significant association with bipolar disorder (BD) and suicidal/aggressive behaviors has also been detected. T. gondii may play a role in the etiopathogenesis of psychiatric disorders affecting neurotransmitters, especially dopamine, that are implicated in the emergence of psychosis and behavioral Toxoplasma-induced abnormalities, and inducing brain inflammation by the direct stimulation of inflammatory cytokines in the central nervous system. Besides this, there is increasing evidence for a prominent role of immune dysregulation in psychosis and BD. The aim of this review is to describe recent evidence suggesting a link between Toxoplasma gondii and BD, focusing on the interaction between immune responses and this infectious agent in the etiopathogenesis of psychiatric symptoms.

KEYWORDS: 

Toxoplasma gondii; antipsychotics and mood stabilizers; bipolar disorder; dopaminergic pathways; immunity; pro-inflammatory cytokines
PMID:
 
28075410
 
DOI:
 
10.3390/pathogens6010003