Wednesday, October 29, 2008

Toxoplasma gondii: Identification and characterization of bradyzoite-specific deoxyribose phosphate aldolase-like gene (TgDPA)

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

Toxoplasma gondii: Identification and characterization of bradyzoite-specific deoxyribose phosphate aldolase-like gene (TgDPA)

Ueno A, Dautu G, Munyaka B, Carmen G, Kobayashi Y, Igarashi M.

National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, 2-13 Inada-cho, Obihiro, Hokkaido 080-8555, Japan.

Toxoplasma gondii undergoes stage conversion from tachyzoites to bradyzoites in intermediate hosts. There have been many reports on bradyzoite-specific genes which are thought to be involved in stage conversion. Here, we described a novel T. gondii deoxyribose phosphate aldolase-like gene (TgDPA) expressing predominantly in bradyzoites. The TgDPA gene encodes 286 amino acids having a predicted molecular weight of 31kDa. Sequence analysis revealed that TgDPA had a deoxyribose phosphate aldolase (DeoC) domain with about 30% homology with its Escherichia coli counterpart. RT- and quantitative PCR analyses showed that the TgDPA gene was more expressed in bradyzoites and that its expression gradually increased during in vitro tachyzoite-to-bradyzoite stage conversion. A polyclonal antibody against recombinant TgDPA protein was raised in rabbits, and immunofluorescent analysis demonstrated that TgDPA was expressed in bradyzoites in vivo and in vitro. These findings indicate that the TgDPA gene is a new bradyzoite-specific marker and might play a role in bradyzoites.

PMID: 18950626 [PubMed - as supplied by publisher]

Rhoptry neck protein RON2 forms a complex with microneme protein AMA1 in Plasmodium falciparum merozoites

Parasitol Int. 2008 Oct 7. [Epub ahead of print]

Rhoptry neck protein RON2 forms a complex with microneme protein AMA1 in Plasmodium falciparum merozoites

Cao J, Kaneko O, Thongkukiatkul A, Tachibana M, Otsuki H, Gao Q, Tsuboi T, Torii M.

Department of Molecular Parasitology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan; Malaria Department, Jiangsu Institute of Parasitic Diseases, Meiyuan, Wuxi, Jiangsu 214064, People's Republic of China.

Erythrocyte invasion is an essential step in the establishment of host infection by malaria parasites, and is a major target of intervention strategies that attempt to control the disease. Recent proteome analysis of the closely-related apicomplexan parasite, Toxoplasma gondii, revealed a panel of novel proteins (RONs) located at the neck portion of the rhoptries. Three of these proteins, RON2, RON4, and RON5 have been shown to form a complex with the microneme protein Apical Membrane Protein 1 (AMA1). This complex, termed the Moving Junction complex, localizes at the interface of the parasite and the host cell during the invasion process. Here we characterized a RON2 ortholog in Plasmodium falciparum. PfRON2 transcription peaked at the mature schizont stage and was expressed at the neck portion of the rhoptry in the merozoite. Co-immunoprecipitation of PfRON2, PfRON4 and PfAMA1 indicated that the complex formation is conserved between T. gondii and P. falciparum, suggesting that co-operative function of the rhoptry and microneme proteins is a common mechanism in apicomplexan parasites during host cell invasion. PfRON2 possesses a region displaying homology with the rhoptry body protein PfRhopH1/Clag, a component of the RhopH complex. However, here we present co-immunoprecipitation studies which suggest that PfRON2 is not a component of the RhopH complex and has an independent role. Nucleotide polymorphism analysis suggested that PfRON2 was under diversifying selective pressure. This evidence suggests that RON2 appears to have a fundamental role in host cell invasion by apicomplexan parasites, and is a potential target for malaria intervention strategies.

PMID: 18952195 [PubMed - as supplied by publisher]

Toxoplasma-safe meat: close to reality?

Trends Parasitol. 2008 Oct 23. [Epub ahead of print]

Toxoplasma-safe meat: close to reality?

Kijlstra A, Jongert E.

Animal Sciences Group, Wageningen UR, PO Box 65, 8200AB Lelystad, The Netherlands; Department of Ophthalmology, Maastricht University, Maastricht, The Netherlands.

In 2008, the centennial of the discovery of Toxoplasma gondii was celebrated. However, toxoplasmosis is still seen as a neglected and underreported disease, despite having a disease burden similar to that of salmonellosis and campylobacteriosis. Human vaccines are not available and current antiparasitic treatment is disappointing. This has led to an urge to focus more on prevention. Food, soil or water contaminated with oocysts from cat faeces and undercooked meat from infected intermediate hosts are important routes of infection. Oocyst contamination is difficult to control, whereas in Western countries, the control of T. gondii in meat should be feasible. Here, we discuss strategies aimed at developing a Toxoplasma-safe meat chain.

PMID: 18951847 [PubMed - as supplied by publisher]

Sunday, October 26, 2008

Host Cell Egress and Invasion Induce Marked Relocations of Glycolytic Enzymes in Toxoplasma gondii Tachyzoites

PLoS Pathog. 2008 Oct;4(10):e1000188. Epub 2008 Oct 24

Host Cell Egress and Invasion Induce Marked Relocations of Glycolytic Enzymes in Toxoplasma gondii Tachyzoites

Pomel S, Luk FC, Beckers CJ.

Department of Cell & Developmental Biology, University of North Carolina, Chapel Hill, North Carolina, United States of America.

Apicomplexan parasites are dependent on an F-actin and myosin-based motility system for their invasion into and escape from animal host cells, as well as for their general motility. In Toxoplasma gondii and Plasmodium species, the actin filaments and myosin motor required for this process are located in a narrow space between the parasite plasma membrane and the underlying inner membrane complex, a set of flattened cisternae that covers most the cytoplasmic face of the plasma membrane. Here we show that the energy required for Toxoplasma motility is derived mostly, if not entirely, from glycolysis and lactic acid production. We also demonstrate that the glycolytic enzymes of Toxoplasma tachyzoites undergo a striking relocation from the parasites' cytoplasm to their pellicles upon Toxoplasma egress from host cells. Specifically, it appears that the glycolytic enzymes are translocated to the cytoplasmic face of the inner membrane complex as well as to the space between the plasma membrane and inner membrane complex. The glycolytic enzymes remain pellicle-associated during extended incubations of parasites in the extracellular milieu and do not revert to a cytoplasmic location until well after parasites have completed invasion of new host cells. Translocation of glycolytic enzymes to and from the Toxoplasma pellicle appears to occur in response to changes in extracellular [K(+)] experienced during egress and invasion, a signal that requires changes of [Ca(2+)](c) in the parasite during egress. Enzyme translocation is, however, not dependent on either F-actin or intact microtubules. Our observations indicate that Toxoplasma gondii is capable of relocating its main source of energy between its cytoplasm and pellicle in response to exit from or entry into host cells. We propose that this ability allows Toxoplasma to optimize ATP delivery to those cellular processes that are most critical for survival outside host cells and those required for growth and replication of intracellular parasites.

PMID: 18949028 [PubMed - in process]

Toxoplasma gondii vs ionizing radiation

Rev Inst Med Trop Sao Paulo. 2008 Sep-Oct;50(5):264

Toxoplasma gondii vs ionizing radiation: cell and humoral immunity in spleen and gut of isogenic mice immunized with 60co irradiated tachyzoites

Galisteo AJ Jr.

Laboratório de Protozoologia, Instituto de Medicina Tropical de São Paulo, São Paulo, SP, Brasil,

PMID: 18949341 [PubMed - in process]

Neuronal injury in brains of mice due to common, persistent, parasitic infection

J Neuroinflammation. 2008 Oct 23;5(1):48. [Epub ahead of print]

Neurological and behavioral abnormalities, ventricular dilatation, altered cellular functions, inflammation, and neuronal injury in brains of mice due to common, persistent, parasitic infection

Hermes G, Ajioka JW, Kelly KA, Mui E, Roberts F, Kasza K, Mayr T, Kirisits MJ, Wollman R, Ferguson DJ, Roberts CW, Hwang JH, Trendler T, Kennan RP, Suzuki Y, Reardon C, Hickey AW, Chen L, McLeod R.

ABSTRACT: BACKGROUND: Worldwide, approximately two billion people are chronically infected with Toxoplasma gondii with largely unknown consequences. METHODS: To better understand long-term effects and pathogenesis of this common, persistent brain infection, mice were infected at a time in human years equivalent to early to mid adulthood and studied 5-12 months later. Appearance, behavior, neurologic function and brain MRIs were studied. Additional analyses of pathogenesis included: correlation of brain weight and neurologic findings; histopathology focusing on brain regions; full genome microarrays; immunohistochemistry characterizing inflammatory cells; determination of presence of tachyzoites and bradyzoites; electron microscopy; and study of markers of inflammation in serum. Histopathology in genetically resistant mice and cytokine and NRAMP knockout mice, effects of inoculation of isolated parasites, and treatment with sulfadiazine or alphaPD1 ligand were studied. RESULTS: Twelve months after infection, a time equivalent to middle to early elderly ages, mice had behavioral and neurological deficits, and brain MRIs showed mild to moderate ventricular dilatation. Lower brain weight correlated with greater magnitude of neurologic abnormalities and inflammation. Full genome microarrays of brains reflected inflammation causing neuronal damage (Gfap), effects on host cell protein processing (ubiquitin ligase), synapse remodeling (Complement 1q), and also increased expression of PD-1L (a ligand that allows persistent LCMV brain infection) and CD 36 (a fatty acid translocase and oxidized LDL receptor that mediates innate immune response to beta amyloid which is associated with pro-inflammation in Alzheimer's disease). Immunostaining detected no inflammation around intra-neuronal cysts, practically no free tachyzoites, and only rare bradyzoites. Nonetheless, there were perivascular, leptomeningeal inflammatory cells, particularly contiguous to the aqueduct of Sylvius and hippocampus, CD4+ and CD8+ T cells, and activated microglia in perivascular areas and brain parenchyma. Genetically resistant, chronically infected mice had substantially less inflammation. CONCLUSION: In outbred mice, chronic, adult acquired T. gondii infection causes neurologic and behavioral abnormalities secondary to inflammation and loss of brain parenchyma. Perivascular inflammation is prominent particularly contiguous to the aqueduct of Sylvius and hippocampus. Even resistant mice have perivascular inflammation. This mouse model of chronic T. gondii infection raises questions of whether persistence of this parasite in brain can cause inflammation or neurodegeneration in genetically susceptible hosts.

PMID: 18947414 [PubMed - as supplied by publisher]

Friday, October 24, 2008

Stem cell transplantation in hematological disorders

Saudi Med J. 2008 Oct;29(10):1484-9

Stem cell transplantation in hematological disorders. A developing country experience-impact of cost considerations

El-Zimaity MM, Hassan EA, Abd El-Wahab SE, Abd El-Ghaffar AA, Mahmoud NA, Elafifi AM, Moussa MM, El-Gohary GM, Hagras MM, Mowafy NM.

10 elsheik Ahmed eldardiry st apt # 303, Golfland Cairo, 11361 Egypt. Tel. +202 22906848/+202 (12) 7460920. Fax. +202 25880592. E-mail:

OBJECTIVE: To describe the experience in setting up a bone marrow transplant program at Ain Shams University, Cairo, Egypt. METHODS: Sixteen patients were transplanted at Ain Shams University Bone Marrow Transplantation unit from March 2005 to January 2008. RESULTS: Sixteen patients were transplanted with a median age of 25 years. Indications for transplantation were chronic myeloid leukemia, acute myeloid leukemia, aplastic anemia, acute lymphoblastic leukemia, and aggressive lymphoma. Seven donors and 6 patients were positive for cytomegalovirus immunoglobulin G (IgG) antibody (Ab) pretransplant. Only one patient was positive for toxoplasma IgG Ab and another had a high titre for toxoplasma IgM Ab pretransplant. Two donors and 2 recipients were positive for hepatitis B antibody markers; however, none were positive for hepatitis B virus DNA by polymerase chain reaction (PCR). None of the patients or donors were positive for hepatitis C virus via PCR pre-transplant. Acute graft versus host disease (GVHD) was seen in 3 patients, while chronic GVHD was seen in 5 patients. Primary cause of death was recurrence in 2 patients and graft failure in one patient. Thirteen are alive and disease free with a median follow-up of 20 months. CONCLUSION: Although our unit is a relatively new unit, these results are comparable to those achieved in the Western world and cost a mean of US$250,000.

PMID: 18946578 [PubMed - in process]

Infection in childhood sensory hearing loss

Saudi Med J. 2008 Oct;29(10):1470-4

Infection in childhood sensory hearing loss

Noorbakhsh S, Farhadi M, Tabatabaei A, Mohamadi S, Jomeh E.

Research Center of Pediatric Infectious Diseases, 8th floor Hazrat Rasul Hospital, Niayesh Street, Satarkhan Avenue, Tehran 14455, Iran.Tel. +98 (21) 66516049. Fax. +98 (21) 66525328/66516049. E-mail:

OBJECTIVE: To evaluate the possible role of infectious agents on the occurrence of idiopathic sensorineural hearing loss (SNHL) in children. METHODS: This case control study was carried out at Rasul Hospital in Tehran, Iran from 2002-2003. We compared specific serum antibodies (IgG, IgM) measured by enzyme linked immunosorbent assay in 95 sensory hearing loss cases and 63 controls. RESULTS: Acute infections (IgM) detected in cases included: cytomegalovirus (CMV) 34.6%, toxoplasma 11.5%, mumps 8.7%, rubella 5.3%, and herpes simplex 5.3%. Previous infections (IgG) detected in cases included CMV 72%, herpes 6.6%, toxoplasma 26%, mumps 23.3%, and rubella 17.2%. Acute CMV and toxoplasma infections were more frequent in cases. Previous CMV, toxoplasma, rubella, and herpes infections were higher in controls. There was no significant difference for acute mumps, rubella, and herpes infections between cases and controls. CONCLUSION: These data are compatible with infectious agents having a significant role in the studied idiopathic SNHL cases, but association does not prove causation. We recommend specific drugs for confirmed active infections (CMV, toxoplasma, herpes) in idiopathic SNHL infants diagnosed before their first birthday. Mumps and rubella induced SNHL are preventable with routine vaccination.

PMID: 18946575 [PubMed - in process]

Role of intermonomer ionic bridges in the stabilization of the actin filament

J Biol Chem. 2008 Oct 21. [Epub ahead of print]

Role of intermonomer ionic bridges in the stabilization of the actin filament

Stokasimov E, McKane M, Rubenstein PA.

Biochemistry, University of Iowa Carver College of Medicine, Iowa City, IA 52242.

Filament formation is required for most of actin's functions. However, the intermonomer interactions that stabilize F-actin have not been elucidated because of a lack of an F-actin crystal structure. The Holmes muscle actin model suggests that an ionic interaction between R39 of one monomer and E167 of an adjacent monomer in the same strand contributes to this stabilization. Yeast actin has an A167 instead. F-actin molecular dynamics modeling predicts another interaction between R39 of one monomer and D275 of an opposing strand monomer. In Toxoplasma gondii actin, which forms short stubby filaments, the D275 equivalent is replaced by R leading to a potential filament-destabilizing charge-charge repulsion. Using yeast actin, we tested the effect of A167E as a potential stabilizer and A167R and D275R as potential filament disruptors. All mutations caused abnormal growth and mitochondrial malfunction. A167E and D275R actins polymerize normally and form relatively normal appearing filaments. A167R nucleates filaments more slowly and forms filament bundles. The R39D/A167R double mutant, which reestablishes an ionic bond in the opposite orientation reverses this polymerization and bundling defect. Stoichiometric amounts of yeast cofilin have little effect on WT and A167E filaments. However, D275R and A167R actin depolymerization is profound with cofilin. While our results suggest that disruption of an interaction between R39 and D275 is not sufficient to cause fragmentation, it suggests that it changes filament stability thereby disposing it for enhanced cofilin depolymerizing effects. A167 results demonstrate the in vivo and in vitro importance of another potential R39 ionic interaction.

PMID: 18945676 [PubMed - as supplied by publisher]

Wednesday, October 22, 2008

The complexity of signaling in host-pathogen interactions revealed by the Toxoplasma gondii-dependent modulation of JNK phosphorylation

Exp Cell Res. 2008 Oct 7. [Epub ahead of print]

The complexity of signaling in host-pathogen interactions revealed by the Toxoplasma gondii-dependent modulation of JNK phosphorylation

Carmen JC, Southard RC, Sinai AP.

Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky College of Medicine, 800 Rose Street, Lexington, KY 40536, USA.

The inhibition of apoptosis by Toxoplasma gondii is governed by its modulation of several signaling cascades including the NFkappaappaB and JNK pathways. This is evident in the dysregulation of JNK activation following treatment with UV and TNFalpha, both apoptogenic stimuli. Infection-mediated interference with the JNK cascade was found to be highly reproducible in HeLa cells. In light of emerging evidence regarding cross talk between the JNK and NFkappaB cascades, we examined the impact of infection in wild type and RelA/p65-/- mouse embryonic fibroblasts (MEF). Remarkably, parasite infection failed to significantly impact both UV and TNFalpha-mediated JNK phosphorylation in both cell lines suggesting a cell type specific effect. Furthermore siRNA-mediated knockdown of RelA/p65 failed to impact the parasite mediated effects on stimulus dependent activation of JNK in HeLa cells. Finally, the infection mediated suppression of JNK phosphorylation in HeLa cells did not result in decreased JNK kinase activity. Rather, the reduced levels of phospho-JNK in infected cells correlated with increased phosphatase activity noted by the partial rescue of the phenotype following treatment with okadaic acid. Taken together the results indicate that manipulation of the JNK pathway does not involve NFkappaB and is furthermore not a central component of the parasite enforced block of apoptosis. It further highlights the complexity of these systems and the danger of extrapolating results both within and across pathogen-host cell systems based on limited studies.

PMID: 18929560 [PubMed - as supplied by publisher]

Immunoregulation by Toxoplasma gondii infection prevents allergic immune responses in mice

Int J Parasitol. 2008 Oct 2. [Epub ahead of print]

Immunoregulation by Toxoplasma gondii infection prevents allergic immune responses in mice

Wagner A, Förster-Waldl E, Garner-Spitzer E, Schabussova I, Kundi M, Pollak A, Scheiner O, Joachim A, Wiedermann U.

Department of Specific Prophylaxis and Tropical Medicine, Center for Physiology, Pathophysiology & Immunology, Medical University of Vienna, Kinderspitalgasse 15, 1090 Vienna, Austria.

Toxoplasma gondii is a ubiquitous intracellular parasite affecting most mammals including humans. In epidemiological studies, infection with T. gondii and allergy development have been postulated to be inversely related. Using a mouse model of birch pollen allergy we investigated whether infection with T. gondii influences allergic immune responses to birch pollen. BALB/c mice were infected with T. gondii oocysts either before or at the end of sensitisation with the major birch pollen allergen Bet v 1 and thereafter aerosol challenged with birch pollen extract. During the acute phase of infection, clinical signs correlated with increased levels of serum TNF-alpha, IL-6, IFN-gamma and anti-Toxoplasma-IgM. In the chronic phase, Toxoplasma-specific serum IgG, brain tissue cysts and high IFN-gamma production in spleen cell cultures were detected. Mice infected prior to allergic sensitisation produced significantly less allergen-specific IgE and IgG1, while IgG2a levels were markedly increased. IL-5 levels in spleen cell cultures and bronchoalveolar lavage fluid were significantly reduced, and airway inflammation was prevented in these mice. Notably, in mice infected at the end of the allergic sensitisation process, systemic and local immune responses to the allergen were markedly reduced. T.gondii infection was associated with up-regulation of Toll-like receptor 2 (TLR2), 4, 9 and 11, as well as T-bet (a differentiation factor for Th1 cells) mRNA expression in splenocytes; moreover, enhanced TGF-beta, IL-10 and Foxp3 mRNA expression in these cells suggested that regulatory mechanisms were involved in suppression of the allergic immune response. Kinetic studies confirmed the induction of Foxp3(+)CD4(+)CD25(+) regulatory T cells preferentially during the chronic phase of T. gondii infection. Our data demonstrate that T. gondii exhibits strong immunomodulating properties which lead to prevention of allergic immune responses and thereby support the hygiene hypothesis.

PMID: 18938169 [PubMed - as supplied by publisher]

Saturday, October 18, 2008

Parasite Stage-Specific Recognition of Endogenous Toxoplasma gondii-Derived CD8(+) T Cell Epitopes

J Infect Dis. 2008 Oct 15. [Epub ahead of print]

Parasite Stage-Specific Recognition of Endogenous Toxoplasma gondii-Derived CD8(+) T Cell Epitopes

Frickel EM, Sahoo N, Hopp J, Gubbels MJ, Craver MP, Knoll LJ, Ploegh HL, Grotenbreg GM.

1Whitehead Institute for Biomedical Research, Cambridge, 2Department of Biology, Boston College, Chestnut Hill, Massachusetts; 3Department of Medical Microbiology and Immunology, University of Wisconsin Medical School, Madison, Wisconsin.

Background. @nbsp; BALB/c mice control infection with the obligate intracellular parasite Toxoplasma gondii and develop a latent chronic infection in the brain, as do immunocompetent humans. Interferon-gamma-producing CD8(+) T cells provide essential protection against T. gondii infection, but the epitopes recognized have so far remained elusive. Methods. @nbsp; We employed caged major histocompatibility complex molecules to generate 250 H-2L(d) tetramers and to distinguish T. gondii-specific CD8(+) T cells in BALB/c mice. Results. @nbsp; We identified 2 T. gondii-specific H-2L(d)-restricted T cell epitopes, one from dense granule protein GRA4 and the other from rhoptry protein ROP7. H-2L(d)/GRA4 reactive T cells from multiple organ sources predominated 2 weeks after infection, while the reactivity of the H-2L(d)/ROP7 T cells peaked 6-8 weeks after infection. BALB/c animals infected with T. gondii mutants defective in establishing a chronic infection showed altered levels of antigen-specific T cells, depending on the T. gondii mutant used. Conclusions. @nbsp; Our results shed light on the identity and the parasite stage-specificity of 2 CD8(+) T cell epitopes recognized in the acute and chronic phase of infection with T. gondii.

PMID: 18922097 [PubMed - as supplied by publisher]

Thursday, October 16, 2008

Toxoplasma gondii antibody profile in HIV-infected pregnant women and the risk of congenital toxoplasmosis

Eur J Clin Microbiol Infect Dis. 2008 Oct 15. [Epub ahead of print]

Toxoplasma gondii antibody profile in HIV-infected pregnant women and the risk of congenital toxoplasmosis

Lago EG, Conrado GS, Piccoli CS, Carvalho RL, Bender AL.

Department of Pediatrics, Pontifícia Universidade Católica do Rio Grande do Sul-School of Medicine, Av. Ipiranga 6690, Porto Alegre, 90610, Rio Grande do Sul, Brazil,

The purpose of this study was to investigate the antibodies to Toxoplasma gondii in human immunodeficiency virus (HIV)-infected pregnant women and to determine the association between serological profile and the risk of congenital toxoplasmosis. The study, conducted in a public maternity ward from May 2002 to April 2005, included all HIV-infected women who delivered live infants during the 36 months, and, as a control group, all HIV-negative women that delivered live infants in the first 12 months of the study. Antibodies to T. gondii were detected in 1,624 of 2,421 HIV-negative women (67%; 95% confidence interval [CI] 65-69%) and in 121 of 168 HIV-infected patients (72%; 95% CI 65-79%). A total of 547 HIV-negative and 103 HIV-infected patients were tested at delivery and had positive T. gondii-specific IgG. In HIV-negative women, the median of the specific IgG concentration was 79 (interquartile range 38-160), and in HIV-infected patients, it was 283 (interquartile range 94-704) (P < 0.001). In the group of co-infected women, the only infant with congenital toxoplasmosis was born to a mother with acute toxoplasmosis infection acquired during pregnancy who did not have a high specific IgG concentration or a positive result for specific IgM. We concluded that high T. gondii-specific IgG values were much more frequent among HIV-infected pregnant women, but it did not translate into an increased risk of maternal-fetal transmission of toxoplasmosis.

PMID: 18855029 [PubMed - as supplied by publisher]

Genotyping of Toxoplasma gondii strains from female patients with toxoplasmosis

J Egypt Soc Parasitol. 2008 Aug;38(2):511-20

Genotyping of Toxoplasma gondii strains from female patients with toxoplasmosis

Abdel-Hameed DM, Hassanein OM.

Department of Parasitology, Faculty of Medicine, Ain-Shams University, Cairo 11566, Egypt.

Thirty-eight female patients with abortion and intrauterine fetal death were selected from the Obstetric and Gynecology Emergency, Ain Shams University Hospitals, with positive PCR results for toxoplasmosis in previous study. In this study, a rapid and efficient procedure was used for genotyping of T. gondii isolates based on PCR-RFLP assay at SAG2 locus. On the basis of the alleles identified at SAG2 locus, the isolates were grouped into three lineages. Type I was determined by resistance of the 3' & 5' end nested product of the SAG2 locus to cleavage by HhaI & Sau3AI respectively. Resistance of 5' end of SAG2 locus to cleavage by HhaI determined type II. Type III was determined by resistance of the 3' end nested-PCR products of SAG2 locus to cleavage by Sau3AI. Of the 38 isolates, type II was the most prevalent genotype found in 33 (87%). Type I was found in 5 (13%) of the isolates, whereas genotype III was not never found.

PMID: 18853624 [PubMed - in process]

Phylogenetic evidence for recombination in SAG5 locus in Toxoplasma gondii

J Egypt Soc Parasitol. 2008 Aug;38(2):371-84

Phylogenetic evidence for recombination in SAG5 locus in Toxoplasma gondii

Elsheikha H, Hafez AO, Zaalouk TK, El Shazly AM, Khalil HH, Morsy TA.

Division of Veterinary Medicine, The School of Veterinary Medicine and Science, The University of Nottingham, College Road, Sutton Bonington Campus, Loughborough, Leicestershire, LE12 5RD, UK.

The present study evaluated the hypothesis that genetic diversity in SAG5 genes was generated by recombination events. Three lines of evidence suggested that recombination occurred in SAG5 genes in T. gondii. The permutation test revealed strong signature of intragenic recombination, pairwise comparisons of nucleotide sequences of SAG5 genes revealed that SAG5A alleles have chimerical structures composed of segments derived through recombination events between different alleles, and phylogenetic trees reconstructed based on SAG5 sequences using neighbor-joining and maximum parsimony methods, showed statistically well-supported consensus clusters of T. gondii strains specific to each SAG5 gene. Topological discrepancies between trees based on the N-terminal variable domain and C-terminal conserved domain sequences, were observed, suggesting intragenic recombinetion between SAG5A and SAG5B/C genes. The results showed that recombination within SAG5 in T. gondii was a major evolutionary mechanism generating both allelic variation at SAG5 locus and contributing to genotypic diversity and to emergence of new T. gondii variants, allowing them to evade the host immune defence mechanism.

PMID: 18853612 [PubMed - in process]

Thursday, October 09, 2008

NSC3852, a redox active anti-proliferative and tumor cell differentiation agent against Toxo

J Parasitol. 2008 Oct 6:1. [Epub ahead of print]

Inhibition of Toxoplasma gondii and Plasmodium falciparum infections in vitro by NSC3852, a redox active anti-proliferative and tumor cell differentiation agent

Strobl JS, Seibert CW, Li Y, Nagarkatti R, Mitchell SM, Rosypal AC, Rathore D, Lindsay DS.

We searched the National Cancer Institute (NCI) compound library for structures related to the anti-tumor quinoline NSC3852 (5-nitroso-8-quinolinol) and used a computer algorithm to predict the anti-protozoan activity of each of 13 structures. Half of these compounds inhibited T. gondii tachyzoite propagation in human fibroblasts at
PMID: 18837587 [PubMed - as supplied by publisher]

Toxoplasma induced foetal resorption in mice involves interferon-gamma-induced apoptosis and spiral artery dilation at the maternofoetal interface

Int J Parasitol. 2008 Sep 22. [Epub ahead of print]

Toxoplasma gondii-induced foetal resorption in mice involves interferon-gamma-induced apoptosis and spiral artery dilation at the maternofoetal interface

Senegas A, Villard O, Neuville A, Marcellin L, Pfaff AW, Steinmetz T, Mousli M, Klein JP, Candolfi E.

Institut de Parasitologie et de Pathologie Tropicale, UPRES-EA 3950, Université Louis Pasteur, Strasbourg F-67000, France.

The severity of congenital toxoplasmosis depends on the stage of the pregnancy at which infection takes place. Infection during the first trimester generally leads to miscarriage, through an unknown mechanism. Toxoplasma gondii infection is normally controlled by a strong Th1-type response with IFN-gamma production. To investigate the mechanisms of foetal resorption induced by T. gondii, pregnant Swiss-Webster mice were infected 1 day post coïtum with the avirulent Me49 strain. Mated recipients were examined at mid-gestation. Few parasites and no cytolytic effects were detected 10 days post coïtum in implantation sites undergoing resorption. Resorption was accompanied by haemorrhage, spiral artery dilation, hypocellularity of the decidua basalis, apoptosis of placental cells, a decline in uterine mature natural killer cell numbers, increased indoleamine 2,3-dioxygenase mRNA levels and reduced IL-15 mRNA levels. Given the role of IFN-gamma in non-infectious abortive processes, IFN-gammaR(-/-) mice were used to investigate its local role in T. gondii-induced foetal resorption. IFN-gammaR(-/-) mice showed 50% less foetal resorption than their wild-type counterparts, and spiral artery dilation and placental cell apoptosis were both abolished. These results strongly suggest that, at least in mice, T. gondii-induced abortion in early gestation is not due to a direct action of the parasite at the maternofoetal interface but rather to massive IFN-gamma release.

PMID: 18838076 [PubMed - as supplied by publisher]

GRA12, a Toxoplasma dense granule protein associated with the intravacuolar membranous nanotubular network

Int J Parasitol. 2008 Sep 20. [Epub ahead of print]

GRA12, a Toxoplasma dense granule protein associated with the intravacuolar membranous nanotubular network

Michelin A, Bittame A, Bordat Y, Travier L, Mercier C, Dubremetz JF, Lebrun M.

INSERM, UMR 55235 CNRS, Université de Montpellier 2, CP 107, Place Eugène Bataillon, 34090 Montpellier, France.

The intracellular protozoan parasite Toxoplasma gondii develops within the parasitophorous vacuole (PV), an intracellular niche in which it secretes proteins from secretory organelles named dense granules and rhoptries. Here, we describe a new dense granule protein that should now be referred to as GRA12, and that displays no homology with other proteins. Immunofluorescence and immuno-electron microscopy showed that GRA12 behaves similarly to both GRA2 and GRA6. It is secreted into the PV from the anterior pole of the parasite soon after the beginning of invasion, transits to the posterior invaginated pocket of the parasite where a membranous tubulovesicular network is first assembled, and finally resides throughout the vacuolar space, associated with the mature membranous nanotubular network. GRA12 fails to localise at the parasite posterior end in the absence of GRA2. Within the vacuolar space, like the other GRA proteins, GRA12 exists in both a soluble and a membrane-associated form. Using affinity chromatography experiments, we showed that in both the parasite and the PV soluble fractions, GRA12 is purified with the complex of GRA proteins associated with a tagged version of GRA2 and that this association is lost in the PV membranous fraction.

PMID: 18840447 [PubMed - as supplied by publisher]

Wednesday, October 08, 2008

Host-cell invasion by malaria parasites: insights from Plasmodium and Toxoplasma

Trends Parasitol. 2008 Oct 1. [Epub ahead of print]

Host-cell invasion by malaria parasites: insights from Plasmodium and Toxoplasma

Baum J, Gilberger TW, Frischknecht F, Meissner M

Division of Infection and Immunity, Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria 3050, Australia.

Recent years have seen tremendous progress in our understanding of malaria parasite molecular biology. To a large extent, this progress follows significant developments in genetic, molecular and chemical tools available to study the malaria parasites and related Apicomplexa, in particular Toxoplasma gondii. One area of major advancement has been in understanding parasite host-cell invasion, a process that utilizes several essential molecular mechanisms that are conserved across the different lifecycle stages. Here, we summarize some of the most recent experimental data that shed light on the events underlying preparation and execution of malaria parasite invasion and how these insights might relate to the development of new antimalarial drugs.

PMID: 18835222 [PubMed - as supplied by publisher]

Double-stranded RNA specific to adenosine kinase and hypoxanthine-xanthine-guanine-phosphoribosyltransferase retards growth of Toxoplasma

Parasitol Res. 2008 Oct 3. [Epub ahead of print]

Double-stranded RNA specific to adenosine kinase and hypoxanthine-xanthine-guanine-phosphoribosyltransferase retards growth of Toxoplasma gondii

Yu L, Gao YF, Li X, Qiao ZP, Shen JL.

Department of Pathogen biology, Anhui Medical University, Hefei, Anhui, 230032, People's Republic of China.

Adenosine kinase (AK) and hypoxanthine-xanthine-guanine phosphoribosyltransferase (HXGPRT) are the two key enzymes involved in the purine salvage pathway in Toxoplasma gondii. In this study, we targeted the genes encoding AK and HXGPRT in T. gondii for inhibition by exposing the parasites to the corresponding double stranded RNAs (dsRNAs). We report here that dsRNAs targeting both AK and HXGPRT were effective at eliciting suppression of the corresponding gene in cultured tachyzoites. When both of these genes were targeted by dsRNA simultaneously, the average doubling for the dsRNA-treated parasites at 24 h, 30 h, and 42 h was 1.85, 2.62, and 4.27, respectively, significantly lower than that of mock-treated parasites. The data show that transfection of dsRNAs into cells can efficiently regulate gene expression in T. gondii. Application of dsRNA to disrupt gene expression in T. gondii would be useful for elucidating gene function as a step towards the development of therapeutic reagents.

PMID: 18836745 [PubMed - as supplied by publisher]

Sunday, October 05, 2008

Restoration of Toxoplasma gondii-specific immune responses in patients with AIDS starting HAART

AIDS. 2008 Oct 18;22(16):2087-96

Restoration of Toxoplasma gondii-specific immune responses in patients with AIDS starting HAART

Furco A, Carmagnat M, Chevret S, Garin YJ, Pavie J, De Castro N, Charron D, Derouin F, Rabian C, Molina JM.

Department of Infectious Diseases, Saint-Louis Hospital, Assistance Publique Hôpitaux de Paris and University of Paris 7, Paris Diderot, Paris, France.

OBJECTIVE: To study the kinetics and identify factors associated with Toxoplasma-specific immune responses in patients with AIDS starting antiretroviral therapy. METHODS: A prospective study of 38 HIV-infected patients seropositive for Toxoplasma who started antiretroviral therapy with CD4 T cells less than 200 cells/microl. T-cell and B-cell phenotypes, anti-Toxoplasma antibodies titers, Th-1 and Th-2 cytokine production and lymphocyte proliferative responses (LPRs) to Toxoplasma were assessed over 12 months. RESULTS: Median CD4 cell count increased from 122 cells/microl at baseline to 260 cells/microl at 12 months, and the incidence of a positive LPR from 18.4 to 70.5%. A Toxoplasma IgG titer more than 150 IU/ml was the only baseline variable associated with a positive LPR (hazard ratio: 4.6, P = 0.003). Among time-dependent covariates, the number of effector memory (CD45RA-CCR7-) CD4 T cells was associated with a positive LPR (P < 0.02) and the number of terminally differentiated (CD45RA+CCR7-) CD8 T cells was associated with in-vitro production of gamma-IFN (P < 0.008). CONCLUSION: Among patients with low CD4 cell counts, high anti-Toxoplasma IgG titers were associated with LPR to Toxoplasma antigen. After starting antiretroviral therapy, the number of effector memory CD4 T cells and terminally differentiated CD8 T cells were associated with the restoration of Toxoplasma LPR and gamma-IFN production, respectively.

Publication Types:
Research Support, Non-U.S. Gov't

PMID: 18832872 [PubMed - in process]

Friday, October 03, 2008

Suppressed Production of Pro-inflammatory Cytokines by LPS-Activated Macrophages after Treatment with Toxoplasma gondii Lysate

Korean J Parasitol. 2008 Sep;46(3):145-51

Suppressed Production of Pro-inflammatory Cytokines by LPS-Activated Macrophages after Treatment with Toxoplasma gondii Lysate

Lee EJ, Heo YM, Choi JH, Song HO, Ryu JS, Ahn MH.

Department of Parasitology and Institute of Biomedical Science, Hanyang University College of Medicine, Seoul 133-791, Korea.

During Toxoplasma gondii infection, macrophages, dendritic cells, and neutrophils are important sources of pro-inflammatory cytokines from the host. To counteract the pro-inflammatory activities, T. gondii is known to have several mechanisms inducing down-regulation of the host immunity. In the present study, we analyzed the production of proand anti-inflammatory cytokines from a human myelomonocytic cell line, THP-1 cells, in response to treatment with T. gondii lysate or lipopolysaccharide (LPS). Treatment of THP-1 cells with LPS induced production of IL-12, TNF-alpha, IL-8, and IL-10. Co-treatment of THP-1 cells with T. gondii lysate inhibited the LPS-induced IL-12, IL-8 and TNF-alpha expression, but increased the level of IL-10 synergistically. IL-12 and IL-10 production was down-regulated by anti-human toll-like receptor (TLR)-2 and TLR4 antibodies. T. gondii lysate triggered nuclear factor (NF)-kappaB-dependent IL-8 expression in HEK293 cells transfected with TLR2. It is suggested that immunosuppression induced by T. gondii lysate treatment might occur via TLR2-mediated NF-kappaB activation.

PMID: 18830053 [PubMed - in process]

Thursday, October 02, 2008

Inhibition of Toxoplasma gondii by Indirubin and Tryptanthrin Analogs

Antimicrob Agents Chemother. 2008 Sep 29. [Epub ahead of print]

Inhibition of Toxoplasma gondii by Indirubin and Tryptanthrin Analogs

Krivogorsky B, Grundt P, Yolken R, Jones-Brando L.

Stanley Laboratory, Department of Pediatrics, Johns Hopkins School of Medicine, Baltimore Md; Department of Chemistry and Biochemistry, University of Minnesota Duluth, Duluth Minn.

New drugs are needed for treatment of Toxoplasma gondii infections. We tested derivatives of principles found in Isatis indigotica for in vitro efficacy against T. gondii. Indirubin-3'-oxime analogs showed modest micromolar activity, while tryptanthrin derivatives displayed ID50s in low nanomolar range. Tryptanthrins have potential as anti-Toxoplasma therapeutics.

PMID: 18824607 [PubMed - as supplied by publisher]

Effects of neuwiedase, a metalloproteinase from snake venom, on the invasion and replication of fibroblasts

Exp Parasitol. 2008 Sep 14. [Epub ahead of print]

Toxoplasma gondii: Effects of neuwiedase, a metalloproteinase from Bothrops neuwiedi snake venom, on the invasion and replication of human fibroblasts in vitro

Bastos LM, Júnior RJ, Silva DA, Mineo JR, Vieira CU, Teixeira DN, Homsi-Brandeburgo MI, Rodrigues VM, Hamaguchi A.

Laboratory of Protein Chemistry and Natural Products, Institute of Genetic and Biochemistry, Federal University of Uberlândia, Av. Pará, 1720, 38400-902 Uberlândia, MG, Brazil.

The major aim to the present study was to determine the effects of neuwiedase, a metalloproteinase isolated from Bothrops neuwiedi snake venom, on invasion and replication of Toxoplasma gondii in human fibroblasts in vitro. Neuwiedase treatment was done on host cells previously infected with T. gondii or on parasite before fibroblast infection. When treatments were done after or before infection, infection rates were inhibited in 71% and 61%, respectively. Considering that therapy protocols currently used in T. gondii infection cause considerable side effects, particularly in immunocompromised individuals and pregnant women, the results of neuwiedase treatment described herein could be taken into account for the development of new synthetic therapeutic agents, mainly due to the capacity of this enzyme to degrade extracellular matrix components, such as laminin, fibronectin and type I collagen, which is important to interfere in T. gondii host cell invasion.

PMID: 18823981 [PubMed - as supplied by publisher]

Wednesday, October 01, 2008

Evolution of the apicoplast and its hosts: From heterotrophy to autotrophy and back again

Int J Parasitol. 2008 Sep 12. [Epub ahead of print]

Evolution of the apicoplast and its hosts: From heterotrophy to autotrophy and back again

Oborník M, Janouškovec J, Chrudimský T, Lukeš J

Biology Centre, Institute of Parasitology, Academy of Sciences of the Czech Republic, University of South Bohemia, Faculty of Sciences, Branisovská 31, 370 05, Ceské Budejovice, Czech Republic.

The photosynthetic origin of apicomplexan parasites was proposed upon the discovery of a reduced non-photosynthetic plastid termed the apicoplast in their cells. Although it is clear that the apicoplast has evolved through a secondary endosymbiosis, its particular origin within the red or green plastid lineage remains controversial. The recent discovery of Chromera velia, the closest known photosynthetic relative to apicomplexan parasites, sheds new light on the evolutionary history of alveolate plastids. Here we review our knowledge on the evolutionary history of Apicomplexa and particularly their plastids, with a focus on the pathway by which they evolved from free-living heterotrophs through photoautotrophs to omnipresent obligatory intracellular parasites. New sequences from C. velia (histones H2A, H2B; GAPDH, TufA) and phylogenetic analyses are also presented and discussed here.

PMID: 18822291 [PubMed - as supplied by publisher]

The apicomplexan whole genome phylogeny: an analysis of incongruence among gene trees

Mol Biol Evol. 2008 Sep 26. [Epub ahead of print]

The apicomplexan whole genome phylogeny: an analysis of incongruence among gene trees

Kuo CH, Wares JP, Kissinger JC.

Department of Genetics.

The protistan phylum Apicomplexa contains many important pathogens and is the subject of intense genome sequencing efforts. Based upon the genome sequences from seven apicomplexan species and a ciliate outgroup, we identified 268 single-copy genes suitable for phylogenetic inference. Both concatenation and consensus approaches inferred the same species tree topology. This topology is consistent with most prior conceptions of apicomplexan evolution based upon ultrastructural and developmental characters, i.e.: the piroplasm genera Theileria and Babesia form the sister group to the Plasmodium; the coccidian genera Eimeria and Toxoplasma are monophyletic and are the sister group to Plasmodium and piroplasms; and Cryptosporidium forms the sister group to the above-mentioned with the ciliate Tetrahymena as the outgroup. The level of incongruence among gene trees appears to be high at first glance; only 19% of the genes support the species tree and a total of 48 different gene tree topologies are observed. Detailed investigations suggest that the low signal-to-noise ratio in many genes may be the main source of incongruence. The probability of being consistent with the species tree increases as a function of the minimum bootstrap support observed at tree nodes for a given gene tree. Moreover, gene sequences that generate high bootstrap support are robust to the changes in alignment parameters or phylogenetic method used. However, caution should be taken that some genes can infer a "wrong" tree with strong support because of paralogy, model violations, or other causes. The importance of examining multiple, unlinked genes that possess a strong phylogenetic signal cannot be overstated.

PMID: 18820254 [PubMed - as supplied by publisher]