Sunday, July 27, 2008

Dissemination of extracellular and intracellular Toxoplasma gondii tachyzoites in the blood flow

Parasitol Int. 2008 Jul 1. [Epub ahead of print]

Dissemination of extracellular and intracellular Toxoplasma gondii tachyzoites in the blood flow

Unno A, Suzuki K, Xuan X, Nishikawa Y, Kitoh K, Takashima Y.

Department of Veterinary Parasitological Diseases, Gifu University, Yanagido 1-1, Gifu 501-1193, Japan.

Toxoplasma gondii is an intracellular parasite. It has been thought that T. gondii can disseminate throughout the body by circulation of tachyzoite-infected leukocytes (intracellular parasite) in the blood flow. However, a small number of parasites exist as free extracellular tachyzoites in the blood flow (extracellular parasite). It is still controversial whether the extracellular parasites in the blood flow disseminate into the peripheral tissues. In this study, we evaluated the dissemination efficiency of the extracellular and intracellular parasites in the blood flow using GFP-expressing transgenic parasite (PLK/GFP) and DsRed Express-expressing transgenic parasite (PLK/RED). When PLK/GFP and PLK/RED tachyzoites were injected, as intracellular and extracellular forms respectively, at the same time into the tail vein of a mouse, many disseminated green fluorescent PLK/GFP tachyzoites were observed in the lung, the spleen, the liver and the brain. However, only a few red fluorescent PLK/RED tachyzoites were detected in these organs. When PLK/GFP and PLK/RED tachyzoites were injected in the opposite manner, that is, as extracellular and intracellular forms respectively, the majority of tachyzoites in these tissues were PLK/RED tachyzoites. Collectively, these results indicate that intracellular tachyzoites mainly disseminate throughout the body and that extracellular tachyzoites hardly contribute to parasite dissemination.

PMID: 18652914 [PubMed - as supplied by publisher]

Friday, July 25, 2008

The mother of all parasites

Future Microbiol. 2008 Aug;3:391-395

The mother of all parasites

Okamoto N, McFadden GI.

School of Botany, University of Melbourne, VIC 3010, Australia and, Department of Botany, University of Birtish Columbia, BC, V6T 1Z4, Canada. , School of Botany, University of Melbourne, VIC 3010, Australia.

Evaluation of: Moore RB, Obornik M, Janouskovec J et al.: A photosynthetic alveolate closely related to apicomplexan parasites. Nature 451(7181), 959-963 (2008). Malaria and related apicomplexan parasites contain a relict plastid (apicoplast) that is a promising drug target. The apicoplast has been argued to derive from either an engulfed red or green alga. The discovery of the first photosynthetic apicomplexan, dubbed Chromera velia, with a fully functional plastid resolves the debate, clearly showing that the relict plastid is derived from a modified red alga. Intriguingly, C. velia is a coral symbiont and thus reminiscent of the closely related dinoflagellate symbionts (zooxanthellae) vital to corals and many other invertebrates. Symbiosis and parasitism are thus wide-spread in both the dinoflagellates and apicomplexans, suggesting that modern parasites like Plasmodium spp. and Toxoplasma likely started out as mutualistic symbionts that initially nourished their animal hosts before turning to parasitism. These symbiotic/parasitic relationships thus extend back in evolutionary time to the earliest origins of the animals, which means that either as parasites or symbionts, these protists have been interacting with the animal immune system since its inception. As a consequence of this protracted dance, malaria parasites are exquisitely well-equipped to evade our immune system: a sobering harbinger for malaria vaccine prospects.

PMID: 18651810 [PubMed - as supplied by publisher]

Thursday, July 24, 2008

Stress response pathways in protozoan parasites

Cell Microbiol. 2008 Jul 17. [Epub ahead of print]

Stress response pathways in protozoan parasites

Vonlaufen N, Kanzok SM, Wek RC, Sullivan Jr WJ

Department of Pharmacology & Toxicology, Indiana University School of Medicine, Indianapolis, IN, USA

Diseases caused by protozoan parasites have a dramatic impact on world health. Emerging drug resistance and a general lack of experimental understanding has created a void in the medicine cabinet used to treat these widespread infections. A novel therapeutic idea that is receiving more attention is centered on targeting the microbe's response to the multitude of environmental stresses it encounters. Protozoan pathogens have complex life cycles, often having to transition from one host to another, or survive in a cyst form in the environment until a new host arrives. The need to respond to environmental cues and stress, and endure in less than optimal conditions, is paramount to their viability and successful progression through their life cycle. This review summarizes the research on parasitic stress responses for Apicomplexa, kinetoplastids, and anaerobic protozoa, with an eye towards how these processes may be exploited therapeutically.

PMID: 18647172 [PubMed - as supplied by publisher]

The proteome of Toxoplasma gondii

Genome Biol. 2008 Jul 21;9(7):R116. [Epub ahead of print]

The proteome of Toxoplasma gondii: integration with the genome provides novel insights into gene expression and annotation

Xia D, Sanderson SJ, Jones AR, Prieto JH, Yates JR 3rd, Bromley E, Tomley FM, Lal K, Sinden RE, Brunk BP, Roos DS, Wastling JM.

ABSTRACT: BACKGROUND: Although the genomes of many of the most important human and animal pathogens have now been sequenced, our understanding of the actual proteins expressed by these genomes and how well they predict protein sequence and expression is still deficient. We have used three complementary approaches (two-dimensional electrophoresis, gel LC-MS/MS and MudPIT) to analyse the proteome of Toxoplasma gondii, a parasite of medical and veterinary significance and have developed a public repository for these data within ToxoDB, making for the first time proteomics data an integral part of this key genome resource. RESULTS: The draft genome for Toxoplasma predicts around 8000 genes with varying degrees of confidence. Our data demonstrate how proteomics can inform these predictions and help discover new genes. We have identified nearly one third (2252) of all the predicted proteins, with 2477 intron-spanning peptides providing supporting evidence for correct splice site annotation. Functional predictions for each protein and key pathways were determined from the proteome. Importantly, we show evidence for many proteins that match alternative gene models, or previously unpredicted genes. For example, approximately 15% of peptides matched more convincingly to alternative gene models. We also compared our data with existing transcriptional data in which we highlight apparent discrepancies between gene transcription and protein expression. CONCLUSIONS: Our data demonstrate the importance of protein data in expression profiling experiments and highlight the necessity of integrating proteomic with genomic data so that iterative refinements of both annotation and expression models are possible.

PMID: 18644147 [PubMed - as supplied by publisher]

Wednesday, July 23, 2008


Molecular and biochemical parasitology:
epigenetics and gene expression

POSTDOCTORAL POSITION available to investigate the molecular basis of epigenetics and gene expression during development of the protozoan parasite Toxoplasma gondii. Related to the malaria parasite, Toxoplasma causes serious disease in immunocompromised (AIDS) patients due to the ability of the parasite to develop into cysts in the body. We have shown that epigenetics plays an important role in this critical developmental transition (Sullivan et al., 2006, Cell Microbiol, 8:1850). Potential projects include biochemical purification of chromatin remodeling complexes and functional characterization of parasite mutants defective in histone modifying enzymes. See for more information.

Position requires a Ph.D., expertise in biochemistry or molecular biology, and excellent communication skills. Experience in molecular parasitology or chromatin biology is a plus. Submit CV and contact information for three references to Dr. Bill Sullivan at

Located in downtown Indianapolis, Indiana University School of Medicine (IUSM) is the second largest medical school in the US and boasts an outstanding intellectual atmosphere and core facilities. IUSM was nationally ranked in the Top 30 Best Places to Work for Postdocs (2008 survey, The Scientist). IUSM is an equal opportunity employer.

Sunday, July 20, 2008

Toxoplasma gondii: An improved rat model of congenital infection

Exp Parasitol. 2008 Jun 25. [Epub ahead of print]

Toxoplasma gondii: An improved rat model of congenital infection

Freyre A, Falcón J, Méndez J, González M.

Laboratory for Toxoplasmosis, Department of Parasitology, College of Veterinary Sciences, Montevideo, Uruguay; Laboratory for Parasitology, College for Chemistry, Montevideo, Uruguay.

The objective of this study was to refine the rat model of congenital toxoplasmosis. In Fischer rates we found that visualization of spermatozoa in vaginal exudates and the detection of at least 6g body weight increase between days 9 and 12 of pregnancy, allowed the diagnosis and timing of pregnancy with 60% specificity and 84% sensitivity. A dose of 10(4)Toxoplasma gondii bradyzoites or 10(2)T. gondii oocysts of the Prugniaud strain resulted in more than 50% of congenital infection of the rat litters. Transmission of T. gondii via lactation was not detected in rats inoculated with either bradyzoites or oocysts. Bioassays of 51 neonates born from mothers inoculated with bradyzoites (in tissue cysts) and 29 neonates from mothers inoculated with oocysts demonstrated that both liver and lungs can be used for the diagnosis of congenital transmission in this model.

PMID: 18634785 [PubMed - as supplied by publisher]

Friday, July 18, 2008

Different effect of Toxoplasma gondii infection on adhesion capacity of fibroblasts and monocytes

Parasite Immunol. 2008 Jul 1. [Epub ahead of print]

Different effect of Toxoplasma gondii infection on adhesion capacity of fibroblasts and monocytes

Pfaff AW, Georges S, Candolfi E.

Institut de Parasitologie et de Pathologie Tropicale, Interaction Cellulaire et Moléculaire Hôte-Parasite (E.A. 3950), Faculté de Médecine, Strasbourg, France.

This study investigated the effect of infection with the apicomplexan parasite Toxoplasma gondii, in combination with the concomitant cytokine environment (IFN-gamma/TNF-alpha), on adhesion of THP-1 monocytic cells to MRC-5 fibroblasts. Surprisingly, infection of THP-1 cells decreased their adhesion to the MRC-5 cell monolayer. This decrease was compensated by IFN-gamma/TNF-alpha stimulation. In contrast, infection of MRC-5 cells significantly increased adhesion, which was synergistically augmented by cytokine stimulation. Levels of ICAM-1 (CD54) on MRC-5 cells, as well as LFA-1 (CD11a) on THP-1 cells, were not changed by infection, neither in resting, nor in cytokine stimulated cells. These results show that T. gondii infection alters adhesion properties and reactivity to cytokine stimulation in a cell-specific way.

PMID: 18627508 [PubMed - as supplied by publisher]

In pregnant mice, the infection of Toxoplasma gondii causes the decrease of CD4(+)CD25(+)-regulatory T cells

Parasite Immunol. 2008 Jul 1. [Epub ahead of print]

In pregnant mice, the infection of Toxoplasma gondii causes the decrease of CD4(+)CD25(+)-regulatory T cells

Ge YY, Zhang L, Zhang G, Wu JP, Tan MJ, Hu W, Liang YJ, Wang Y.

Laboratory of Reproductive Medicine, Nanjing Medical University, Nanjing, China.

Acute maternal infection with Toxoplasma gondii during pregnancy is associated with adverse pregnancy outcomes. Although previous reports have indicated that T. gondii may result in abortion without direct transmission of the parasite to the foetus, the molecular mechanism remains unclear. CD4(+)CD25(+)-regulatory T cells are known to be involved in maternal tolerance toward the foetus-bearing alloantigens. With a model of pregnant mice infected with T. gondii, we found that Foxp3 mRNA expression levels in both splenocytes and placenta were reduced markedly during the process of infection. Furthermore, the numbers of splenic CD4(+)CD25(+)-regulatory T cells and placental Foxp3(+) cells decreased synchronously in the infected mice, and the reduction of splenic CD4(+)CD25(+)-regulatory T cells were associated with apoptosis induced by the infection. Additionally, injection of pregnant mice with excretory-secretory antigens (ESA) of T. gondii also resulted in foetal loss, which could be partly prevented by adoptive transfer of CD4(+)CD25(+)-regulatory T cells from normal pregnant mice. These data suggest that foetal loss caused by T. gondii can be independent of vertical infection and that the decrease of CD4(+)CD25(+)-regulatory T cells during infection may represent a previously unrecognized mechanism for the pathogenesis of abortion caused by this parasite.

PMID: 18627509 [PubMed - as supplied by publisher]

Unusual Sorting Mechanisms to Deliver Transmembrane Proteins into the Host-Cell Vacuole

Traffic. 2008 Jul 7. [Epub ahead of print]

Toxoplasma gondii Uses Unusual Sorting Mechanisms to Deliver Transmembrane Proteins into the Host-Cell Vacuole
Gendrin C, Mercier C, Braun L, Musset K, Dubremetz JF, Cesbron-Delauw MF.

Laboratoire Adaptation et Pathogénie des Micro-organismes, CNRS UMR 5163, Université Joseph Fourier GRENOBLE 1, France.

A critical step in infection by the apicomplexan parasite Toxoplasma gondii, is the formation of a membrane-bound compartment within which the parasite proliferates. This process relies on a set of secretory organelles that discharge their contents into the host-cell upon invasion. Among these organelles, the dense granules are specialized in the export of transmembrane GRA proteins which are major components of the mature parasitophorous vacuole (PV) membrane. How eukaryotic pathogens export and sort membrane-bound proteins destined to the host cell is still poorly understood at the mechanistic level. In the present study, we show that soluble trafficking of the PV-targeted GRA5 transmembrane protein is parasite-specific: when expressed in mammalian cells, GRA5 is targeted to the plasma membrane and behaves as an integral membrane protein with a type I toplogy. We also demonstrate the dual role of the GRA5 N-terminal ectodomain, which is sufficient to prevent membrane integration within the parasite and is essential for both sorting and post-secretory membrane insertion into the vacuolar membrane. These results contrast with the general rule which states that information contained within the cytoplasmic tail and/or the transmembrane domain of integral membrane proteins dictates their cellular localization. They also highlight the diversity of sorting mechanisms that leads to the specialization of secretory processes uniquely adapted to intracellular parasitism.

PMID: 18631244 [PubMed - as supplied by publisher]

Stress-related and spontaneous stage differentiation of Toxo

Mol Biosyst. 2008 Aug;4(8):824-834. Epub 2008 Jun 2

Stress-related and spontaneous stage differentiation of Toxoplasma gondii

Ferreira da Silva MD, Barbosa HS, Groß U, Lüder CG.

Institute for Medical Microbiology, Georg-August-University, Kreuzbergring 57, D-37075, Göttingen, Germany.

Toxoplasma gondii is an obligatory intracellular parasitic protozoan that infects a variety of avian and mammalian hosts including up to one third of the human population worldwide. Developmental differentiation between distinct stages, i.e. sporozoites, tachyzoites and bradyzoites is fundamental for the parasite life cycle and for transmission between hosts. It is also interconnected with the pathogenesis of overt toxoplasmosis and makes T. gondii an important opportunistic pathogen of humans. In order to delineate the underlying mechanisms, several cell culture differentiation systems have been developed which mimic the transition from fast-replicating tachyzoites to slowly proliferating bradyzoites in vitro. Since exogenous stress factors, i.e. alkaline pH, IFN-gamma and other proinflammatory cytokines, chemicals or drugs, heat shock, and deprivation of nutrients have been shown to increase the efficacy of bradyzoite development in vitro, Toxoplasma stage differentiation is largely viewed as a stress-related response to hostile environmental conditions. However, tachyzoite to bradyzoite differentiation also occurs spontaneously in vitro and this raises questions about the importance of stress conditions for triggering stage conversion. High frequencies of spontaneous bradyzoite development in primary and permanent skeletal muscle cells, i.e. cells that preferentially harbour bradyzoite-containing tissue cysts in vivo suggest that the host cell type may be critical. Furthermore, the host cell transcriptome, including the expression of distinct host cell genes, has recently been shown to trigger bradyzoite development and cyst formation. Together, these results strongly indicate that the complex cellular environment, besides exogenous stress factors, may govern the developmental differentiation of T. gondii.

PMID: 18633484 [PubMed - as supplied by publisher]

Thursday, July 17, 2008

Protein Ser/Thr phosphatases of parasitic protozoa

Mol Biochem Parasitol. 2008 Jun 21. [Epub ahead of print]

Protein Ser/Thr phosphatases of parasitic protozoa

Kutuzov MA, Andreeva AV.

Department of Pharmacology (MC 868), University of Illinois at Chicago, 909 S. Wolcott Avenue, Chicago, IL 60612, USA.

Protein phosphorylation is an important mechanism implicated in physiology of any organism, including parasitic protozoa. Enzymes that control protein phosphorylation (kinases and phosphatases) are considered promising targets for drug development. This review attempts to provide the first account of the current understanding of the structure, regulation and biological functions of protein Ser/Thr phosphatases in unicellular parasites. We have examined the complements of phosphatases ("phosphatomes") of the PPP and PPM families in several species of Apicomplexa (including malaria parasite Plasmodium), as well as Giardia lamblia, Entamoeba histolytica, Trichomonas vaginalis and a microsporidium Encephalitozoon cuniculi. Apicomplexans have homologues (in most cases represented by single isoforms) of all human PPP subfamilies. Some apicomplexan PPP phosphatases have no orthologues in their vertebrate hosts, including a previously unrecognised group of pseudo-phosphatases with putative Ca(2+)-binding domains, which we designate as EFPP. We also describe the presence of previously undetected Zn finger motifs in PPEF phosphatases from kinetoplastids, and a likely case of convergent evolution of tetratricopeptide repeat domain-containing phosphatases in G. lamblia. Among the parasites examined, E. cuniculi has the smallest Ser/Thr phosphatome (5 PPP and no PPM), while T. vaginalis shows the largest expansion of the PPP family (169 predicted phosphatases). Most protozoan PPM phosphatases cluster separately from human sequences. The structural peculiarities or absence of human orthologues of a number of protozoan protein Ser/Thr phosphatases makes them potentially suitable targets for chemotherapy and thus warrants their functional assessment.

PMID: 18619495 [PubMed - as supplied by publisher]

Longitudinal Study of New Eye Lesions in Children with Toxoplasmosis Who Were Not Treated During the First Year of Life

Am J Ophthalmol. 2008 Jul 9. [Epub ahead of print]

Longitudinal Study of New Eye Lesions in Children with Toxoplasmosis Who Were Not Treated During the First Year of Life

Phan L, Kasza K, Jalbrzikowski J, Noble AG, Latkany P, Kuo A, Mieler W, Meyers S, Rabiah P, Boyer K, Swisher C, Mets M, Roizen N, Cezar S, Sautter M, Remington J, Meier P, McLeod R.

University of Chicago School of Medicine, University of Chicago.

PURPOSE: To determine the incidence of new chorioretinal lesions in children with toxoplasmosis diagnosed after, and therefore not treated during, their first year. DESIGN: Prospective longitudinal cohort study. METHODS: Thirty-eight children were evaluated in Chicago between 1981 and 2005 for new chorioretinal lesions. Thirty-eight children and mothers had serum IgG antibody to Toxoplasma gondii. RESULTS: Twenty-eight of 38 children had one of the following: diagnosis with serum antibody to T. gondii indicative of chronic infection at age 24 months, central nervous system calcifications, hydrocephalus, illness compatible with congenital toxoplasmosis perinatally but not diagnosed at that time. Twenty-five returned for follow-up during 1981 to 2005. Their mean (range) age at last exam was 10.9 +/- 5.7 (range, 3.5 to 27.2) years and mean follow-up was 5.7 +/- 2.9 years. Eighteen (72%) children developed at least one new lesion. Thirteen (52%) had new central lesions, 11 (44%) had new peripheral lesions, and six (24%) had both. Thirteen (52%) had new lesions diagnosed at age >/=10 years. New lesions were found at more than one visit in four (22%), and bilateral new lesions developed in seven (39%) of 18 children who developed new lesions. Of 10 additional children with eye findings and serologic tests indicative of chronic infection, six returned for follow-up, four (67%) developing new lesions at >/=10 years of age. CONCLUSIONS: More than 70% developed new chorioretinal lesions. New lesions were commonly diagnosed after the first decade of life.

PMID: 18619570 [PubMed - as supplied by publisher]

Anti-Toxoplasma gondii RH strain activity of herbal extracts used in traditional medicine

Int J Antimicrob Agents. 2008 Jul 9. [Epub ahead of print]

Anti-Toxoplasma gondii RH strain activity of herbal extracts used in traditional medicine

Choi KM, Gang J, Yun J.

Ecodigm Co., Ltd., 305DTV Post BI, 707 Tamnip-dong, Yuseong-gu, Daejeon 305-510, South Korea.

Methanolic extracts of 15 traditional medicines used to treat Toxoplasma gondii were tested in vitro for their anti-T. gondii activity and cytotoxicity. The median effective concentration (EC(50)) values for the herbal extracts ranged from 0.11mg/mL to 2.28mg/mL. Significant anti-T. gondii RH strain activity was observed with Zingiber officinale extracts (EC(50)=0.18mg/mL), which displayed a highly selective toxicity (selectivity=10.1). Sophora flavescens Aiton extracts also showed high anti-T. gondii activity (EC(50)=0.20mg/mL) and a high selective toxicity (4.6). This indicates that Z. officinale and S. flavescens Aiton extracts may be sources of new anti-T. gondii compounds.

PMID: 18619816 [PubMed - as supplied by publisher]

Specificity of the Toxoplasma gondii-altered behaviour to definitive versus non-definitive host predation risk

Parasitology. 2008 Jul 14:1-8. [Epub ahead of print]

Specificity of the Toxoplasma gondii-altered behaviour to definitive versus non-definitive host predation risk

Lamberton PH, Donnelly CA, Webster JP.

Department of Zoology, University of Oxford, South Parks Road, Oxford OX1 3SY, UK.

SUMMARYThe hypothesis that the parasite Toxoplasma gondii manipulates the behaviour of its intermediate rat host in order to increase its chance of being predated specifically by its feline definitive host, rather than a non-definitive host predator species, was tested. The impact of a range of therapeutic drugs, previously demonstrated to be effective in preventing the development of T. gondii-associated behavioural and cognitive alterations in rats, on definitive-host predator specificity was also tested. Using a Y-shaped maze design, we demonstrated that T. gondii-associated behavioural changes, apparently aimed to increase predation rate, do appear to be specific to that of the feline definitive host - there were significant and consistent differences between the (untreated) infected and uninfected rats groups where T. gondii-infected rats tended to choose the definitive host feline-predator-associated maze arm and nest-box significantly more often than a maze arm or nest-box treated with non-definitive host predator (mink) odour. Drug treatment of infected rats prevented any such host-specificity from being displayed. We discuss our results in terms of their potential implications both for T. gondii epidemiology and the evolution of parasite-altered behaviour.

PMID: 18620624 [PubMed - as supplied by publisher]

Management of Toxoplasma gondii Infection during Pregnancy

Clin Infect Dis. 2008 Jul 14. [Epub ahead of print]

Management of Toxoplasma gondii Infection during Pregnancy

Montoya JG, Remington JS

Palo Alto Medical Foundation Toxoplasma Serology Laboratory, Palo Alto, and Department of Medicine and Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California.

Acute infection with Toxoplasma gondii during pregnancy and its potentially tragic outcome for the fetus and newborn continue to occur in the United States, as well as worldwide, despite the fact that it can be prevented. The infection can be acquired through ingestion of infected, undercooked meat or contaminated food or water. Transmission to the fetus occurs almost solely in women who acquire their primary infection during gestation and can result in visual and hearing loss, mental and psychomotor retardation, seizures, hematological abnormalities, hepatosplenomegaly, or death. Systematic education and serological screening of pregnant women are the most reliable and currently available strategies for the prevention, diagnosis, and early treatment of the infection in the offspring; this is largely because toxoplasmosis in pregnant women most often goes unrecognized. Treatment of the infection in the fetus and infant during the first year of life has been demonstrated to significantly improve the clinical outcome.

PMID: 18624630 [PubMed - as supplied by publisher]

Monday, July 14, 2008

Toll-like receptors and their role in host resistance to Toxoplasma

Immunol Lett. 2008 Jun 18. [Epub ahead of print]

Toll-like receptors and their role in host resistance to Toxoplasma gondii

Yarovinsky F

Department of Immunology, University of Texas Southwestern Medical Center, 5323 Harry Hines Blvd, Dallas, TX 75390-9093, United States

Toxoplasma gondii and other apicomplexan parasites are widely distributed obligate intracellular protozoa. A critical host mediator produced in response to T. gondii infection is IL-12. This cytokine is synthesized by dendritic cells, macrophages and neutrophils and plays a pivotal role in the production of IFN-gamma, which in turn activates anti-microbial effector cells. In the past several years, many of the receptors and signaling pathways that link pathogen detection to induction of IL-12 have been identified and characterized. Among these receptors the Toll-like receptor (TLR) family can recognize all classes of pathogens and induce different types of immune responses. In the following review, I summarize the evidence for specific TLR function in host resistance to T. gondii.

PMID: 18617274 [PubMed - as supplied by publisher]

Wednesday, July 09, 2008

Anti-Toxoplasma gondii activity of GAS in vitro

J Ethnopharmacol. 2008 Jul 4. [Epub ahead of print]

Anti-Toxoplasma gondii activity of GAS in vitro

Chen SX, Wu L, Jiang XG, Feng YY, Cao JP.

School of Medical Technology, Jiangsu University, Zhenjiang 212013, People's Republic of China.

ETHNOPHARMACOLOGICAL RELEVANCE: The sarcotesta of Ginkgo biloba is a Chinese herbal medicine used for treating toxoplasmosis, a serious disease requiring treatment with antibiotics that can have serious side effects. AIM OF THE STUDY: To investigate the anti-Toxoplasmagondii activity of ginkgolic acids (GAs) isolated from the Ginkgo biloba sarcotesta in Toxoplasmagondii-infected human foreskin fibroblast (HFF) cells in vitro. MATERIALS AND METHODS: The safe concentration of GAs for HFF cells was determined by methyl thiazolyl tetrazolium (MTT) cell proliferation assay. The presence of Toxoplasmagondii was measured by [(3)H]-thymine deoxyriboside ([(3)H]-TdR) and [(3)H]-leucine ([(3)H]-Leu) incorporation, as well as Giemsa staining. The positive control was the commonly used and highly effective antibiotic azithromycin. RESULTS: Light microscopy revealed that most HFF cells were infected after 4h of exposure to Toxoplasmagondii. After 48h of exposure to either GAs or azithromycin, Toxoplasmagondii DNA and protein synthesis were minimal, there were no visible parasites in HFF cells, and the HFF cells had no significant morphological changes. CONCLUSIONS: These results demonstrate that GAs have significant anti-Toxoplasma activity with low toxicity to HFF cells, suggesting that GAs could be an alternative treatment for toxoplasmosis.

PMID: 18602775 [PubMed - as supplied by publisher]

Antifolates as Inhibitors of Dihydrofolate Reductase

J Med Chem. 2008 Jul 8. [Epub ahead of print]

The Effect of 5-Alkyl Modification on the Biological Activity of Pyrrolo[2,3-d]pyrimidine Containing Classical and Nonclassical Antifolates as Inhibitors of Dihydrofolate Reductase and as Antitumor and/or Antiopportunistic Infection Agents-

Gangjee A, Jain HD, Queener SF, Kisliuk RL.

Division of Medicinal Chemistry, Graduate School of Pharmaceutical Sciences, Duquesne University, Pittsburgh, Pennsylvania 15282, Department of Pharmacology and Toxicology, School of Medicine, Indiana University, Indianapolis, Indiana 46202, Department of Biochemistry, Tufts University School of Medicine, Boston, Massachusetts 02111

Novel classical antifolates ( 3 and 4) and 17 nonclassical antifolates ( 11- 27) were synthesized as antitumor and/or antiopportunistic infection agents. Intermediates for the synthesis of 3, 4, and 11- 27 were 2,4-diamino-5-alkylsubstituted-7 H-pyrrolo[2,3- d]pyrimidines, 31 and 38, prepared by a ring transformation/ring annulation sequence of 2-amino-3-cyano-4-alkyl furans to which various aryl thiols were attached at the 6-position via an oxidative addition reaction using I 2. The condensation of alpha-hydroxy ketones with malonodinitrile afforded the furans. For the classical analogues 3 and 4, the ester precursors were deprotected, coupled with diethyl- l-glutamate, and saponified. Compounds 3 (IC 50 = 60 nM) and 4 (IC 50 = 90 nM) were potent inhibitors of human DHFR. Compound 3 inhibited tumor cells in culture with GI 50 500-fold selectivity over human DHFR. Analogue 17 was 50-fold more potent than trimethoprim and about twice as selective against T. gondii DHFR.

PMID: 18605720 [PubMed - as supplied by publisher]

Macrophage migration inhibitory factor (MIF) is critical for the host resistance against Toxo

FASEB J. 2008 Jul 7. [Epub ahead of print]

Macrophage migration inhibitory factor (MIF) is critical for the host resistance against Toxoplasma gondii

Flores M, Saavedra R, Bautista R, Viedma R, Tenorio EP, Leng L, Sánchez Y, Juárez I, Satoskar AA, Shenoy AS, Terrazas LI, Bucala R, Barbi J, Satoskar AR, Rodriguez-Sosa M.

*Unidad de Biomedicina, FES-Iztacala and Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México (UNAM), Mexico City, Mexico; Department of Medicine, Yale University School of Medicine, New Haven, Connecticut, USA; Department of Pathology, Seth G. S. Medical College and KEM Hospital, Mumbai, India; and paragraph signDepartment of Pathology, The Ohio State University Medical Center, and ||Department of Microbiology, The Ohio State University College of Medicine, Columbus, Ohio, USA.

Macrophage migration inhibitory factor (MIF) exerts either a protective or a deleterious role in the immune response to different pathogens. We analyzed herein the role of MIF in the host control of toxoplasmosis using MIF(-/-) mice backcrossed to either the BALB/c or the C57BL/6 genetic backgrounds. Both, wild-type (WT) BALB/c and MIF(-/-) BALB/c mice were susceptible to infection with highly virulent RH as well as moderately virulent ME49 strains of T. gondii. MIF(-/-) mice, however, showed greater liver damage and more brain cysts, produced less proinflammatory cytokines, and succumbed significantly faster than WT mice. Bone marrow-derived dendritic cells (BMDCs) from MIF(-/-) mice produced less interleukin-1beta, interleukin-12, and tumor necrosis factor-alpha than WT BMDCs after stimulation with soluble Toxoplasma antigen (STAg). Similar observations were made in CD11c(+) low-density cells isolated from the spleens of MIF(-/-) mice challenged with STAg. MIF(-/-) C57BL/6 mice succumbed to ME49 infection faster than their WT counterparts. C57BL/6 mice that succumbed to infection with the ME49 strain produced less MIF than resistant BALB/c mice similarly infected. Interestingly, an analysis of brains from patients with cerebral toxoplasmosis showed low levels of MIF expression. Together, these findings demonstrate that MIF plays a critical role in mediating host resistance against T. Gondii.-Flores, M., Saavedra, R., Bautista, R., Viedma, R., Tenorio, E. P., Leng, L., Sánchez, Y., Juárez, I., Satoskar, A. A., Shenoy, A. S., Terrazas, L. I., Bucala, R., Barbi J., Satoskar, A. R., Rodriguez-Sosa, M. Macrophage migration inhibitory factor (MIF) is critical for the host resistance against Toxoplasma gondii..

PMID: 18606868 [PubMed - as supplied by publisher]

Thursday, July 03, 2008

Protective effect of a DNA vaccine delivered in attenuated Salmonella typhimurium against Toxoplasma gondii infection in mice

Vaccine. 2008 Jun 27. [Epub ahead of print]

Protective effect of a DNA vaccine delivered in attenuated Salmonella typhimurium against Toxoplasma gondii infection in mice

Qu D, Wang S, Cai W, Du A.

Institute of Preventive Veterinary Medicine, Zhejiang University, Hangzhou, Zhejiang 310029, China.

Infection with the intracellular protozoan parasite Toxoplasma gondii causes serious public health problems and is of great economic importance worldwide. The T. gondii surface antigen protein SAG1 is a significant candidate vaccine against toxoplasmosis. In this study, we evaluated safety, stability of ZJ111/pcDNA3-SAG1, a DNA vaccine delivered in attenuated Salmonella typhimurium, and immune responses induced by immunizing ICR mice orally with ZJ111/pcDNA3-SAG1 of different doses. Mice had no significant differences in body weight between the groups before immunization and at week 4 after the booster immunization. The ZJ111/pcDNA3-SAG1 was eventually eliminated from the spleen and liver on week 6 post-immunization. The plasmid pcDNA3-SAG1 was stably maintained over 90% of the attenuated S. typhimurium population after 100 generations of growth in antibiotic-free media. Oral immunization of mice with ZJ111/pcDNA3-SAG1 elicited specific humoral responses and stimulated proliferation of splenocytes (P<0.05). The cellular immune response was associated with the production of IFN-gamma, indicating that a Th-1 type response was elicited, which was confirmed by the production of large amounts of IgG2a (P<0.01). Mice immunized with ZJ111/pcDNA3-SAG1 displayed significant protection against an intraperitoneally challenge with 500 tachyzoite forms of T. gondii RH strain. Vaccination at 10(7) and 10(8)CFU per mice provided a 20% and 10% survival rate comparing 100% mortality of the non-immunized mice, exhibiting longer living time and better survival rate. These results confirmed a DNA vaccine delivered in attenuated S. typhimurium, ZJ111/pcDNA3-SAG1, can elicit specific immune response as well as provide effective protection against T. gondii infection, and the dosage of 10(7)CFU was a most considerate one.

PMID: 18590785 [PubMed - as supplied by publisher]

Wednesday, July 02, 2008

In vivo study of toxoplasmic parasitemia using interferon-gamma-deficient mice

Parasitol Int. 2008 Jun 8. [Epub ahead of print]

In vivo study of toxoplasmic parasitemia using interferon-gamma-deficient mice: Absolute cell number of leukocytes, parasite load and cell susceptibility

Norose K, Naoi K, Fang H, Yano A.

Department of Infection and Host Defense, Graduate School of Medicine, Chiba University, 1-8-1, Inohana, Chuo-ku, Chiba 260-8670, Japan.

Toxoplasma gondii infection was induced in wild type (WT) C57BL/6 and BALB/c mice and same-background interferon-gamma knockout (GKO) mice by peroral inoculation of Fukaya strain cysts. We studied parasitemia, absolute cell number of leukocytes, and cell susceptibility in peripheral blood leukocyte (PBL) subsets in vivo. Parasitemia was observed in both WT and GKO mice, although the pattern of the parasite load was totally different among them. After infection, the absolute number of neutrophils in peripheral blood increased in both GKO C57BL/6 and GKO BALB/c mice with statistical significance, while it rose up slightly in susceptible WT C57BL/6 mice, but declined moderately in resistant WT BALB/c mice. The absolute number of lymphocytes in both WT and GKO mice decreased significantly after infection. Although leukocyte number per mul decreased significantly in both strains of WT mice, it increased in GKO BALB/c mice. There were no differences in susceptibility of PBL subsets to T. gondii infection as assessed by quantitative competitive polymerase chain reaction in both WT and GKO mice. These results indicate that each of the PBL subsets was infected in vivo. The increase of neutrophils only among immunocompromised or susceptible strains suggests that the neutrophil may be involved in the lethal process of T. gondii infection. The lack of any difference in cell susceptibility per mug gDNA among the PBL subsets examined implies that the neutrophil may contribute to the whole body dissemination process of the parasite in vivo more than other PBL subsets through the increase in number.

PMID: 18585475 [PubMed - as supplied by publisher]

A thioredoxin family protein of the apicoplast periphery identifies abundant candidate transport vesicles in Toxoplasma

Eukaryot Cell. 2008 Jun 27. [Epub ahead of print]

A thioredoxin family protein of the apicoplast periphery identifies abundant candidate transport vesicles in Toxoplasma

Derocher AE, Coppens I, Karnataki A, Gilbert LA, Rome ME, Feagin JE, Bradley PJ, Parsons M.

Seattle Biomedical Research Institute, 307 Westlake Ave. N., Seattle, WA 98109, USA; Department of Molecular Microbiology and Immunology, Johns Hopkins University Bloomberg School of Public Health, 615 N. Wolfe Street, Baltimore, MD 21205, USA; Interdisciplinary Program in Pathobiology, Department of Global Health, University of Washington, Seattle, WA 98195, USA; and Department of Microbiology, Immunology, and Molecular Genetics, 609 Charles E. Young Dr. East, University of California, Los Angeles, CA, 90095, USA.

Toxoplasma gondii, which causes toxoplasmic encephalitis and birth defects, contains an essential chloroplast-related organelle to which proteins are trafficked via the secretory system. This organelle, the apicoplast, is bounded by multiple membranes. In this report we identify a novel apicoplast-associated thioredoxin family protein, ATrx1, which is predominantly soluble or peripherally associated with membranes, and which localizes primarily to the outer compartments of the organelle. As such, it represents the first protein to be identified as residing in the apicoplast intermembrane spaces. ATrx1 lacks the apicoplast targeting sequences typical of luminal proteins. However, sequences near the N-terminus are required for proper targeting of ATrx1, which is proteolytically processed from a larger precursor to multiple smaller forms. This protein reveals a population of vesicles, hitherto unrecognized as being highly abundant in the cell, which may serve to transport proteins to the apicoplast.

PMID: 18586952 [PubMed - as supplied by publisher]

Immunodominant, protective response to the parasite Toxoplasma gondii requires antigen processing in the endoplasmic reticulum

Nat Immunol. 2008 Jun 29. [Epub ahead of print]

Immunodominant, protective response to the parasite Toxoplasma gondii requires antigen processing in the endoplasmic reticulum

Blanchard N, Gonzalez F, Schaeffer M, Joncker NT, Cheng T, Shastri AJ, Robey EA, Shastri N.

Division of Immunology and Pathogenesis, Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA.

The parasite Toxoplasma gondii replicates in a specialized intracellular vacuole and causes disease in many species. Protection from toxoplasmosis is mediated by CD8(+) T cells, but the T. gondii antigens and host genes required for eliciting protective immunity are poorly defined. Here we identified GRA6, a polymorphic protein secreted in the parasitophorous vacuole, as the source of the immunodominant and protective decapeptide HF10 presented by the H-2L(d) major histocompatibility complex class I molecule. Presentation of the HF10-H-2L(d) ligand required proteolysis by ERAAP, the endoplasmic reticulum aminopeptidase associated with antigen processing. Consequently, expansion of protective CD8(+) T cell populations was impaired in T. gondii-infected ERAAP-deficient mice, which were more susceptible to toxoplasmosis. Thus, endoplasmic reticulum proteolysis is critical for eliciting protective immunity to a vacuolar parasite.

PMID: 18587399 [PubMed - as supplied by publisher]

Efficient expression of a Toxoplasma gondii dense granule Gra4 antigen in tobacco leaves

Exp Parasitol. 2008 Jun 10. [Epub ahead of print]

Efficient expression of a Toxoplasma gondii dense granule Gra4 antigen in tobacco leaves

Ferraro G, Becher ML, Angel SO, Zelada A, Mentaberry AN, Clemente M.

IIB-INTECH, Camino Circunvalación Laguna km. 6, Chascomús, prov. de Bs. As 7130, Argentina.

A His-tagged truncated version of Toxoplasma gondii dense granule 4 protein (Gra4(163-345)) was transiently expressed in tobacco leaves. Two genetic constructions were used to accomplish this goal. In one of them, based in a Potato virus X (PVX) amplicon, the sequence encoding His-Gra4(163-345) was placed under control of an additional PVX coat protein subgenomic promoter. In the other, the same sequence was fused to an apoplastic transport signal and placed under the direction of the cauliflower mosaic virus 35S promoter. His-Gra4(163-345) accumulation in agroinfiltrated tobacco leaves was estimated by Western blot analysis using mouse anti-Gra4 antibody and a seropositive human serum. Here, we demonstrated the feasibility of producing a Gra4 antigen using transient expression methods in plants.

PMID: 18588877 [PubMed - as supplied by publisher]