toxplasma news feeds

A Glycosylphosphatidylinositol-Anchored Carbonic Anhydrase-Related Protein of Toxoplasma gondii Is Important for Rhoptry Biogenesis and Virulence

The Anti-Toxo Blog - 23 May 2017 - 12:43pm
 2017 May 17;2(3). pii: e00027-17. doi: 10.1128/mSphere.00027-17. eCollection 2017 May-Jun.
Chasen NM1,2, Asady B1, Lemgruber L3, Vommaro RC4, Kissinger JC1,5,6, Coppens I7, Moreno SNJ1,8.AbstractCarbonic anhydrase-related proteins (CARPs) have previously been described as catalytically inactive proteins closely related to α-carbonic anhydrases (α-CAs). These CARPs are found in animals (both vertebrates and invertebrates) and viruses as either independent proteins or domains of other proteins. We report here the identification of a new CARP (TgCA_RP) in the unicellular organism Toxoplasma gondii that is related to the recently described η-class CA found in Plasmodium falciparum. TgCA_RP is posttranslationally modified at its C terminus with a glycosylphosphatidylinositol anchor that is important for its localization in intracellular tachyzoites. The protein localizes throughout the rhoptry bulbs of mature tachyzoites and to the outer membrane of nascent rhoptries in dividing tachyzoites, as demonstrated by immunofluorescence and immunoelectron microscopy using specific antibodies. T. gondii mutant tachyzoites lacking TgCA_RP display a growth and invasion phenotype in vitro and have atypical rhoptry morphology. The mutants also exhibit reduced virulence in a mouse model. Our results show that TgCA_RP plays an important role in the biogenesis of rhoptries. IMPORTANCEToxoplasma gondii is an intracellular pathogen that infects humans and animals. The pathogenesis of T. gondii is linked to its lytic cycle, which starts when tachyzoites invade host cells and secrete proteins from specialized organelles. Once inside the host cell, the parasite creates a parasitophorous vacuole (PV) where it divides. Rhoptries are specialized secretory organelles that contain proteins, many of which are secreted during invasion. These proteins have important roles not only during the initial interaction between parasite and host but also in the formation of the PV and in the modification of the host cell. We report here the identification of a new T. gondii carbonic anhydrase-related protein (TgCA_RP), which localizes to rhoptries of mature tachyzoites. TgCA_RP is important for the morphology of rhoptries and for invasion and growth of parasites. TgCA_RP is also critical for parasite virulence. We propose that TgCA_RP plays a role in the biogenesis of rhoptries.KEYWORDS: Toxoplasma gondii; carbonic anhydrase; glycosylphosphatidylinositols; infectivity; organelle structure; rhoptry
PMID:
 
28529974
 
PMCID:
 
PMC5437132
 
DOI:
 
10.1128/mSphere.00027-17
Categories: toxoplasma news feeds

Characterization of Cytosine Methylation and the DNA Methyltransferases of Toxoplasma gondii

The Anti-Toxo Blog - 23 May 2017 - 12:42pm
 2017 Mar 11;13(4):458-470. doi: 10.7150/ijbs.18644. eCollection 2017.
Wei H1, Jiang S1, Chen L1, He C1, Wu S1, Peng H1.AbstractDNA methylation is a key epigenetic modification which confers phenotypic plasticity and adaptation. Cyst-forming strains of Toxoplasma gondii undergo tachyzoite to bradyzoite conversion after initial acute infection of a host, and the reverse conversion may occur in immune-suppressed hosts. The formation of m5C is catalyzed by DNA methyltransferase (DNMT). We identified two functional DNA methyltransferases, TgDNMTa and TgDNMTb, in T. gondii that may mediate DNA methylation. The recombinant proteins showed intrinsic methyltransferase activity; both have higher transcription levels in bradyzoites than that in tachyzoites. We performed genome-wide analysis of DNA methylation in tachyzoites and bradyzoites. The results showed more methylation sites in bradyzoites than that in tachyzoites. The most significantly enriched GO-terms of genes with DNA methylation were associated with basal cellular processes such as energy metabolism and parasite resistance to host immunity. Tachyzoite proliferation in parasitophorous vacuoles (PV) can be inhibited by the DNA methyltransferase inhibitor 5-azacytidine, a chemical analogue of the nucleotide cytosine that can inactivate DNA methyltransferases. These findings provide the first confirmation of DNA methylation in T. gondii.KEYWORDS: 5-azacytidine.; 5-cytosine methyltransferase; DNA methylation; Toxoplasma gondii; bradyzoite; tachyzoite
PMID:
 
28529454
 
PMCID:
 
PMC5436566
 
DOI:
 
10.7150/ijbs.18644
Categories: toxoplasma news feeds

The merozoite-specific protein, TgGRA11B, identified as a component of the Toxoplasma gondii parasitophorous vacuole in a tachyzoite expression model

The Anti-Toxo Blog - 22 May 2017 - 2:27pm
2017 May 16. pii: S0020-7519(17)30130-3. doi: 10.1016/j.ijpara.2017.04.001. [Epub ahead of print]
Ramakrishnan C1, Walker RA2, Eichenberger RM2, Hehl AB2, Smith NC3.
The apicomplexan, Toxoplasma gondii, infects all warm-blooded animals as intermediate hosts but only felids as definitive hosts. Dense granule proteins are critical for the survival of Toxoplasma within host cells but, whilst these proteins have been studied intensively in tachyzoites, little is known about their expression in the coccidian stages in the cat intestine. Transcriptomic profiling indicates that two putative dense granule proteins, TgGRA11A and TgGRA11B, are expressed uniquely in merozoites. Immunofluorescent microscopy of Toxoplasma-infected cat intestine and tachyzoites engineered to express TgGRA11B, reveals that it is a dense granule protein that traffics into the parasitophorous vacuole and its membrane.
Copyright © 2017. Published by Elsevier Ltd.KEYWORDS: Coccidia; Dense granule; Merozoite; Parasitophorous vacuole; TgGRA11B; Toxoplasma gondii
PMID:
28526607
DOI:
10.1016/j.ijpara.2017.04.001
Categories: toxoplasma news feeds

Neurophysiological Changes Induced by Chronic Toxoplasma gondii Infection

The Anti-Toxo Blog - 18 May 2017 - 3:44pm
 2017 May 17;6(2). pii: E19. doi: 10.3390/pathogens6020019.
Tedford E1, McConkey G2.AbstractAlthough the parasite Toxoplasma gondii is one of the most pervasive neurotropic pathogens in the world, the host-parasite interactions during CNS infection and the consequences of neurological infection are just beginning to be unraveled. The chronic stages of infection have been considered dormant, although several studies have found correlations of infection with an array of host behavioral changes. These may facilitate parasite transmission and impact neurological diseases. During infection, in addition to the presence of the parasites within neurons, host-mediated neuroimmune and hormonal responses to infection are also present. T. gondii induces numerous changes to host neurons during infection and globally alters host neurological signaling pathways, as discussed in this review. Understanding the neurophysiological changes in the host brain is imperative to understanding the parasitic mechanisms and to delineate the effects of this single-celled parasite on health and its contribution to neurological disease.KEYWORDS: Toxoplasma gondii; catecholamine; dopamine; glutamatergic; host-parasite interaction; neuroimmune; neurophysiology; testosterone
PMID:
 
28513566
 
DOI:
 
10.3390/pathogens6020019
Categories: toxoplasma news feeds

Toxoplasma gondii and schizophrenia: a review of published RCTs

The Anti-Toxo Blog - 17 May 2017 - 12:33pm
2017 May 15. doi: 10.1007/s00436-017-5478-y. [Epub ahead of print]
Chorlton SD1.
Over the last 60 years, accumulating evidence has suggested that acute, chronic, and maternal Toxoplasma gondii infections predispose to schizophrenia. More recent evidence suggests that chronically infected patients with schizophrenia present with more severe disease. After acute infection, parasites form walled cysts in the brain, leading to lifelong chronic infection and drug resistance to commonly used antiparasitics. Chronic infection is the most studied and closely linked with development and severity of schizophrenia. There are currently four published randomized controlled trials evaluating antiparasitic drugs, specifically azithromycin, trimethoprim, artemisinin, and artemether, in patients with schizophrenia. No trials have demonstrated a change in psychopathology with adjunctive treatment. Published trials have either selected drugs without evidence against chronic infection or used them at doses too low to reduce brain cyst burden. Furthermore, trials have failed to achieve sufficient power or account for confounders such as previous antipsychotic treatment, sex, age, or rhesus status on antiparasitic effect. There are currently no ongoing trials of anti-Toxoplasma therapy in schizophrenia despite ample evidence to justify further testing.KEYWORDS: Chronic; Schizophrenia; Toxoplasma; Treatment
PMID:
28508166
DOI:
10.1007/s00436-017-5478-y
Categories: toxoplasma news feeds

Purification Toxoplasma gondii Tissue Cysts Using Percoll Gradients

The Anti-Toxo Blog - 17 May 2017 - 12:32pm
2017 May 16;45:20C.2.1-20C.2.19. doi: 10.1002/cpmc.30.
Watts EA1,2, Dhara A1, Sinai AP1.
The protozoan parasite Toxoplasma gondii is capable of infecting all warm-blooded animals and humans. Infectious, transmissible forms of the parasite include oocysts produced by the sexual cycle within the definitive feline host and tissue cysts that form Toxoplasma in the central nervous system and muscle during the asexual cycle within all chronically infected warm-blooded hosts. These tissue cysts are populated with slow-growing bradyzoites, which until recently have been thought to be dormant entities in the context of immune sufficiency. Reactivation to active growth during immune suppression is of critical clinical importance. However, little is known about tissue cysts or the bradyzoites they house, as the diversity of tissue cysts cannot be replicated in cell culture systems. This protocol for optimization of tissue cyst purification from the brains of infected mice using Percoll gradients provides an efficient means to recover in vivo-derived tissue cysts that can be applied to imaging, cell biological, biochemical, transcriptomic, and proteomic analyses. © 2017 by John Wiley & Sons, Inc.
Copyright © 2017 John Wiley & Sons, Inc.KEYWORDS: Percoll; Toxoplasma gondii; bradyzoite; tissue cyst
PMID:
28510363
DOI:
10.1002/cpmc.30
Categories: toxoplasma news feeds

The common parasite Toxoplasma gondii induces prostatic inflammation and microglandular hyperplasia in a mouse model

The Anti-Toxo Blog - 13 May 2017 - 5:50pm
2017 May 12. doi: 10.1002/pros.23362. [Epub ahead of print]
Colinot DL1, Garbuz T1,2, Bosland MC3, Wang L1, Rice SE4, Sullivan WJ Jr1,2, Arrizabalaga G1,2, Jerde TJ1,4,5.BACKGROUND: Inflammation is the most prevalent and widespread histological finding in the human prostate, and associates with the development and progression of benign prostatic hyperplasia and prostate cancer. Several factors have been hypothesized to cause inflammation, yet the role each may play in the etiology of prostatic inflammation remains unclear. This study examined the possibility that the common protozoan parasite Toxoplasma gondii induces prostatic inflammation and reactive hyperplasia in a mouse model.
METHODS: Male mice were infected systemically with T. gondii parasites and prostatic inflammation was scored based on severity and focality of infiltrating leukocytes and epithelial hyperplasia. We characterized inflammatory cells with flow cytometry and the resulting epithelial proliferation with bromodeoxyuridine (BrdU) incorporation.
RESULTS: We found that T. gondii infects the mouse prostate within the first 14 days of infection and can establish parasite cysts that persist for at least 60 days. T. gondii infection induces a substantial and chronic inflammatory reaction in the mouse prostate characterized by monocytic and lymphocytic inflammatory infiltrate. T. gondii-induced inflammation results in reactive hyperplasia, involving basal and luminal epithelial proliferation, and the exhibition of proliferative inflammatory microglandular hyperplasia in inflamed mouse prostates.
CONCLUSIONS: This study identifies the common parasite T. gondii as a new trigger of prostatic inflammation, which we used to develop a novel mouse model of prostatic inflammation. This is the first report that T. gondii chronically encysts and induces chronic inflammation within the prostate of any species. Furthermore, T. gondii-induced prostatic inflammation persists and progresses without genetic manipulation in mice, offering a powerful new mouse model for the study of chronic prostatic inflammation and microglandular hyperplasia.
© 2017 Wiley Periodicals, Inc.KEYWORDS: Toxoplasma gondii; hyperplasia; inflammation; parasite; prostate
PMID:
28497488
DOI:
10.1002/pros.23362
Categories: toxoplasma news feeds

Microbial Vertical Transmission during Human Pregnancy

The Anti-Toxo Blog - 13 May 2017 - 5:49pm
2017 May 10;21(5):561-567. doi: 10.1016/j.chom.2017.04.007.
Arora N1, Sadovsky Y2, Dermody TS3, Coyne CB4.
Congenital infections with pathogens such as Zika virus, Toxoplasma gondii, Listeria monocytogenes, Treponema pallidium, parvovirus, HIV, varicella zoster virus, Rubella, Cytomegalovirus, and Herpesviruses are a major cause of morbidity and mortality worldwide. Despite the devastating impact of microbial infections on the developing fetus, relatively little is known about how pathogens associated with congenital disease breach the placental barrier to transit vertically during human pregnancy. In this Review, we focus on transplacental transmission of pathogens during human gestation. We introduce the structure of the human placenta and describe the innate mechanisms by which the placenta restricts microbial access to the intrauterine compartment. Based on current knowledge, we also discuss the potential pathways employed by microorganisms to overcome the placental barrier and prospects for the future.
Copyright © 2017 Elsevier Inc. All rights reserved.KEYWORDS: Cytomegalovirus; Listeria monocytogenes; Toxoplasma gondii; Zika virus; placenta; trophoblast; vertical transmission
PMID:
28494237
DOI:
10.1016/j.chom.2017.04.007
Categories: toxoplasma news feeds

The coccidian parasites Toxoplasma and Neospora dysregulate mammalian lipid droplet biogenesis

The Anti-Toxo Blog - 12 May 2017 - 3:28pm
2017 May 9. pii: jbc.M116.768176. doi: 10.1074/jbc.M116.768176. [Epub ahead of print]
Hu X1, Binns D1, Reese ML2.
Upon infection, the intracellular parasite Toxoplasma gondii co-opts critical functions of its host cell to avoid immune clearance and gain access to nutritional resources. One route by which Toxoplasma co-opts its host cell is through hijacking host organelles, many of which have roles in immunomodulation. Here we demonstrate that Toxoplasma infection results in increased biogenesis of host lipid droplets through rewiring of multiple components of host neutral lipid metabolism. These metabolic changes cause an increased responsiveness of host cells to free fatty acid, leading to a radical increase in the esterification of free fatty acids into triacylglycerol. We identified c-Jun kinase and mammalian target of rapamycin (mTOR) as components of two distinct host signaling pathways that modulate the parasite-induced lipid droplet accumulation. We also found that, unlike many host processes dysregulated during Toxoplasma infection, the induction of lipid droplet generation is conserved not only during infection with genetically diverse Toxoplasma strains, but also with Neospora caninum, which is closely related to Toxoplasma, but has a restricted host range and uses different effector proteins to alter host signaling. Finally, by showing that a Toxoplasma strain deficient in exporting a specific class of effectors is unable to induce lipid droplet accumulation, we demonstrate that the parasite plays an active role in this process. These results indicate that despite their different host ranges, Toxoplasma and Neospora use a conserved mechanism to co-opt these host organelles, which suggests that lipid droplets play a critical role at the coccidian host-pathogen interface.
Copyright © 2017, The American Society for Biochemistry and Molecular Biology.KEYWORDS: Toxoplasma gondii; host-pathogen interaction; lipid droplet; lipid metabolism; parasite
PMID:
28487365
DOI:
10.1074/jbc.M116.768176
Categories: toxoplasma news feeds

The antifungal Aureobasidin A and an analogue are active against the protozoan parasite Toxoplasma gondii but do not inhibit sphingolipid biosynthesis

The Anti-Toxo Blog - 12 May 2017 - 3:27pm

2017 May 10:1-8. doi: 10.1017/S0031182017000506. [Epub ahead of print]
Alqaisi AQI1, Mbekeani AJ1, Llorens MB1, Elhammer AP2, Denny PW1.
Toxoplasma gondii is an obligate intracellular protozoan parasite of the phylum Apicomplexa, and toxoplasmosis is an important disease of both humans and economically important animals. With a limited array of drugs available there is a need to identify new therapeutic compounds. Aureobasidin A (AbA) is an antifungal that targets the essential inositol phosphorylceramide (IPC, sphingolipid) synthase in pathogenic fungi. This natural cyclic depsipeptide also inhibits Toxoplasma proliforation, with the protozoan IPC synthase orthologue proposed as the target. The data presented here show that neither AbA nor an analogue (Compound 20), target the protozoan IPC synthase orthologue or total parasite sphingolipid synthesis. However, further analyses confirm that AbA exhibits significant activity against the proliferative tachyzoite form of Toxoplasma, and Compound 20, whilst effective, has reduced efficacy. This difference was more evident on analyses of the direct effect of these compounds against isolated Toxoplasma, indicating that AbA is rapidly microbicidal. Importantly, the possibility of targeting the encysted, bradyzoite, form of the parasite with AbA and Compound 20 was demonstrated, indicating that this class of compounds may provide the basis for the first effective treatment for chronic toxoplasmosis.KEYWORDS: Toxoplasma ; Aureobasidin A; bradyzoite; sphingolipid biosynthesis
PMID:
28486997
DOI:
10.1017/S0031182017000506
Categories: toxoplasma news feeds

Brains and Brawn: Toxoplasma Infections of the Central Nervous System and Skeletal Muscle

The Anti-Toxo Blog - 10 May 2017 - 11:25am
 2017 May 5. pii: S1471-4922(17)30102-2. doi: 10.1016/j.pt.2017.04.001. [Epub ahead of print]
Wohlfert EA1, Blader IJ2, Wilson EH3.AbstractToxoplasma gondii is a widespread parasitic pathogen that infects over a third of the world's population. Following an acute infection, the parasite can persist within its mammalian host as intraneuronal or intramuscular cysts. Cysts will occasionally reactivate, and - depending on the host's immune status and site of reactivation - encephalitis or myositis can develop. Because these diseases have high levels of morbidity and can be lethal, it is important to understand how Toxoplasma traffics to these tissues, how the immune response controls parasite burden and contributes to tissue damage, and what mechanisms underlie neurological and muscular pathologies that toxoplasmosis patients present with. This review aims to summarize recent important developments addressing these critical topics.Copyright © 2017 Elsevier Ltd. All rights reserved.KEYWORDS: Plasmodium; blood–brain barrier; chronic infections; myositis; parasite; seizures
PMID:
 
28483381
 
DOI:
 
10.1016/j.pt.2017.04.001
Categories: toxoplasma news feeds

Nanomedicine advances in toxoplasmosis: diagnostic, treatment, and vaccine applications

The Anti-Toxo Blog - 10 May 2017 - 11:24am
 2017 May 5. doi: 10.1007/s00436-017-5458-2. [Epub ahead of print]
Assolini JP1, Concato VM1, Gonçalves MD1, Carloto ACM1, Conchon-Costa I1, Pavanelli WR1, Melanda FN1, Costa IN2,3.AbstractToxoplasmosis is an infectious disease caused by the intracellular parasite Toxoplasma gondii that affects about one third of the world's population. The diagnosis of this disease is carried out by parasite isolation and host antibodies detection. However, the diagnosis presents problems in regard to test sensitivity and specificity. Currently, the most effective T. gondii treatment is a combination of pyrimethamine and sulfadiazine, although both drugs are toxic to the host. In addition to the problems that compromise the effective diagnosis and treatment of toxoplasmosis, there are no reports or indications of any vaccine capable of fully protecting against this infection. Nanomaterials, smaller than 1000 nm, are currently being investigated as an alternative tool in the management of T. gondii infection. This article reviews how recent nanotechnology advances indicate the utility of nanomaterials in toxoplasmosis diagnosis, treatment, and vaccine development.KEYWORDS: Infection; Nanomedicine; Nanoparticles; Nanotechnology; Toxoplasma gondii
PMID:
 
28477099
 
DOI:
 
10.1007/s00436-017-5458-2
Categories: toxoplasma news feeds

Acute infection with an avirulent strain of Toxoplasma gondii causes decreasing and atrophy of nitrergic myenteric neurons of rats

The Anti-Toxo Blog - 10 May 2017 - 11:24am
 2017 May 4. pii: S0065-1281(17)30020-X. doi: 10.1016/j.acthis.2017.04.008. [Epub ahead of print]
Sant'Ana DMG1, Gois MB2, Hermes-Uliana C3, Pereira-Severi LS1, Baptista EM4, Mantovani LC4, da Silva AV5, de Almeida Araújo EJ6.AbstractIn the enteric nervous system (ENS), nitrergic neurons produce and use nitric oxide (NO) as an inhibitory motor neurotransmitter in response to parasitic infections, including those caused by Toxoplasma gondii. However, damage to the host caused by NO has been reported by various authors, and the role of NO in protection or cytotoxicity continues to be extensively studied. In this study, nitrergic neurons were investigated in the myenteric plexus of the jejunum and the distal colon of rats infected with 500 oocysts of the M7741 strain of T. gondii. Ten rats were randomly assigned into a control group (CG) and infected group (IG; received 500 sporulated oocysts of T. gondii orally). After 24h, the rats were euthanized, and samples of the jejunum and distal colon were obtained and processed for NADPH-diaphorase histochemical analysis. Quantitative and morphometric analysis of the nitrergic neurons in whole mounts containing the myenteric plexus was performed. There was a numeric reduction of nitrergic neurons per mm2 in both jejunum and distal colon. The remaining nitrergic neurons suffered atrophy in the areas of the cell body and nucleus, which resulted in a decrease in cytoplasm. Thus, we conclude that an avirulent strain of T. gondii in a short time causes neuroplastic changes in the small and large intestine of rats.Copyright © 2017 Elsevier GmbH. All rights reserved.KEYWORDS: Enteric nervous system; NADPH-diaphorase; Neuroplasticity; Nitric oxide; Toxoplasmosis
PMID:
 
28478954
 
DOI:
 
10.1016/j.acthis.2017.04.008
Categories: toxoplasma news feeds

Toxoplasma gondii Infection Induces High Mobility Group Box 1 Released from Mouse Macrophages

The Anti-Toxo Blog - 10 May 2017 - 11:23am
 2017 Apr 24;8:658. doi: 10.3389/fmicb.2017.00658. eCollection 2017.
Wang H1,2, Li M1, Liu J1, Xu J1, Han Q3, Liu Q1.AbstractHigh mobility group box 1 (HMGB1) is abundantly expressed in intracellular engaged DNA binding ability. However, more importantly, it is a weapon against infection through proinflammatory response and immune regulation while released to extracellular. Toxoplasma gondii causes inflammatory pathological changes including ileitis and encephalitis in chronic infection. To investigate whether HMGB1 contributes to the toxoplasmosis lesions, we examined HMGB1 changes during T. gondii infection. The results showed that HMGB1 transcription was down-regulated in the murine macrophage ANA1 cell line and mouse peritoneal macrophages (PMΦs) after T. gondii inoculation, but up-regulated in the IFN-γ treated macrophages and the intraperitoneal exudate cells from the T. gondii infected mice. The content of intracellular HMGB1 are basically consistent with the transcription levels in ANA1 assay, while there were no obvious changes in the mouse PMΦs. Both ANA1 and mouse PMΦs released HMGB1 after parasites infection, and no obvious HMGB1 aggregation in cytoplasm compare to the IFN-γ treatment group. Furthermore, we demonstrated that T. gondiiinvasion led to HMGB1 release, which was dependent on the Caspase 1 activity. These finding should promote to further investigate the functions of extracellular HMGB1 in the toxoplasmosis.KEYWORDS: HMGB1; IFN-γ; Toxoplasma gondii; caspase-1; inflammasome; proinflammatory response
PMID:
 
28484433
 
PMCID:
 
PMC5402041
 
DOI:
 
10.3389/fmicb.2017.00658
Categories: toxoplasma news feeds

A murine vertical transmission model for Toxoplasma gondii oocyst infection and studies on the efficacy of bumped kinase inhibitor (BKI)-1294 and the naphthoquinone buparvaquone against congenital toxoplasmosis

The Anti-Toxo Blog - 10 May 2017 - 11:23am
 2017 May 9. doi: 10.1093/jac/dkx134. [Epub ahead of print]
Müller J1, Aguado-Martínez A1, Ortega-Mora LM2, Moreno-Gonzalo J2, Ferre I2, Hulverson MA3, Choi R3, McCloskey MC3, Barrett LK3, Maly DJ4, Ojo KK3, Van Voorhis W3, Hemphill A1.AbstractOBJECTIVES: Establishment of a mouse model for congenital toxoplasmosis based on oral infection with oocysts from Toxoplasma gondii ME49 and its application for investigating chemotherapeutic options against congenital toxoplasmosis.METHODS: CD1 mice were mated, orally infected with 5, 25, 100, 500 or 2000 oocysts and monitored for clinical signs and survival of dams and pups until 4 weeks post partum . The parasite burden in infected mice was quantified by real-time PCR in lungs, brains and, in the case of surviving pups, also in eyes. Seroconversion was assessed by ELISA. T. gondii cysts in brain were identified by immunofluorescence. In a second experiment, pregnant CD1 mice challenged with 20 oocysts/mouse were treated with buparvaquone or the calcium-dependent protein kinase 1 inhibitor bumped kinase inhibitor (BKI)-1294 and the outcome of infection was analysed.RESULTS: T. gondii DNA was detected in the brain of all infected animals, irrespective of the infection dose. Seroconversion occurred at 3 weeks post-infection. Most pups born to infected dams died within 1 week post partum , but a small fraction survived until the end of the experiment. T. gondii DNA was detected in the brain of all survivors and half of them exhibited ocular infection. Chemotherapy with both compounds led to dramatically increased numbers of surviving pups and reduced cerebral infection. Most efficient were treatments with BKI-1294, with 100% survivors and only 7% brain-positive pups.CONCLUSIONS: BKI-1294 and buparvaquone exert excellent activities against transplacental transmission in pregnant mice.
PMID:
 
28486633
 
DOI:
 
10.1093/jac/dkx134
Categories: toxoplasma news feeds

Calmodulin-like proteins localized to the conoid regulate motility and cell invasion by Toxoplasma gondii

The Anti-Toxo Blog - 8 May 2017 - 4:00pm
2017 May 5;13(5):e1006379. doi: 10.1371/journal.ppat.1006379. [Epub ahead of print]
Long S1, Brown KM1, Drewry LL1, Anthony B1, Phan IQH2, Sibley LD1.
Toxoplasma gondii contains an expanded number of calmodulin (CaM)-like proteins whose functions are poorly understood. Using a combination of CRISPR/Cas9-mediated gene editing and a plant-like auxin-induced degron (AID) system, we examined the roles of three apically localized CaMs. CaM1 and CaM2 were individually dispensable, but loss of both resulted in a synthetic lethal phenotype. CaM3 was refractory to deletion, suggesting it is essential. Consistent with this prediction auxin-induced degradation of CaM3 blocked growth. Phenotypic analysis revealed that all three CaMs contribute to parasite motility, invasion, and egress from host cells, and that they act downstream of microneme and rhoptry secretion. Super-resolution microscopy localized all three CaMs to the conoid where they overlap with myosin H (MyoH), a motor protein that is required for invasion. Biotinylation using BirA fusions with the CaMs labeled a number of apical proteins including MyoH and its light chain MLC7, suggesting they may interact. Consistent with this hypothesis, disruption of MyoH led to degradation of CaM3, or redistribution of CaM1 and CaM2. Collectively, our findings suggest these CaMs may interact with MyoH to control motility and cell invasion.
PMID:
28475612
DOI:
10.1371/journal.ppat.1006379
Categories: toxoplasma news feeds

Plasma Membrane Association by N-Acylation Governs PKG Function in Toxoplasma gondii

The Anti-Toxo Blog - 4 May 2017 - 2:53pm

2017 May 2;8(3). pii: e00375-17. doi: 10.1128/mBio.00375-17.
Brown KM1, Long S1, Sibley LD2.
Cyclic GMP (cGMP)-dependent protein kinase (protein kinase G [PKG]) is essential for microneme secretion, motility, invasion, and egress in apicomplexan parasites, However, the separate roles of two isoforms of the kinase that are expressed by some apicomplexans remain uncertain. Despite having identical regulatory and catalytic domains, PKGI is plasma membrane associated whereas PKGII is cytosolic in Toxoplasma gondii To determine whether these isoforms are functionally distinct or redundant, we developed an auxin-inducible degron (AID) tagging system for conditional protein depletion in T. gondii By combining AID regulation with genome editing strategies, we determined that PKGI is necessary and fully sufficient for PKG-dependent cellular processes. Conversely, PKGII is functionally insufficient and dispensable in the presence of PKGI The difference in functionality mapped to the first 15 residues of PKGI, containing a myristoylated Gly residue at position 2 that is critical for membrane association and PKG function. Collectively, we have identified a novel requirement for cGMP signaling at the plasma membrane and developed a new system for examining essential proteins in T. gondiiIMPORTANCEToxoplasma gondii is an obligate intracellular apicomplexan parasite and important clinical and veterinary pathogen that causes toxoplasmosis. Since apicomplexans can only propagate within host cells, efficient invasion is critically important for their life cycles. Previous studies using chemical genetics demonstrated that cyclic GMP signaling through protein kinase G (PKG)-controlled invasion by apicomplexan parasites. However, these studies did not resolve functional differences between two compartmentalized isoforms of the kinase. Here we developed a conditional protein regulation tool to interrogate PKG isoforms in T. gondii We found that the cytosolic PKG isoform was largely insufficient and dispensable. In contrast, the plasma membrane-associated isoform was necessary and fully sufficient for PKG function. Our studies identify the plasma membrane as a key location for PKG activity and provide a broadly applicable system for examining essential proteins in T. gondii.
Copyright © 2017 Brown et al.
PMID:
28465425
DOI:
10.1128/mBio.00375-17
Categories: toxoplasma news feeds

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

The Anti-Toxo Blog - 3 May 2017 - 12:44pm

2017 May 1;12(5):e0176700. doi: 10.1371/journal.pone.0176700. eCollection 2017.
Milovanović I1, Busarčević M2, Trbovich A1,2, Ivović V2, Uzelac A2, Djurković-Djaković O2.
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.
PMID:
28459857
DOI:
10.1371/journal.pone.0176700
Categories: toxoplasma news feeds

Mechanisms and pathways of Toxoplasma gondii transepithelial migration

The Anti-Toxo Blog - 1 May 2017 - 12:32pm

2017 Jan 2;5(1):e1273865. doi: 10.1080/21688370.2016.1273865. Epub 2016 Dec 20.
Jones EJ1,2, Korcsmaros T1,2, Carding SR1,3.
Toxoplasma gondii is a ubiquitous parasite and a prevalent food-borne parasitic pathogen. Infection of the host occurs principally through oral consumption of contaminated food and water with the gastrointestinal tract being the primary route for entry into the host. To promote infection, T. gondii has evolved highly specialized strategies for rapid traversal of the single cell thick intestinal epithelial barrier. Parasite transmigration via the paracellular pathway between adjacent cells enables parasite dissemination to secondary sites of infection where chronic infection of muscle and brain tissue is established. It has recently been proposed that parasite interactions with the integral tight junction (TJ) protein occludin influences parasite transmigration of the intestinal epithelium. We review here the emerging mechanisms of T. gondii transmigration of the small intestinal epithelium alongside the developing role played in modulating the wider TJ-associated proteome to rewire host cell regulatory systems for the benefit of the parasite.KEYWORDS: Toxoplasma gondii; intestinal epithelial cells; occludin; paracellular transmigration; tight junction
PMID:
28452683
DOI:
10.1080/21688370.2016.1273865
Categories: toxoplasma news feeds

Determination of a Key Antigen for Immunological Intervention To Target the Latent Stage of Toxoplasma gondii

The Anti-Toxo Blog - 28 April 2017 - 12:30pm
2017 Apr 26. pii: 1700062. doi: 10.4049/jimmunol.1700062. [Epub ahead of print]
Sa Q1, Ochiai E1, Tiwari A1, Mullins J1, Shastri N2, Mercier C3, Cesbron-Delauw MF3, Suzuki Y4.
Toxoplasma gondii, an obligate intracellular protozoan parasite, establishes a chronic infection by forming cysts preferentially in the brain. Up to one third of the human population worldwide is estimated to be chronically infected with this parasite. However, there is currently no drug effective against the cyst form of the parasite. In addition, the protective immunity against the cysts remains largely unknown. We analyzed the molecular mechanisms by which the immune system detects host cells harboring the cysts to eliminate the latent stage of the parasite using mice with the H-2d haplotype, which are genetically resistant to the infection. Our study revealed that CD8+ immune T cells bearing TCR Vβ8.1, 8.2 chain have a potent activity to remove T. gondii cysts from the brain. Our studies also uncovered that H-2Ld is the major Ag-presenting molecule to CD8+ T cells for initiating cyst elimination, and that CD8+Vβ8.1, 8.2+ immune T cells recognize the N-terminal region (aa 41-152) of dense granule protein 6 (GRA6Nt) of the parasite presented by the H-2Ld molecule. Furthermore, CD8+ immune T cells induced by immunization with recombinant GRA6Nt were eventually capable of removing the cysts from the brain when transferred to infected immunodeficient mice lacking T cells. Thus, GRA6Nt is a novel and potent Ag to activate CD8+ T cells capable of removing T. gondii cysts. These observations offer a basis for immunological intervention to combat chronic infection with T. gondii by targeting the persistent cysts of the parasite.
Copyright © 2017 by The American Association of Immunologists, Inc.
PMID:
28446567
DOI:
10.4049/jimmunol.1700062
Categories: toxoplasma news feeds

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