Saturday, December 29, 2007

N-linked glycosylation of proteins in the protozoan parasite Toxoplasma

Mol Biochem Parasitol. 2007 Nov 9 [Epub ahead of print]

N-linked glycosylation of proteins in the protozoan parasite Toxoplasma gondii

Luk FC, Johnson TM, Beckers CJ

Department of Cell & Developmental Biology, University of North Carolina, Chapel Hill, NC 27599-7090, USA.

Toxoplasma gondii is an obligate intracellular parasite of animal cells. Infection of humans is common and may result in devastating disease, especially in immunocompromised individuals. Despite previous reports that N-glycosylation of proteins may be a rare post-translational modification in this and related organisms, we demonstrate that it is actually quite prevalent in Toxoplasma. N-Glycosylation is completely inhibited by treatment of parasites with tunicamycin, but this does not appear to exert its major effect on the parasites until they have egressed from their host cells. Although the tunicamycin-treated parasites appear structurally normal at this time they are not motile and mostly incapable of invading new host cells. The few tunicamycin-treated parasites that do invade are severely affected in their ability to replicate and accumulate with a distended endoplasmic reticulum, deformed nuclei, and without recognizable late secretory organelles. We provide experimental evidence that indicate that Toxoplasma N-glycans differ structurally from those in other eukaryotes.

PMID: 18096254 [PubMed - as supplied by publisher]

First isolate of Toxoplasma gondii from arctic fox

Vet Parasitol. 2007 Nov 17 [Epub ahead of print]

First isolate of Toxoplasma gondii from arctic fox (Vulpes lagopus) from Svalbard

Prestrud KW, Dubey JP, Asbakk K, Fuglei E, Su C

Norwegian School of Veterinary Science (NVH), Section of Arctic Veterinary Medicine, Stakkevollveien 23, N-9010, Norway.

Cats are considered essential for the maintenance of Toxoplasma gondii in nature. However, T. gondii infection has been reported in arctic fox (Vulpes lagopus) from the Svalbard high arctic archipelago where felids are virtually absent. To identify the potential source of T. gondii, we attempted to isolate and genetically characterize the parasite from arctic foxes in Svalbard. Eleven foxes were trapped live in Grumant (78 degrees 11'N, 15 degrees 09'E), Svalbard, in September 2005 and 2006. One of the foxes was found to be seropositive to T. gondii by the modified agglutination test (MAT). The fox was euthanized and its heart and brain were bioassayed in mice for the isolation of T. gondii. All 10 mice inoculated with brain tissue and one of the five inoculated with heart developed MAT antibodies, and tissue cysts were found in the brains of seropositive mice. Two cats fed tissues from infected mice shed T. gondii oocysts. Genotyping using 10 PCR-RFLP markers and DNA sequencing of gene loci BSR4, GRA6, UPRT1 and UPRT2 determined the isolate to be Type II strain, the predominant T. gondii lineage in the world.

PMID: 18096319 [PubMed - as supplied by publisher]

Targeting farnesyl diphosphate synthase

Bioorg Med Chem. 2007 Dec 10 [Epub ahead of print]

Synthesis and biological evaluation of 2-alkylaminoethyl-1,1-bisphosphonic acids against Trypanosoma cruzi and Toxoplasma gondii targeting farnesyl diphosphate synthase

Szajnman SH, Garcı A Liñares GE, Li ZH, Jiang C, Galizzi M, Bontempi EJ, Ferella M, Moreno SN, Docampo R, Rodriguez JB

Departamento de Quı´mica Orgánica and UMYMFOR (CONICET–FCEyN), Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Pabellón 2, Ciudad Universitaria, C1428EHA Buenos Aires, Argentina.

The effect of a series of 2-alkylaminoethyl-1,1-bisphosphonic acids against proliferation of the clinically more relevant form of Trypanosoma cruzi, the etiologic agent of American trypanosomiasis (Chagas' disease), and against tachyzoites of Toxoplasma gondii has been studied. Most of these drugs exhibited an extremely potent inhibitory action against the intracellular form of T. cruzi, exhibiting IC(50) values at the low micromolar level. This cellular activity was associated with a strong inhibition of the enzymatic activity of T. cruzi farnesyl diphosphate synthase (TcFPPS), which constitutes a valid target for Chagas' disease chemotherapy. Compound 17 was an effective agent against amastigotes exhibiting an IC(50) value of 0.84muM, while this compound showed an IC(50) value of 0.49muM against the target enzyme TcFPPS. Interestingly, compound 19 was very effective against both T. cruzi and T. gondii exhibiting IC(50) values of 4.1muM and 2.6muM, respectively. In this case, 19 inhibited at least two different enzymes of T. cruzi (TcFPPS and solanesyl diphosphate synthase (TcSPPS); 1.01muM and 0.25muM, respectively), while it inhibited TgFPPS in T. gondii. In general, this family of drugs was less effective against the activity of T. cruzi SPPS and against T. gondii growth in vitro. As bisphosphonate-containing compounds are FDA-approved drugs for the treatment of bone resorption disorders, their potential low toxicity makes them good candidates to control tropical diseases.

PMID: 18096393 [PubMed - as supplied by publisher]

Evaluation of an intranasal vaccine using recombinant proteins

Exp Parasitol. 2007 Oct 11 [Epub ahead of print]

Toxoplasma gondii: Evaluation of an intranasal vaccine using recombinant proteins against brain cyst formation in BALB/c mice

Igarashi M, Kano F, Tamekuni K, Machado RZ, Navarro IT, Vidotto O, Vidotto MC, Garcia JL

Protozoology Laboratory, Departamento de Medicina Veterinária Preventiva, Universidade Estadual de Londrina, Postal Box 6001, 86050-970, Londrina, PR, Brazil.

The purpose of this work was to evaluate protective activity against brain cyst formation in BALB/c mice intranasally vaccinated with recombinant proteins from Toxoplasma gondii. The recombinant proteins rROP2, rGRA5 and rGRA7 were used in vaccine preparation. Thirty-three female mice were divided into three groups, these animals received two doses by intranasal route at days 0 and 21 as follows; group 1 (G1, n=11) received 12.5mug of each recombinant protein plus 0.5mug of cholera toxin, group 2 (G2, n=11) received phosphate buffer saline (PBS) plus 0.5mug of cholera toxin, and group 3 (G3, n=11) received PBS only. At challenge day (day 33) three animals from each group were euthanatized for IgA measure from intestine. Mice were infected orally with 50 cysts from the VEG strain at day 33. At challenge day the G1 animals had high immunoglobulin A levels, however, they only showed IgG antibody titers against rROP2 and rGRA7. Animals from G1 also exhibited strong resistance to cyst formation compared with the control group (G3, P<0.05). However, we did not observe differences in protection against brain cyst formation between G1 and G2 (P>0.1). These results indicate that intranasal immunization in BALB/c mice with recombinant proteins rROP2, rGRA5 and rGRA7 associated with cholera toxin induced partial protection, when compared with G3, against tissue cyst formation after oral infection with tissue cysts from T. gondii.

PMID: 18154953 [PubMed - as supplied by publisher]

Genetic diversity among sea otter isolates of Toxoplasma gondii

Vet Parasitol. 2007 Nov 17 [Epub ahead of print]

Genetic diversity among sea otter isolates of Toxoplasma gondii

Sundar N, Cole RA, Thomas NJ, Majumdar D, Dubey JP, Su C

United States Department of Agriculture, Agricultural Research Service, Animal Natural Resources Institute, Animal Parasitic Diseases Laboratory, Building 1001, Beltsville, MD 20705-2350, USA.

Sea otters (Enhydra lutris) have been reported to become infected with Toxoplasma gondii and at times succumb to clinical disease. Here, we determined genotypes of 39 T. gondii isolates from 37 sea otters in two geographically distant locations (25 from California and 12 from Washington). Six genotypes were identified using 10 PCR-RFLP genetic markers including SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico, and by DNA sequencing of loci SAG1 and GRA6 in 13 isolates. Of these 39 isolates, 13 (33%) were clonal Type II which can be further divided into two groups at the locus Apico. Two of the 39 isolates had Type II alleles at all loci except a Type I allele at locus L358. One isolate had Type II alleles at all loci except the Type I alleles at loci L358 and Apico. One isolate had Type III alleles at all loci except Type II alleles at SAG2 and Apico. Two sea otter isolates had a mixed infection. Twenty-one (54%) isolates had an unique allele at SAG1 locus. Furthermore genotyping or DNA sequence analysis for 18 of these 21 isolates at loci SAG1 and GRA6 revealed that there were two different genotypes, including the previously identified Type X (four isolates) and a new genotype named Type A (14 isolates). The results from this study suggest that the sea otter isolates are genetically diverse.

PMID: 18155841 [PubMed - as supplied by publisher]

Studies on synchronous egress of coccidian parasites

Vet Res Commun. 2007 Dec 25 [Epub ahead of print]

Studies on synchronous egress of coccidian parasites (Neospora caninum, Toxoplasma gondii, Eimeria bovis) from bovine endothelial host cells mediated by calcium ionophore A23187

Behrendt JH, Taubert A, Zahner H, Hermosilla C.
Institute of Parasitology, Justus Liebig University Giessen, D-35392, Giessen, Germany.

Neospora caninum, Toxoplasma gondii and Eimeria bovis are coccidian parasites of veterinary importance. Tachyzoites of N. caninum and T. gondii and sporozoites of E. bovis are able to invade and replicate in endothelial cells in vivo and in vitro. As it holds true for all eukaryotic cells, the survival of parasitized host cells and the parasites themselves should be dependent on ion balances, especially on extra- and intracellular calcium concentrations. Addition of the calcium ionophore A23187 reliably did release merozoites from mature N. caninum and T. gondii meronts grown in cultured primary bovine umbilical vein endothelial cells (BUVEC). Extent and time course of merozoite release depended on both, maturity of the meronts and concentration of the calcium ionophore. Attempts, however, to achieve synchronous release of merozoites from E. bovis first generation meronts by ionophore treatment failed, suggesting a different biological behaviour of this parasite. According to microscopical observations, the quite variable time of E. bovis macromeront maturation and a hampered merozoite exit owing to dense parasite-induced cytoskeleton elements surrounding the meront may be a reason for the lack of inducible synchronous release.

PMID: 18158611 [PubMed - as supplied by publisher]

Thursday, December 20, 2007

Effect of hyperprolactinaemia on Toxo prevalence in humans

Parasitol Res. 2007 Dec 20 [Epub ahead of print]

Effect of hyperprolactinaemia on Toxoplasma gondii prevalence in humans

Dzitko K, Malicki S, Komorowski J

Department of Immunoparasitology, Institute of Microbiology and Immunology, University of Łódź, ul. Banacha 12/16, 90-237, Łódź, Poland,

Recent studies have shown that hormones could induce anti-parasitic functions of the host immune system; thus, the aim of the present study was to estimate the seroprevalence of Toxoplasma gondii antibodies by an enzyme-linked immunosorbent assay in a Polish population of women and men with hyperprolactinaemia (n = 234) and hypoprolactinaemia (n = 41) and in a control group (n = 281) with the physiological level of prolactin (PRL). Women with hyperprolactinaemia revealed lower seroprevalence than those with normal PRL level (33.90% and 45.58%, respectively; p = 0.025). Detailed analysis of the results showed that twofold, threefold, fourfold and fivefold increase of the PRL concentration above the normal was correlated to the decrease of the T. gondii seroprevalence, but only in the group of women with a very high PRL level (>86 ng/ml) seroprevalence (12.50%) was significantly lower (p = 0.0004) than in the control subjects. These results confirm previously described suggestions on the relationship between hyperprolactinaemia and parasitic infection frequency. We postulate that a high level of PRL may be one of the important factors preventing T. gondii infection in women.

PMID: 18092180 [PubMed - as supplied by publisher]

Thursday, December 13, 2007

Azurin-like protein blocks invasion of Toxo through potential interactions with SAG1

Antimicrob Agents Chemother. 2007 Dec 10 [Epub ahead of print]

Azurin-like protein blocks invasion of Toxoplasma gondii through potential interactions with parasite surface antigen SAG1

Naguleswaran A, Fialho AM, Chaudhari A, Hong CS, Chakrabarty AM, Sullivan WJ Jr

Department of Pharmacology & Toxicology, Indiana University School of Medicine, Indianapolis, Indiana; Department of Microbiology & Immunology, University of Illinois College of Medicine, Chicago, Illinois; Institute for Biotechnology and Bioengineering (IBB), Centre for Biological and Chemical Engineering, Instituto Superior Tecnico, Lisbon, Portugal.

Some pathogenic bacteria produce factors that have evolved a capacity to neutralize competing microbes. The cupredoxin family protein azurin, produced by Pseudomonas aeruginosa, exhibits a remarkable ability to impede invasion of a number of diverse intracellular pathogens, including human AIDS virus HIV-1 and the protozoan parasite Plasmodium falciparum (malaria). Here we report that azurin and an azurin-like protein (Laz) from gonococci/meningococci have activity against Toxoplasma, an apicomplexan parasite that causes opportunistic infection in immunocompromised individuals. We demonstrate that the mechanism of action for Laz involves interfering with the ability of Toxoplasma to adhere to host cells. Computer structural analysis reveals that azurin shares structural features with the predominant surface antigen SAG1, which is known to play an important role in parasite attachment. Interestingly, azurin also has structural similarities with a monoclonal antibody to SAG1. Surface plasmon resonance binding studies validate that SAG1 interacts strongly with Laz and, to lesser extent, azurin. Moreover, Toxoplasma mutants lacking SAG1 are not as susceptible to the growth inhibitory effects of Laz. Collectively, our data show that Toxoplasma adhesion can be significantly impaired by Laz, and to some extent by azurin, via interactions with SAG1. These observations indicate that Laz can serve as an important tool in the study of host-pathogen interactions, and are worthy of further study for development into potential therapeutic agents.

PMID: 18070964 [PubMed - as supplied by publisher]

A technique for dating toxoplasmosis in pregnancy and comparison with the Vidas anti-toxoplasma IgG avidity test

Clin Microbiol Infect. 2007 Dec 7 [Epub ahead of print]

A technique for dating toxoplasmosis in pregnancy and comparison with the Vidas anti-toxoplasma IgG avidity test

Flori P, Bellete B, Crampe C, Maudry A, Patural H, Chauleur C, Hafid J, Raberin H, Tran Manh Sung R

Pôle de Biologie-Pathologie, Laboratoire de Parasitologie et Mycologie, Hôpital Nord, CHU de Saint Etienne, France.

A comparative evaluation of 384 selected sera was performed using the Beckman Coulter Access and Abbott Axsym Toxo-IgG assays. The Axsym assay yields positive early results following infection, while the Access assay gives higher titres during chronic infection. The ratio between the two complementary tests, Axsym Toxo-IgG/Access Toxo-IgG (Ax/Ac), was compared with the Vidas anti-Toxoplasma IgG avidity index (AI). The Ax/Ac ratio decreased progressively as the time between infection and sampling increased. The mean Ax/Ac values (+/-SE) were 2.50 (+/-0.26), 2.14 (+/-0.13), 2.33 (+/-0.22), 1.34 (+/-0.09), 1.32 (+/-0.10), 0.92 (+/-0.08) and 0.74 (+/-0.07) for groups of sera sampled at 1, 2, 3, 4-5, 6-8, 9-12 and 13-24 months, respectively, after infection in pregnant women. These values were much smaller for cases with chronic infection (>24 months), i.e., 0.56 (+/-0.03), 0.44 (+/-0.04) and 0.53 (+/-0.04), respectively, for pregnant women and immunodepressed patients with and without reactivation. Taking a ratio of 1 as a threshold for recent infection, the patients in the groups sampled at 1, 2 and 3 months had Ax/Ac ratios >1 in 49/50 (98%), 53/55 (96.4%) and 36/36 (100%) cases, respectively. Thus, an Ax/Ac ratio of <1 in serum from a pregnant woman allows a recent infection (<3 months) to be excluded. This technique has the advantage of yielding positive results that develop much more rapidly than the AI, thereby helping to reassure large numbers of pregnant women and avoiding costly and unnecessary prophylactic treatment and follow-up.

PMID: 18070124 [PubMed - as supplied by publisher]

Functional expression of ribozymes in Apicomplexa: Towards exogenous control of gene expression by inducible RNA-cleavage

Int J Parasitol. 2007 Nov 6 [Epub ahead of print]

Functional expression of ribozymes in Apicomplexa: Towards exogenous control of gene expression by inducible RNA-cleavage

Agop-Nersesian C, Pfahler J, Lanzer M, Meissner M

Hygiene-Institute, Department of Parasitologie, University Hospital Heidelberg, Im Neuenheimer Feld 324, D-69120 Heidelberg, Germany.

The ability to control expression of a specific gene is a prerequisite to understand the function of essential genes. Many gene regulation systems operate on the transcriptional level by employing heterologous cis- and trans-acting elements. Recently, novel approaches employing autocatalytic RNA have been reported for different organisms. Here we show specific downregulation of gene expression in the apicomplexan parasites Toxoplasma gondii and Plasmodium falciparum, employing self-cleaving ribozymes integrated into the transcriptional unit of different genes. Moreover, we demonstrate the potential to specifically upregulate reporter gene expression by employment of the recently identified ribozyme inhibitor toyocamycin. At the RNA-level, we were able to significantly stabilise the mRNA in T. gondii. Furthermore we show that the adenosine analogue toyocamycin needs to be phosphorylated by adenosine kinase in order to act as an inhibitor for hammerhead ribozymes, since neither upregulation of reporter gene expression nor a toxic effect of toyocamycin can be detected in parasites that do not express the enzyme adenosine kinase.

PMID: 18062972 [PubMed - as supplied by publisher]

Diagnosis of Sarcocystis cruzi, Neospora caninum, and Toxoplasma gondii infections in cattle

Parasitol Res. 2007 Dec 8 [Epub ahead of print]

Diagnosis of Sarcocystis cruzi, Neospora caninum, and Toxoplasma gondii infections in cattle

Moré G, Basso W, Bacigalupe D, Venturini MC, Venturini L

Laboratorio de Inmunoparasitología, Cátedra de Parasitología y Enfermedades Parasitarias, Facultad de Ciencias Veterinarias, Universidad Nacional de La Plata, 60 y 118, 1900, La Plata, Argentina,

The aim of the study was to diagnose Sarcocystis sp. infections in cattle and to detect coinfections by Toxoplasma gondii and/or Neospora caninum. Blood, diaphragm, esophagus, and myocardium from 90 beef cattle from Argentina were collected. Histopathological, immunohistochemical, polymerase chain reaction assays, and direct microscopical examination were carried out. Sarcocysts from myocardium were measured and counted. Indirect fluorescent antibody test (IFAT) for the three protozoans was performed. Sarcocystis cruzi sarcocysts were found in 100% of myocardium samples. Sarcocysts per gram ranged from 8 to 380 with higher values found in adult cattle (p < 0.001). T. gondii and N. caninum were not detected by immunohistochemistry. T. gondii DNA was found in myocardium of 2/20 seropositive animals, while N. caninum DNA was not found. Antibodies against S. cruzi were detected in all samples, those against N. caninum in 73% and against T. gondii in 91% of the samples (IFAT titer >/=25). It is concluded that serology by IFAT is a suitable method to diagnose these protozoan infections due to its specific IgG detection; therefore, IFAT may be a useful tool to evaluate the impact of each protozoan infection in coinfected animals.

PMID: 18066600 [PubMed - as supplied by publisher]

Saturday, December 08, 2007

Pre-transplant Toxo seropositivity among heart transplant recipients is associated with an increased risk of all-cause and cardiac mortality

J Am Coll Cardiol. 2007 Nov 13;50(20):1967-72. Epub 2007 Oct 29

Pre-transplant Toxoplasma gondii seropositivity among heart transplant recipients is associated with an increased risk of all-cause and cardiac mortality

Arora S, Jenum PA, Aukrust P, Rollag H, Andreassen AK, Simonsen S, Gude E, Fiane AE, Geiran O, Gullestad L

Department of Cardiology, Rikshospitalet-Radiumhospitalet Medical Center, Oslo, Norway.

OBJECTIVES: We evaluated the risk of mortality, development of cardiac allograft vasculopathy (CAV), and acute cellular rejection among Toxoplasma gondii (T. gondii) seropositive heart transplant (HTx) recipients and the 4 donor/recipient seropairing groups. BACKGROUND: Chronic T. gondii infection is known to trigger potentially adverse immunoregulatory changes, but the long-term implication for HTx recipients has not been assessed previously. METHODS: Frozen pre-HTx serum samples of 288 recipients and 246 donors were evaluated for T. gondii serostatus using Platelia immunoglobulin G immunoassay. Patients had undergone prospective serotesting using alternative assays, and results determined by the 2 methods were compared. Data regarding mortality, CAV, and acute cellular rejection were available for all patients. RESULTS: Overall, 211 recipients (73%) were seronegative and 77 (27%) were seropositive. In total, 82 recipients died, 76 developed CAV, and 82 had 1 or more episode of treated cellular rejection. Recipient seropositivity was associated with a significantly higher risk of all-cause (hazard ratio [HR] 1.9, 95% confidence interval [CI] 1.1 to 3.4; p = 0.02) and CAV mortality (HR 4.4, 95% CI 1.3 to 15.6; p = 0.02) and a higher risk of developing advanced CAV (HR 2.7, 95% CI 1.2 to 5.8; p = 0.01). Seropositivity did not influence the number of rejection episodes, and donor/recipient seropairing was not a risk factor for any end point. CONCLUSIONS: T. gondii seropositivity among HTx recipients is associated with an increased risk of all-cause and CAV mortality and of development of advanced CAV. This may be mediated via immunoregulatory changes triggered by chronic T. gondii infection and needs to be explored further.

PMID: 17996562 [PubMed - indexed for MEDLINE]

Functional domains of GRA2 in the formation of the membranous nanotubular network

Int J Parasitol. 2007 Nov 1 [Epub ahead of print]

Functional domains of the Toxoplasma GRA2 protein in the formation of the membranous nanotubular network of the parasitophorous vacuole

Travier L, Mondragon R, Dubremetz JF, Musset K, Mondragon M, Gonzalez S, Cesbron-Delauw MF, Mercier C

Laboratoire Adaptation et Pathogénie des Micro-organismes, Université Joseph Fourier GRENOBLE 1, Centre National de la Recherche Scientifique UMR 5163, BP 170, Campus Santé, Domaine de la Merci, 38042 Grenoble cedex 9, France.

Amphipathic alpha-helices have been proposed as the general means used by soluble proteins to induce membrane tubulation. Previous studies had shown that the GRA2 dense granule protein of Toxoplasma gondii would be a crucial protein for the formation of the intravacuolar membranous nanotubular network (MNN) and that one of the functions of the MNN is to organise the parasites within the parasitophorous vacuole. GRA2 is a small protein (185 amino acids), predicted to contain three amphipathic alpha-helices (alpha1: 70-92; alpha2: 95-110 and alpha3: 119-139) when using the standard programs of secondary structure prediction. To investigate the respective contribution of each alpha-helix in the GRA2 functions, we used DeltaGRA2-HXGPRT knock-out complementation: eight truncated forms of GRA2 were expressed in the deleted recipient and the phenotypes of these mutants were analysed. This study showed that: (i) alpha3, when associated with the N-terminal region (NT) and the C-terminal region (CT), is sufficient to target the protein to the parasite posterior end and to induce formation of membranous vesicles within the vacuole. However, when associated only with CT, alpha3 is not sufficient to provide the hydrophobicity required for membrane association; (ii) the alpha1alpha2 region is alone not sufficient to induce membrane tubulation within the PV; and (iii) only one mutant, NT-alpha1alpha2alpha3, restores most of the biochemical and functional properties of GRA2, including traffic to the dense granules, secretion into the vacuole, association with vacuolar membranes, induction of the MNN formation and organisation of the parasites within the vacuole.

PMID: 18061598 [PubMed - as supplied by publisher]

Identification of novel bradyzoite-specific genes with domains for protein-protein interactions

Mol Biochem Parasitol. 2007 Oct 24 [Epub ahead of print]

Identification of novel bradyzoite-specific Toxoplasma gondii genes with domains for protein-protein interactions by suppression subtractive hybridization

Friesen J, Fleige T, Gross U, Bohne W

Institute of Medical Microbiology, University of Göttingen, Kreuzbergring 57, Göttingen D-37075, Germany.

By using suppression subtractive hybridization we identified five so far uncharacterized stage specific genes in Toxoplasma gondii, which are induced during tachyzoite-to-bradyzoite differentiation. The mRNA level of a putative zinc-finger protein was increased 23-fold in bradyzoites, while the remaining four genes displayed induction levels >100-fold. Two of these genes predict proteins with domains for protein-protein interactions. One protein (ANK1) contains both, a TPR-domain and an ankyrin motif, which consists of seven repeats. ANK1 was shown by epitope tagging experiments to be localized in the cytosol. In a fraction of parasites, the myc-tagged fusion protein was additionally localized in the nucleus, which is in agreement with the presence of a bipartite nuclear targeting sequence in ANK1. The identification of bradyzoite-specific proteins with TPR- and ankyrin-domains supports the concept that during stage conversion a variety of proteins which are involved in protein-protein interactions are induced, thereby assisting the rebuilding of the proteome.

PMID: 18061287 [PubMed - as supplied by publisher]

Diagnosis of Ocular Toxocariasis by Establishing Intraocular Antibody Production

Am J Ophthalmol. 2007 Nov 29 [Epub ahead of print]

Diagnosis of Ocular Toxocariasis by Establishing Intraocular Antibody Production

de Visser L, Rothova A, de Boer JH, van Loon AM, Kerkhoff FT, Canninga-van Dijk MR, Weersink AY, de Groot-Mijnes JD

Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands; Donders Institute of Ophthalmology, University Medical Center Utrecht, Utrecht, The Netherlands.

PURPOSE: To investigate the role of Toxocara canis in posterior uveitis of undetermined origin. DESIGN: Retrospective case-study. METHODS: Paired ocular fluid (47 aqueous humor [AH] and two vitreous fluids) and serum samples of 37 adults and 12 children with undetermined posterior uveitis were retrospectively analyzed for intraocular IgG antibody production against Toxocara canis by enzyme-linked immunosorbent assay and Goldmann-Witmer coefficient (GWC) determination. Previous diagnostic investigation by polymerase chain reaction and GWC for Herpes simplex virus, Varicella zoster virus, and Toxoplasma gondii had not provided a cause of the posterior uveitis. RESULTS: Three of 12 (25%) children showed intraocular IgG production against Toxocara canis. One child had vitritis, one presented with a low-grade uveitis and a peripheral retinal lesion, and the third had posterior uveitis and a chorioretinal scar. All three children had AH IgG titers exceeding those of the corresponding serum. In fact, two children had low Toxocara serum IgG titers (<1:32) and would have been considered seronegative upon routine serology screening. Intraocular antibody production against Toxocara canis was absent in all 37 adults, including five seropositive patients. CONCLUSIONS: Our results indicate that ocular toxocariasis is mainly a pediatric disease. Serological screening is not informative for the diagnosis of intraocular Toxocara infection. Toxocara GWC analysis, however, can be of value when diagnosing patients with posterior focal lesions or vitritis of unknown etiology.

PMID: 18061138 [PubMed - as supplied by publisher]

Kiss and spit: the dual roles of Toxoplasma rhoptries

Nat Rev Microbiol. 2007 Dec 3 [Epub ahead of print]

Kiss and spit: the dual roles of Toxoplasma rhoptries

Boothroyd JC, Dubremetz JF

Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California 94305-5124, USA.

Toxoplasma gondii is a single-celled, eukaryotic parasite that can only reproduce inside a host cell. Upon entry, this Apicomplexan parasite co-opts host functions for its own purposes. An unusual set of apical organelles, named rhoptries, contain some of the machinery that is used by T. gondii both for invasion and to commandeer host functions. Of particular interest are a group of injected protein kinases that are among the most variable of all the T. gondii proteins. At least one of these kinases has a major effect on host-gene expression, including the modulation of key regulators of the immune response. Here, we discuss these recent findings and use them to propose a model in which an expansion of host range is a major force that drives rhoptry-protein evolution.

PMID: 18059289 [PubMed - as supplied by publisher]

Death Receptor Ligation or Exposure to Perforin Trigger Rapid Egress of the Intracellular Parasite Toxoplasma

J Immunol. 2007 Dec 15;179(12):8357-65

Death Receptor Ligation or Exposure to Perforin Trigger Rapid Egress of the Intracellular Parasite Toxoplasma gondii

Persson EK, Agnarson AM, Lambert H, Hitziger N, Yagita H, Chambers BJ, Barragan A, Grandien A

Center for Infectious Medicine

The obligate intracellular parasite Toxoplasma gondii chronically infects up to one-third of the global population, can result in severe disease in immunocompromised individuals, and can be teratogenic. In this study, we demonstrate that death receptor ligation in T. gondii-infected cells leads to rapid egress of infectious parasites and lytic necrosis of the host cell, an active process mediated through the release of intracellular calcium as a consequence of caspase activation early in the apoptotic cascade. Upon acting on infected cells via death receptor- or perforin-dependent pathways, T cells induce rapid egress of infectious parasites able to infect surrounding cells, including the Ag-specific effector cells.

PMID: 18056381 [PubMed - in process]

Detection of Toxoplasma gondii-like oocysts in cat feces and estimates of the environmental oocyst burden

J Am Vet Med Assoc. 2007 Dec 1;231(11):1676-84

Detection of Toxoplasma gondii-like oocysts in cat feces and estimates of the environmental oocyst burden

Dabritz HA, Miller MA, Atwill ER, Gardner IA, Leutenegger CM, Melli AC, Conrad PA

Departments of Pathology, Microbiology, and Immunology, School of Veterinary Medicine, University of California, Davis, CA 95616.

Objective-To estimate the analytic sensitivity of microscopic detection of Toxoplasma gondii oocysts and the environmental loading of T gondii oocysts on the basis of prevalence of shedding by owned and unowned cats. Design-Cross-sectional survey. Sample Population-326 fecal samples from cats. Procedures-Fecal samples were collected from cat shelters, veterinary clinics, cat-owning households, and outdoor locations and tested via ZnSO(4) fecal flotation. Results-Only 3 (0.9%) samples of feces from 326 cats in the Morro Bay area of California contained T gondii-like oocysts. On the basis of the estimated tonnage of cat feces deposited outdoors in this area, the annual burden in the environment was estimated to be 94 to 4,671 oocysts/m(2) (9 to 434 oocysts/ft(2)). Conclusions and Clinical Relevance-Despite the low prevalence and short duration of T gondii oocyst shedding by cats detected in the present and former surveys, the sheer numbers of oocysts shed by cats during initial infection could lead to substantial environmental contamination. Veterinarians may wish to make cat owners aware of the potential threats to human and wildlife health posed by cats permitted to defecate outdoors.

PMID: 18052801 [PubMed - in process]

Thursday, December 06, 2007

Inhibition of Lewis Lung Carcinoma Growth by Toxoplasma

J Korean Med Sci. 2007 Sep;22 Suppl:S38-46.

Inhibition of Lewis Lung Carcinoma Growth by Toxoplasma gondii through Induction of Th1 Immune Responses and Inhibition of Angiogenesis

Kim JO, Jung SS, Kim SY, Kim TY, Shin DW, Lee JH, Lee YH

Department of Internal Medicine, College of Medicine, Chungnam National University, Daejeon, Korea.

Toxoplasma gondii is an obligate intracellular protozoan parasite that induces antitumor activity against certain types of cancers. However, little information is available regarding the immunologic mechanisms that regulate these effects. For this purpose, C57BL/6 mice were administered either the T. gondii Me49 strain orally or Lewis lung carcinoma (LLC) cells intramuscularly. Survival rates, tumor size, histopathology, and immune responses were determined for each group, and angiogenesis was evaluated by in vivo Matrigel plug assay. Toxoplasma-infected (TG-injected) mice survived the entire experimental period, whereas cancer cell-bearing (LLC-injected) mice died within six weeks. Mice injected with both T. gondii and cancer cells (TG/LLC-injected group) showed significantly increased survival rates, CD8+ T-cell percentages, IFN-gamma mRNA expression levels, serum IgG2a titers, and CTL responses as compared to the LLC-injected mice. In addition, angiogenesis in the TG/LLC-injected mice was notably inhibited. These effects in TG/LCC-injected mice were similar or were increased by the addition of an adjuvant, Quil-A. However, TG/LLC-injected mice showed decreased percentages of CD4+ and CD8+ T cells, IFN-gamma mRNA expression levels, and serum IgG1 and IgG2a titers as compared to TG-injected mice. Taken together, our results demonstrate that T. gondii infection inhibits tumor growth in the Lewis lung carcinoma mouse model through the induction of Th1 immune responses and antiangiogenic activity.

PMID: 17923753 [PubMed - in process]