Saturday, March 31, 2012

Toxoplasma gondii and the blood-brain barrier

Virulence. 2012 Mar 1;3(2). [Epub ahead of print]

Toxoplasma gondii and the blood-brain barrier.

Feustel S, Meissner M, Liesenfeld O.

SourceInstitute for Microbiology and Hygiene; Charité Medical School; Berlin, Germany.

Infection with the protozoan parasite Toxoplasma gondii is characterized by asymptomatic latent infection in the central nervous system and skeletal muscle tissue in the majority of immunocompentent individuals. Life-threatening reactivation of the infection in immunocompromized patients originates from rupture of Toxoplasma cysts in the brain. While major progress has been made in our understanding of the immunopathogenesis of infection the mechanism(s) of neuroinvasion of the parasite remains poorly understood. The present review presents the current understanding of blood-brain barrier (patho)physiology and the interaction of Toxoplasma gondii with cells of the blood-brain barrier.

PMID: 22460645 [PubMed - as supplied by publisher]

Passage of parasites across the blood-brain barrier

Virulence. 2012 Mar 1;3(2). [Epub ahead of print]

Passage of parasites across the blood-brain barrier.

Masocha W, Kristensson K

SourceDepartment of Applied Therapeutics; Faculty of Pharmacy; Kuwait University; Kuwait City, Kuwait.

The blood-brain barrier (BBB) is a structural and functional barrier that protects the central nervous system (CNS) from invasion by blood-borne pathogens including parasites. However, some intracellular and extracellular parasites can traverse the BBB during the course of infection and cause neurological disturbances and/or damage which are at times fatal. The means by which parasites cross the BBB and how the immune system controls the parasites within the brain are still unclear. In this review we present the current understanding of the processes utilized by two human neuropathogenic parasites, Trypanosoma brucei spp and Toxoplasma gondii, to go across the BBB and consequences of CNS invasion. We also describe briefly other parasites that can invade the brain and how they interact with or circumvent the BBB. The roles played by parasite-derived and host-derived molecules during parasitic and white blood cell invasion of the brain are discussed.

PMID: 22460639 [PubMed - as supplied by publisher]

Friday, March 30, 2012

Comparative Genomics of the Apicomplexan Parasites Toxoplasma gondii and Neospora caninum: Coccidia Differing in Host Range and Transmission Strategy

PLoS Pathog. 2012 Mar;8(3):e1002567. Epub 2012 Mar 22

Comparative Genomics of the Apicomplexan Parasites Toxoplasma gondii and Neospora caninum: Coccidia Differing in Host Range and Transmission Strategy.

Reid AJ, Vermont SJ, Cotton JA, Harris D, Hill-Cawthorne GA, Könen-Waisman S, Latham SM, Mourier T, Norton R, Quail MA, Sanders M, Shanmugam D, Sohal A, Wasmuth JD, Brunk B, Grigg ME, Howard JC, Parkinson J, Roos DS, Trees AJ, Berriman M, Pain A, Wastling JM.

SourceWellcome Trust Sanger Institute, Hinxton, Cambridgshire, United Kingdom.

Toxoplasma gondii is a zoonotic protozoan parasite which infects nearly one third of the human population and is found in an extraordinary range of vertebrate hosts. Its epidemiology depends heavily on horizontal transmission, especially between rodents and its definitive host, the cat. Neospora caninum is a recently discovered close relative of Toxoplasma, whose definitive host is the dog. Both species are tissue-dwelling Coccidia and members of the phylum Apicomplexa; they share many common features, but Neospora neither infects humans nor shares the same wide host range as Toxoplasma, rather it shows a striking preference for highly efficient vertical transmission in cattle. These species therefore provide a remarkable opportunity to investigate mechanisms of host restriction, transmission strategies, virulence and zoonotic potential. We sequenced the genome of N. caninum and transcriptomes of the invasive stage of both species, undertaking an extensive comparative genomics and transcriptomics analysis. We estimate that these organisms diverged from their common ancestor around 28 million years ago and find that both genomes and gene expression are remarkably conserved. However, in N. caninum we identified an unexpected expansion of surface antigen gene families and the divergence of secreted virulence factors, including rhoptry kinases. Specifically we show that the rhoptry kinase ROP18 is pseudogenised in N. caninum and that, as a possible consequence, Neospora is unable to phosphorylate host immunity-related GTPases, as Toxoplasma does. This defense strategy is thought to be key to virulence in Toxoplasma. We conclude that the ecological niches occupied by these species are influenced by a relatively small number of gene products which operate at the host-parasite interface and that the dominance of vertical transmission in N. caninum may be associated with the evolution of reduced virulence in this species.

PMID: 22457617 [PubMed - as supplied by publisher]

Thursday, March 29, 2012

Conditional mutagenesis of a novel choline kinase demonstrates the plasticity of phosphatidylcholine biogenesis and gene expression in Toxoplasma

J Biol Chem. 2012 Mar 26. [Epub ahead of print]

Conditional mutagenesis of a novel choline kinase demonstrates the plasticity of phosphatidylcholine biogenesis and gene expression in Toxoplasma gondii.

Sampels V, Hartmann A, Dietrich I, Coppens I, Sheiner L, Striepen B, Herrmann A, Lucius R, Gupta N.

SourceHumboldt University, Germany;

The obligate intracellular and promiscuous protozoan parasite Toxoplasma gondii needs an extensive membrane biogenesis that must be satisfied irrespective of its host-cell milieu. We show that the synthesis of the major lipid in T. gondii, phosphatidylcholine (PtdCho), is initiated by a choline kinase (TgCK, ~70-kDa). Full-length TgCK displayed a low affinity for choline (Km ~0.77 mM), and harbors a unique N-terminal hydrophobic peptide that is required for the formation of enzyme oligomers in the parasite cytosol but not for activity. Conditional mutagenesis of the TgCK gene in T. gondii attenuated the protein level by ~60%, which was abolished in the off state of the mutant (Δtgcki). Unexpectedly, the mutant was not impaired in its growth, and also exhibited a normal PtdCho biogenesis. The parasite circumvented the loss of full-length TgCK by a cryptic promoter identified within exon 1 sequence, which can express two putative 53-kDa and 44-kDa isoforms. TgCK-Exon1 alone was also sufficient in driving the expression of GFP in E. coli. The presence of a cryptic promoter correlated with the persistent enzyme activity, PtdCho synthesis and susceptibility of T. gondii to a choline analog, dimethylethanolamine. Quite notably, the mutant displayed a regular growth in the off state despite a 35% decline in PtdCho content and lipid synthesis suggesting a compositional flexibility in the parasite membranes. The observed plasticity of gene expression and membrane biogenesis can ensure a faithful replication and adaptation of T. gondii in disparate host or nutrient environments.

PMID: 22451671 [PubMed - as supplied by publisher]

Wednesday, March 21, 2012

Chronic toxoplasma infection modifies the structure and the risk of host behavior

PLoS One. 2012;7(3):e32489. Epub 2012 Mar 14.

Chronic toxoplasma infection modifies the structure and the risk of host behavior.

Afonso C, Paixão VB, Costa RM.

Champalimaud Neuroscience Programme, Instituto Gulbenkian de Ciência, Oeiras, Portugal.

The intracellular parasite Toxoplasma has an indirect life cycle, in which felids are the definitive host. It has been suggested that this parasite developed mechanisms for enhancing its transmission rate to felids by inducing behavioral modifications in the intermediate rodent host. For example, Toxoplasma-infected rodents display a reduction in the innate fear of predator odor. However, animals with Toxoplasma infection acquired in the wild are more often caught in traps, suggesting that there are manipulations of intermediate host behavior beyond those that increase predation by felids. We investigated the behavioral modifications of Toxoplasma-infected mice in environments with exposed versus non-exposed areas, and found that chronically infected mice with brain cysts display a plethora of behavioral alterations. Using principal component analysis, we discovered that most of the behavioral differences observed in cyst-containing animals reflected changes in the microstructure of exploratory behavior and risk/unconditioned fear. We next examined whether these behavioral changes were related to the presence and distribution of parasitic cysts in the brain of chronically infected mice. We found no strong cyst tropism for any particular brain area but found that the distribution of Toxoplasma cysts in the brain of infected animals was not random, and that particular combinations of cyst localizations changed risk/unconditioned fear in the host. These results suggest that brain cysts in animals chronically infected with Toxoplasma alter the fine structure of exploratory behavior and risk/unconditioned fear, which may result in greater capture probability of infected rodents. These data also raise the possibility that selective pressures acted on Toxoplasma to broaden its transmission between intermediate predator hosts, in addition to felid definitive hosts.

PMID: 22431975 [PubMed - in process]

Globally diverse Toxoplasma gondii isolates comprise six major clades originating from a small number of distinct ancestral lineages

Proc Natl Acad Sci U S A. 2012 Mar 19. [Epub ahead of print]

Globally diverse Toxoplasma gondii isolates comprise six major clades originating from a small number of distinct ancestral lineages.

Su C, Khan A, Zhou P, Majumdar D, Ajzenberg D, Dardé ML, Zhu XQ, Ajioka JW, Rosenthal BM, Dubey JP, Sibley LD.

Department of Microbiology, University of Tennessee, Knoxville, TN 37996.

Marked phenotypic variation characterizes isolates of Toxoplasma gondii, a ubiquitous zoonotic parasite that serves as an important experimental model for studying apicomplexan parasites. Progress in identifying the heritable basis for clinically and epidemiologically significant differences requires a robust system for describing and interpreting evolutionary subdivisions in this prevalent pathogen. To develop such a system, we have examined more than 950 isolates collected from around the world and genotyped them using three independent sets of polymorphic DNA markers, sampling 30 loci distributed across all nuclear chromosomes as well as the plastid genome. Our studies reveal a biphasic pattern consisting of regions in the Northern Hemisphere where a few, highly clonal and abundant lineages predominate; elsewhere, and especially in portions of South America are characterized by a diverse assemblage of less common genotypes that show greater evidence of recombination. Clustering methods were used to organize the marked genetic diversity of 138 unique genotypes into 15 haplogroups that collectively define six major clades. Analysis of gene flow indicates that a small number of ancestral lineages gave rise to the existing diversity through a process of limited admixture. Identification of reference strains for these major groups should facilitate future studies on comparative genomics and identification of genes that control important biological phenotypes including pathogenesis and transmission.

PMID: 22431627 [PubMed - as supplied by publisher]

Tuesday, March 20, 2012

Toxoplasma gondii Chromodomain Protein 1 Binds to Heterochromatin and Colocalises with Centromeres and Telomeres at the Nuclear Periphery

PLoS One. 2012;7(3):e32671. Epub 2012 Mar 9.

Toxoplasma gondii Chromodomain Protein 1 Binds to Heterochromatin and Colocalises with Centromeres and Telomeres at the Nuclear Periphery.

Gissot M, Walker R, Delhaye S, Huot L, Hot D, Tomavo S.

Center for Infection and Immunity of Lillle, CNRS UMR 8204, INSERM U 1019, Université Lille Nord de France, Institut Pasteur de Lille, Lille, France.


Apicomplexan parasites are responsible for some of the most deadly parasitic diseases afflicting humans, including malaria and toxoplasmosis. These obligate intracellular parasites exhibit a complex life cycle and a coordinated cell cycle-dependant expression program. Their cell division is a coordinated multistep process. How this complex mechanism is organised remains poorly understood.

In this study, we provide evidence for a link between heterochromatin, cell division and the compartmentalisation of the nucleus in Toxoplasma gondii. We characterised a T. gondii chromodomain containing protein (named TgChromo1) that specifically binds to heterochromatin. Using ChIP-on-chip on a genome-wide scale, we report TgChromo1 enrichment at the peri-centromeric chromatin. In addition, we demonstrate that TgChromo1 is cell-cycle regulated and co-localised with markers of the centrocone. Through the loci-specific FISH technique for T. gondii, we confirmed that TgChromo1 occupies the same nuclear localisation as the peri-centromeric sequences.

We propose that TgChromo1 may play a role in the sequestration of chromosomes at the nuclear periphery and in the process of T. gondii cell division.

PMID: 22427862 [PubMed - in process]

Identification of a New Rhoptry Neck Complex RON9/RON10 in the Apicomplexa Parasite Toxoplasma gondii

PLoS One. 2012;7(3):e32457. Epub 2012 Mar 12.

Identification of a New Rhoptry Neck Complex RON9/RON10 in the Apicomplexa Parasite Toxoplasma gondii.

Lamarque MH, Papoin J, Finizio AL, Lentini G, Pfaff AW, Candolfi E, Dubremetz JF, Lebrun M.

UMR 5235 CNRS, Université de Montpellier 2, 34095 Montpellier, France.

Apicomplexan parasites secrete and inject into the host cell the content of specialized secretory organelles called rhoptries, which take part into critical processes such as host cell invasion and modulation of the host cell immune response. The rhoptries are structurally and functionally divided into two compartments. The apical duct contains rhoptry neck (RON) proteins that are conserved in Apicomplexa and are involved in formation of the moving junction (MJ) driving parasite invasion. The posterior bulb contains rhoptry proteins (ROPs) unique to an individual genus and, once injected in the host cell act as effector proteins to co-opt host processes and modulate parasite growth and virulence. We describe here two new RON proteins of Toxoplasma gondii, RON9 and RON10, which form a high molecular mass complex. In contrast to the other RONs described to date, this complex was not detected at the MJ during invasion and therefore was not associated to the MJ complex RON2/4/5/8. Disruptions of either RON9 or RON10 gene leads to the retention of the partner in the ER followed by subsequent degradation, suggesting that the RON9/RON10 complex formation is required for proper sorting to the rhoptries. Finally, we show that the absence of RON9/RON10 has no significant impact on the morphology of rhoptry, on the invasion and growth in fibroblasts in vitro or on virulence in vivo. The conservation of RON9 and RON10 in Coccidia and Cryptosporidia suggests a specific relation with development in intestinal epithelial cells.

PMID: 22427839 [PubMed - in process]

Structure-activity studies of some berberine analogs as inhibitors of Toxoplasma gondii

Bioorg Med Chem Lett. 2012 Feb 22. [Epub ahead of print]

Structure-activity studies of some berberine analogs as inhibitors of Toxoplasma gondii.

Krivogorsky B, Pernat JA, Douglas KA, Czerniecki NJ, Grundt P.

Department of Chemistry & Biochemistry, University of Minnesota Duluth, 1039 University Drive, Duluth, MN 55812, United States.

The shortfalls of the current treatment options against infections with the parasite Toxoplasma gondii (T. gondii) necessitates the development of non-toxic and well-tolerated alternatives. To address this problem a structurally diverse panel of berberine alkaloids was synthesized and evaluated for in vitro inhibition of T. gondii tachyzoites. Inhibitory doses (ID(50)) of less than 50nM and therapeutic indices (TI) up to 4000 were observed.

Copyright © 2012 Elsevier Ltd. All rights reserved.

PMID: 22425568 [PubMed - as supplied by publisher]

Wednesday, March 14, 2012

Epidemiology of ocular toxoplasmosis

Ocul Immunol Inflamm. 2012 Apr;20(2):68-75.

Epidemiology of ocular toxoplasmosis.

Petersen E, Kijlstra A, Stanford M.

Department of Infectious Diseases, Aarhus University Hospital Skejby , Aarhus , Denmark.

Retinal infection with Toxoplasma gondii is the most important cause of posterior uveitis, whereby prevalence and incidence of ocular symptoms after infection depend on socio-economic factors and the circulating parasite genotypes. Ocular toxoplasmosis is more common in South America, Central America, and the Caribbean and parts of tropical Africa as compared to Europe and Northern America, and is quite rare in China. Ocular disease in South America is more severe than in other continents due to the presence of extremely virulent genotypes of the parasite. Drinking untreated water is considered the major source of Toxoplasma infection in developing countries, whereas in the Western world the consumption of raw or undercooked meat (products) is the most important cause. Since acquired infection with T. gondii is currently a more important cause of ocular toxoplasmosis compared to congenital infection, prevention should be directed not only toward pregnant women but toward the general population.

PMID: 22409558 [PubMed - in process]

Friday, March 09, 2012

Galactose recognition by the apicomplexan parasite Toxoplasma gondii

J Biol Chem. 2012 Mar 7. [Epub ahead of print]

Galactose recognition by the apicomplexan parasite Toxoplasma gondii.

Marchant J, Cowper B, Liu Y, Lai L, Pinzan C, Marq JB, Friedrich N, Sawmynaden K, Liew L, Chai W, Childs RA, Saouros S, Simpson P, Roque Barreira MC, Feizi T, Soldati-Favre D, Matthews S.

Imperial College London, United Kingdom;

Toxosplasma gondii is the model parasite of the phylum Apicomplexa, which contains numerous obligate intracellular parasites of medical and veterinary importance, including Eimeria, Sarcocystis, Cryptosporidium, Cyclospora and Plasmodium species. Members of this phylum actively enter host cells by a multi-step process with the help of microneme protein (MIC) complexes that play important roles in motility, host cell attachment, moving junction formation and invasion. Toxoplasma gondii (Tg)MIC1-4-6 complex is the most extensively investigated microneme complex which contributes to host cell recognition and attachment via the action of TgMIC1, a sialic acid-binding adhesin. Here, we report the structure of TgMIC4 and reveal its carbohydrate-binding specificity to a variety of galactose-containing carbohydrate ligands. The lectin is composed of six apple domains in which the fifth domain displays a potent galactose-binding activity, and which is cleaved from the complex during parasite invasion. We propose that galactose recognition by TgMIC4 masks the recognition of the parasite thereby providing protection from galectin-mediated activation of the host immune system.

PMID: 22399295 [PubMed - as supplied by publisher]

Toxoplasma gondii profilin acts primarily to sequester G-actin while formins efficiently nucleate actin filament formation in vitro

Biochemistry. 2012 Mar 7. [Epub ahead of print]

Toxoplasma gondii profilin acts primarily to sequester G-actin while formins efficiently nucleate actin filament formation in vitro.

Skillman KM, Daher W, Ma CI, Soldati-Favre D, Sibley LD.

Apicomplexan parasites employ gliding motility that depends on the polymerization of parasite actin filaments for host cell entry. Despite this requirement, parasite actin remains almost entirely unpolymerized at steady state; formation of filaments required for motility relies on a small repertoire of actin-binding proteins. Previous studies have shown that apicomplexan formins and profilin exhibit canonical functions on heterologous actins from higher eukaryotes; however, their biochemical properties on parasite actins are unknown. We therefore analyzed the impact of T. gondii profilin (TgPRF) and FH1-FH2 domains of two formin isoforms in T. gondii (TgFRM1 and TgFRM2) on the polymerization of T. gondii actin (TgACTI). Our findings based on in vitro assays demonstrate that TgFRM1-FH1-FH2 and TgFRM2-FH1-FH2 dramatically enhanced TgACTI polymerization in the absence of profilin, making them the sole protein factors known to initiate polymerization of this normally unstable actin. In addition, T. gondii formin domains were shown to both initiate polymerization and induce bundling of TgACTI filaments; however, they did not rely on TgPRF for these activities. In contrast, TgPRF sequestered TgACTI monomers, thus inhibiting polymerization even in the presence of formins. Collectively, these findings provide insight into the unusual control mechanisms of actin dynamics within the parasite.

PMID: 22397711 [PubMed - as supplied by publisher]

Wednesday, March 07, 2012

Discovery of a Novel Toxoplasma gondii Conoid-Associated Protein Important for Parasite Resistance to Reactive Nitrogen Intermediates

J Immunol. 2012 Mar 2. [Epub ahead of print]

Discovery of a Novel Toxoplasma gondii Conoid-Associated Protein Important for Parasite Resistance to Reactive Nitrogen Intermediates.

Skariah S, Bednarczyk RB, McIntyre MK, Taylor GA, Mordue DG.

Department of Microbiology and Immunology, New York Medical College, Valhalla, NY 10595;

Toxoplasma gondii modifies its host cell to suppress its ability to become activated in response to IFN-γ and TNF-α and to develop intracellular antimicrobial effectors, including NO. Mechanisms used by T. gondii to modulate activation of its infected host cell likely underlie its ability to hijack monocytes and dendritic cells during infection to disseminate to the brain and CNS where it converts to bradyzoites contained in tissue cysts to establish persistent infection. To identify T. gondii genes important for resistance to the effects of host cell activation, we developed an in vitro murine macrophage infection and activation model to identify parasite insertional mutants that have a fitness defect in infected macrophages following activation but normal invasion and replication in naive macrophages. We identified 14 independent T. gondii insertional mutants out of >8000 screened that share a defect in their ability to survive macrophage activation due to macrophage production of reactive nitrogen intermediates (RNIs). These mutants have been designated counter-immune mutants. We successfully used one of these mutants to identify a T. gondii cytoplasmic and conoid-associated protein important for parasite resistance to macrophage RNIs. Deletion of the entire gene or just the region encoding the protein in wild-type parasites recapitulated the RNI-resistance defect in the counter-immune mutant, confirming the role of the protein in resistance to macrophage RNIs.

PMID: 22387554 [PubMed - as supplied by publisher]

Sunday, March 04, 2012

Role of ATG3 in the parasite Toxoplasma gondii: Autophagy in an early branching eukaryote

Autophagy. 2012 Mar 1;8(3). [Epub ahead of print]

Role of ATG3 in the parasite Toxoplasma gondii: Autophagy in an early branching eukaryote.

Besteiro S.

DIMNP; UMR 5235 CNRS; Université de Montpellier 2; Montpellier, France.

Toxoplasma gondii belongs to the phylum Apicomplexa, a diverse group of early branching unicellular eukaryotes related to dinoflagellates and ciliates. Like several other Apicomplexa such as Plasmodium (the causative agent of malaria), T. gondii is a human pathogen responsible for a potentially lethal disease called toxoplasmosis. Most Apicomplexa have complex life cycles, involving intermediate hosts and vectors, which include obligatory intracellular developmental stages. In the case of malaria and toxoplasmosis, it is that replicative process, leading to the ultimate lysis of the host cell, which is causing the symptoms of the disease. For Toxoplasma, the invasive and fast-replicating form of the parasite is called the tachyzoite. While autophagy has been a fast-growing field of research in recent years, not much was known about the relevance of this catabolic process in medically important apicomplexan parasites. Vesicles resembling autophagosomes had been described in drug-treated Plasmodium parasites in the early 1970s and a potential role for autophagy in organelle recycling during differentiation between Plasmodium life stages has also been recently described. Interestingly, recent database searches have identified putative orthologs of the core machinery responsible for the formation of autophagosomes in several protists, including Toxoplasma. In spite of an apparently reduced machinery (only about one-third of the yeast ATG genes appear to be conserved), T. gondii seemed thus able to perform macroautophagy, but the cellular functions of the pathway for this parasite remained to be demonstrated.

PMID: 22361579 [PubMed - as supplied by publisher]

Interactions between the ROP18 kinase and host cell proteins that aid in the parasitism of Toxoplasma gondii

Acta Trop. 2012 Feb 16. [Epub ahead of print]

Interactions between the ROP18 kinase and host cell proteins that aid in the parasitism of Toxoplasma gondii.

Cheng L, Chen Y, Chen L, Shen Y, Shen J, An R, Luo Q, Du J.

Department of Biochemistry and Molecular Biology, Anhui Medical University, Hefei 230032, China.

Serine/threonine kinases secreted from rhoptry organelles are important virulence factors for Toxoplasma gondii. Among rhoptry proteins, the ROP18 kinase has been identified as a key virulence determinant mediating pathogenesis in T. gondii; however, the molecular mechanisms by which this kinase exerts its pathogenic action remain poorly understood. In this study, the interactions between the ROP18 kinase of Toxoplasma gondii and the host cell proteins were analyzed using a yeast two-hybrid technique. The cMyc-ROP18(25-251) fusion proteins expressed by pGBKT7 plasmids in AH109 yeast were bound to host cell proteins from a human fetal brain cDNA library transformed to AH109 yeast using a mating method. Using these selection procedures, we identified seven host proteins that had not previously been reported to interact with ROP18 such as DDB1, TOR1AIP1, integrin, SLC3A2, TPST2, DERL2 and OCIAD1. These host proteins are associated with DNA repair, transcriptional regulation, translation modification, protein degradation and cell adhesion. Our data strongly support the hypothesis that the secreted kinase ROP18 is involved in several complex cellular pathways for the invasion and commandeering of host functions.

Copyright © 2012. Published by Elsevier B.V.

PMID: 22365922 [PubMed - as supplied by publisher]

The impaired pregnancy outcome in murine congenital toxoplasmosis is associated with a pro-inflammatory immune response

Int J Parasitol. 2012 Feb 17. [Epub ahead of print]

The impaired pregnancy outcome in murine congenital toxoplasmosis is associated with a pro-inflammatory immune response, but not correlated with decidual inducible nitric oxide synthase expression.

Coutinho LB, Gomes AO, Araújo EC, Barenco PV, Santos JL, Caixeta DR, Silva DA, Junior JP, Ferro EA, Silva NM.

Laboratory of Immunopathology, Institute of Biomedical Sciences, Federal University of Uberlândia, Av. Pará 1720, Uberlândia, MG 38400-902, Brazil.

Congenital toxoplasmosis is associated with adverse pregnancy outcome. Despite the type 1 immune response, C57BL/6 mice are more susceptible than BALB/c mice to Toxoplasma gondii infection. Additionally, successful pregnancy appears to be correlated with type 2 T helper maternal immunity and regulatory T cells. In order to investigate the mechanisms of susceptibility/resistance to congenital toxoplasmosis in mice with different genetic backgrounds and the influence of inducible nitric oxide synthase in pregnancy outcome, groups of C57BL/6, BALB/c and C57BL/6 iNOS(-/-) females were orally infected with T. gondii ME-49 strain on day 1 of pregnancy and were sacrificed on day 8 p.i. and day 19 p.i. The uterus and placenta were evaluated for the foetal resorption rate, parasite load, immunological and histological changes. C57BL/6 mice presented inflammatory foci in the decidua (endometrium) of the uterus at a higher frequency than BALB/c mice on day 8 p.i., and a large number of pregnant C57BL/6 mice presented necrotic implantation sites. The parasite was seldom found in the uterus or placenta of either lineage of mice. Interestingly, there was no observed difference in inducible nitric oxide synthase expression in the uterus and placenta of infected mice. In addition, higher levels of TNF-α were detected in serum samples from C57BL/6 mice compared with BALB/c mice. Accordingly, C57BL/6 mice presented with levels of 90% abortion compared with 50% in BALB/c mice on day 19 p.i. C57BL/6 iNOS(-/-) mice showed low placental parasite counts and high absorption rates, similar to wild type mice. The data suggest that the impaired pregnancy outcome due to T. gondii infection in C57BL/6 mice could be associated with a higher inflammatory response leading to cell apoptosis and necrosis of implantation sites compared with BALB/c mice, and this phenomenon was not due to inducible nitric oxide synthase expression in the decidua.

Copyright © 2012 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

PMID: 22366549 [PubMed - as supplied by publisher]

Metabolic pathway analysis and molecular docking analysis for identification of putative drug targets in Toxoplasma gondii: novel approach

Bioinformation. 2012;8(3):134-41. Epub 2012 Feb 3.

Metabolic pathway analysis and molecular docking analysis for identification of putative drug targets in Toxoplasma gondii: novel approach.

Gautam B, Singh G, Wadhwa G, Farmer R, Singh S, Singh AK, Jain PA, Yadav PK.

Toxoplasma gondii is an obligate intracellular apicomplexan parasite that can infect a wide range of warm-blooded animals including humans. In humans and other intermediate hosts, toxoplasma develops into chronic infection that cannot be eliminated by host's immune response or by currently used drugs. In most cases, chronic infections are largely asymptomatic unless the host becomes immune compromised. Thus, toxoplasma is a global health problem and the situation has become more precarious due to the advent of HIV infections and poor toleration of drugs used to treat toxoplasma infection, having severe side effects and also resistance have been developed to the current generation of drugs. The emergence of these drug resistant varieties of T. gondii has led to a search for novel drug targets. We have performed a comparative analysis of metabolic pathways of the host Homo sapiens and the pathogen T. gondii. The enzymes in the unique pathways of T. gondii, which do not show similarity to any protein from the host, represent attractive potential drug targets. We have listed out 11 such potential drug targets which are playing some important work in more than one pathway. Out of these, one important target is Glutamate dehydrogenase enzyme; it plays crucial part in oxidation reduction, metabolic process and amino acid metabolic process. As this is also present in the targets of tropical diseases of TDR (Tropical disease related Drug) target database and no PDB and MODBASE 3D structural model is available, homology models for Glutamate dehydrogenase enzyme were generated using MODELLER9v6. The model was further explored for the molecular dynamics simulation study with GROMACS, virtual screening and docking studies with suitable inhibitors against the NCI diversity subset molecules from ZINC database, by using AutoDock-Vina. The best ten docking solutions were selected (ZINC01690699, ZINC17465979, ZINC17465983, ZINC18141294_03, ZINC05462670, ZINC01572309, ZINC18055497_01, ZINC18141294, ZINC05462674 and ZINC13152284_01). Further the Complexes were analyzed through LIGPLOT. On the basis of Complex scoring and binding ability it is deciphered that these NCI diversity set II compounds, specifically ZINC01690699 (as it has minimum energy score and one of the highest number of interactions with the active site residue), could be promising inhibitors for T. gondii using Glutamate dehydrogenase as Drug target.

PMID: 22368385 [PubMed - in process] PMCID: PMC3283885

Multiple determinants for selective inhibition of apicomplexan calcium-dependent protein kinase CDPK1

J Med Chem. 2012 Feb 27. [Epub ahead of print]

Multiple determinants for selective inhibition of apicomplexan calcium-dependent protein kinase CDPK1.

Larson ET, Ojo KK, Murphy RC, Johnson SM, Zhang Z, Kim JE, Leibly DJ, Fox AM, Reid MC, Dale EJ, Perera BG, Kim J, Hewitt SN, Hol WG, Verlinde CL, Fan E, Van Voorhis WC, Maly DJ, Merritt EA.

Diseases caused by the apicomplexan protozoans Toxoplasma gondii and Cryptosporidium parvum are a major health concern. The life cycle of these parasites is regulated by a family of calcium-dependent protein kinases (CDPKs) that have no direct homologs in the human host. Fortuitously, CDPK1 from both parasites contains a rare glycine gatekeeper residue adjacent to the ATP-binding pocket. This has allowed creation of a series of C3-substituted pyrazolopyrimidine compounds that are potent inhibitors selective for CDPK1 over a panel of human kinases. Here we demonstrate that selectivity is further enhanced by modification of the scaffold at the C1 position. The explanation for this unexpected result is provided by crystal structures of the inhibitors bound to CDPK1 and the human kinase c-SRC. Furthermore, the insight gained from these studies was applied to transform an alternative ATP-competitive scaffold lacking potency and selectivity for CDPK1 into a low nanomolar inhibitor of this enzyme with no activity against SRC.

PMID: 22369268 [PubMed - as supplied by publisher]

Vaccination with a DNA vaccine coding Perforin-like Protein 1 (TgPLP1) and MIC6 Induces Significant Protective Immunity Against Toxoplasma

Clin Vaccine Immunol. 2012 Feb 29. [Epub ahead of print]

Vaccination with a DNA vaccine coding Perforin-like Protein 1 (TgPLP1) and MIC6 Induces Significant Protective Immunity Against Toxoplasma gondii.

Yan HK, Yuan ZG, Song HQ, Petersen E, Zhou Y, Ren D, Zhou DH, Li HX, Lin RQ, Yang GL, Zhu XQ.

State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, CAAS, Lanzhou, Gansu Province 730046, People's Republic of China.

Host cell invasion by Toxoplasma gondii is tightly related to micronemal protein 6 (MIC6) and T. gondii perforin-like protein 1 (TgPLP1). In this study, we constructed a DNA vaccine expressing a fusion protein of TgPLP1/MIC6 using vector pIRESneo, and evaluated the immune response it induced in Kunming mice. Levels of IgG antibody, IFN-γ, IL-2, IL-12, IL-4 and IL-10 were examined. Five mice were randomly chosen from every group (vaccinated or non-vaccinated control groups) and were intragastrically challenged with 80 cysts of the T. gondii PRU strain (genotype II) strain to observe mortality daily. To analyze protection against less virulent challenge, eight mice of each group were orally infected with 20 cysts of the PRU strain at the 14th day after the last immunization. The brain parasite load was evaluated 6 weeks after infection. The results demonstrated that the immunization with pIRESneo/MIC6/PLP1 resulted in the lowest brain cyst count and prolonged the survival time of immunized mice. The levels of Toxoplasma-specific IgG, IFN-γ, IL-2 and IL-12 increased significantly, and the cysts in brains decreased more obviously in the group immunized with the plasmid pIRESneo/MIC6/PLP1 than the other groups (P < 0.05). Compared with pIRESneo/MIC6/PLP1, co-immunization of pIRESneo/MIC6/PLP1 with adjuvant murine IL-18 promoted cellular and humoral immune responses, but did not contribute significantly in cyst reduction (65.43% vs 61.60%) or survival of immunized mice (45.0 ± 2.9 days vs 42.8 ± 2.9 days) (P > 0.05). Furthermore, the study also showed that immune efficacy induced by pIRESneo/MIC6/PLP1 was better than that induced by pVAX/PLP1 or pVAX/MIC6 alone.

PMID: 22379063 [PubMed - as supplied by publisher]