Friday, November 30, 2012

Library of Apicomplexan Metabolic Pathways: a manually curated database for metabolic pathways of apicomplexan parasites

Nucleic Acids Res. 2012 Nov 27. [Epub ahead of print]

Library of Apicomplexan Metabolic Pathways: a manually curated database for metabolic pathways of apicomplexan parasites

Shanmugasundram A, Gonzalez-Galarza FF, Wastling JM, Vasieva O, Jones AR.

Department of Functional and Comparative Genomics, Institute of Integrative Biology, University of Liverpool, Biosciences Building, Crown Street, Liverpool L69 7ZB and Department of Infection Biology, Institute of Infection and Global Health, University of Liverpool, Liverpool Science Park Innovation Centre 2, 146 Brownlow Hill, Liverpool L3 5RF, UK.

The Library of Apicomplexan Metabolic Pathways (LAMP, http://www.llamp.net) is a web database that provides near complete mapping from genes to the central metabolic functions for some of the prominent intracellular parasites of the phylum Apicomplexa. This phylum includes the causative agents of malaria, toxoplasmosis and theileriosis-diseases with a huge economic and social impact. A number of apicomplexan genomes have been sequenced, but the accurate annotation of gene function remains challenging. We have adopted an approach called metabolic reconstruction, in which genes are systematically assigned to functions within pathways/networks for Toxoplasma gondii, Neospora caninum, Cryptosporidium and Theileria species, and Babesia bovis. Several functions missing from pathways have been identified, where the corresponding gene for an essential process appears to be absent from the current genome annotation. For each species, LAMP contains interactive diagrams of each pathway, hyperlinked to external resources and annotated with detailed information, including the sources of evidence used. We have also developed a section to highlight the overall metabolic capabilities of each species, such as the ability to synthesize or the dependence on the host for a particular metabolite. We expect this new database will become a valuable resource for fundamental and applied research on the Apicomplexa.

PMID: 23193253 [PubMed - as supplied by publisher]

Toxoplasma gondii infection enhances testicular steroidogenesis in rats

Mol Ecol. 2012 Nov 28. doi: 10.1111/mec.12042. [Epub ahead of print]

Toxoplasma gondii infection enhances testicular steroidogenesis in rats

Lim A, Kumar V, Hari Dass SA, Vyas A.

School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore, 637551, Singapore.

The protozoan parasite Toxoplasma gondii enhances the sexual attractiveness of infected male rats and attenuates the innate fear of cat odour in infected individuals. These behavioural changes plausibly lead to greater transmission of parasites through sexual and trophic routes, respectively. Testosterone, a testicular steroid, is known to reduce fear and enhance sexual attractiveness in males. Here, we show that Toxoplasma gondii infection enhances expression of genes involved in facilitating synthesis of testosterone, resulting in greater testicular testosterone production in male rats. In several species, testosterone mediates trade-offs between sexually selected traits and life history decisions. Augmentation of testosterone synthesis by Toxoplasma gondii suggests that parasites may manipulate these trade-offs in rats.

PMID: 23190313 [PubMed - as supplied by publisher]

Thursday, November 29, 2012

Subversion of host cellular functions by the apicomplexan parasites

FEMS Microbiol Rev. 2012 Nov 27. doi: 10.1111/1574-6976.12013. [Epub ahead of print]

Subversion of host cellular functions by the apicomplexan parasites

Kemp LE, Yamamoto M, Soldati-Favre D.

Department of Microbiology and Molecular Medicine, Faculty of Medicine, University of Geneva, 1211, Geneva 4, Switzerland.

Rhoptries are club-shaped secretory organelles located at the anterior pole of species belonging to the phylum of Apicomplexa. Parasites of this phylum are responsible for a huge burden of disease in humans and animals and a loss of economic productivity. Members of this elite group of obligate intracellular parasites include Plasmodium spp. that cause malaria and Cryptosporidium spp. that cause diarrhoeal disease. Although rhoptries are present almost ubiquitously throughout the phylum, the relevance and role of the proteins contained within the rhoptries varies. Rhoptry contents separates into two intra-organellar compartments, the neck and the bulb. A number of rhoptry neck proteins are conserved between species and are involved in functions such as host cell invasion. The bulb proteins are less well conserved and likely evolved for a particular lifestyle. In the majority of species studied to date, rhoptry content is involved in formation and maintenance of the parasitophorous vacuole however some species live free within the host cytoplasm. In this review we will summarise the knowledge available regarding rhoptry proteins. Specifically we will discuss the role of the rhoptry kinases that are used by Toxoplasma gondii and other coccidian parasites to subvert the host cellular functions and prevent parasite death. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

PMID: 23186105 [PubMed - as supplied by publisher]

Theoretical investigation on structural, functional and epitope of a 12 kDa excretory-secretory protein from Toxoplasma gondii

BMC Struct Biol. 2012 Nov 27;12(1):30. [Epub ahead of print]

Theoretical investigation on structural, functional and epitope of a 12 kDa excretory-secretory protein from Toxoplasma gondii

Tommy YB, Lim TS, Noordin R, Saadatnia G, Choong YS.

BACKGROUND: Toxoplasma gondii is an intracellular coccidian parasite that causes toxoplasmosis. It was estimated that more than one third of the world population is infected by T. gondii, and the disease is critical in fetuses and immunosuppressed patients. Thus, early detection is crucial for disease diagnosis and therapy. However, the current available toxoplasmosis diagnostic tests vary in their accuracy and the better ones are costly.
RESULTS:
An earlier published work discovered a highly antigenic 12 kDa excretory-secretory (ES) protein of T. gondii which may potentially be used for the development of an antigen detection test for toxoplasmosis. However, the three-dimensional structure of the protein is unknown. Since epitope identification is important prior to designing of a specific antibody for an antigen-detection based diagnostic test, the structural elucidation of this protein is essential. In this study, we constructed a three dimensional model of the 12 kDa ES protein. The built structure possesses a thioredoxin backbone which consists of four alpha-helices flanking five beta-strands at the center. Three potential epitopes (6--8 residues) which can be combined into one "single" epitope have been identified from the built structure as the most potential antibody binding site.
CONCLUSION:
Together with specific antibody design, this work could contribute towards future development of an antigen detection test for toxoplasmosis.

PMID: 23181504 [PubMed - as supplied by publisher]

Tuesday, November 20, 2012

SNPs in toll-like receptor (TLR) genes as new genetic alterations associated with congenital toxoplasmosis?

Eur J Clin Microbiol Infect Dis. 2012 Nov 17. [Epub ahead of print]

SNPs in toll-like receptor (TLR) genes as new genetic alterations associated with congenital toxoplasmosis?

Wujcicka W, Wilczyński J, Nowakowska D.

Department of Fetal-Maternal Medicine and Gynecology, Polish Mother's Memorial Hospital Research Institute, 281/289 Rzgowska Street, Lodz, 93-338, Poland.

Nearly 40 % of pregnant women are infected with Toxoplasma gondii. Primary infections in pregnant women result, in approximately 30-50 % of patients, in transmission of T. gondii through the placenta to the fetus and then in congenital infections with severe, sometimes fatal course. Studies still do not provide sufficient data on the genetic bases of the immunity in fetuses, newborns, and infants with congenital toxoplasmosis. Previous research showed the contribution of toll-like receptors (TLRs) to non-specific immunity against T. gondii invasion, observed in T. gondii-infected animals, especially mice. So far, the activity of TLRs in defense against T. gondii infections was observed particularly for TLR2, TLR4, and TLR9 molecules. Differential TLR activity associates with both cell types, including a variety of placental cells and stage of pregnancy. Several single-nucleotide polymorphisms (SNPs) residing in three genes encoding these receptors were reported as significant genetic modifications of TLRs associated with different pregnancy disorders. Despite those data, genetic alterations of TLRs which have contributed to innate immune response against T. gondii infections are still not precisely described. In this article, we present reasons for the research of the plausible role of SNPs residing in TLR2, TLR4, and TLR9 genes in congenital toxoplasmosis development.

PMID: 23161283 [PubMed - as supplied by publisher]

Mitochondrial Metabolism of Glucose and Glutamine Is Required for Intracellular Growth of Toxoplasma gondii

Cell Host Microbe. 2012 Nov 15;12(5):682-92. doi: 10.1016/j.chom.2012.09.013.

Mitochondrial Metabolism of Glucose and Glutamine Is Required for Intracellular Growth of Toxoplasma gondii

Macrae JI, Sheiner L, Nahid A, Tonkin C, Striepen B, McConville MJ.

Department of Biochemistry and Molecular Biology, Bio21 Institute of Molecular Science and Biotechnology, 30 Flemington Road, University of Melbourne, Parkville, VIC 3010, Australia.

Toxoplasma gondii proliferates within host cell vacuoles where the parasite relies on host carbon and nutrients for replication. To assess how T. gondii utilizes these resources, we mapped the carbon metabolism pathways in intracellular and egressed parasite stages. We determined that intracellular T. gondii stages actively catabolize host glucose via a canonical, oxidative tricarboxylic acid (TCA) cycle, a mitochondrial pathway in which organic molecules are broken down to generate energy. These stages also catabolize glutamine via the TCA cycle and an unanticipated γ-aminobutyric acid (GABA) shunt, which generates GABA and additional molecules that enter the TCA cycle. Chemically inhibiting the TCA cycle completely prevents intracellular parasite replication. Parasites lacking the GABA shunt exhibit attenuated growth and are unable to sustain motility under nutrient-limited conditions, suggesting that GABA functions as a short-term energy reserve. Thus, T. gondii tachyzoites have metabolic flexibility that likely allows the parasite to infect diverse cell types.

PMID: 23159057 [PubMed - in process]

Friday, November 16, 2012

Integrated Bioinformatic and Targeted Deletion Analyses of the SRS Gene Superfamily Identify SRS29C as a Negative Regulator of Toxoplasma Virulence

MBio. 2012 Nov 13;3(6). pii: e00321-12. doi: 10.1128/mBio.00321-12.

Integrated Bioinformatic and Targeted Deletion Analyses of the SRS Gene Superfamily Identify SRS29C as a Negative Regulator of Toxoplasma Virulence

Wasmuth JD, Pszenny V, Haile S, Jansen EM, Gast AT, Sher A, Boyle JP, Boulanger MJ, Parkinson J, Grigg ME.

Program in Molecular Structure and Function, Hospital for Sick Children, Toronto, Ontario, Canada.

ABSTRACT The Toxoplasma gondii SRS gene superfamily is structurally related to SRS29B (formerly SAG1), a surface adhesin that binds host cells and stimulates host immunity. Comparative genomic analyses of three Toxoplasma strains identified 182 SRS genes distributed across 14 chromosomes at 57 genomic loci. Eight distinct SRS subfamilies were resolved. A core 69 functional gene orthologs were identified, and strain-specific expansions and pseudogenization were common. Gene expression profiling demonstrated differential expression of SRS genes in a developmental-stage- and strain-specific fashion and identified nine SRS genes as priority targets for gene deletion among the tissue-encysting coccidia. A Δsag1 sag2A mutant was significantly attenuated in murine acute virulence and showed upregulated SRS29C (formerly SRS2) expression. Transgenic overexpression of SRS29C in the virulent RH parent was similarly attenuated. Together, these findings reveal SRS29C to be an important regulator of acute virulence in mice and demonstrate the power of integrated genomic analysis to guide experimental investigations. IMPORTANCE Parasitic species employ large gene families to subvert host immunity to enable pathogen colonization and cause disease. Toxoplasma gondii contains a large surface coat gene superfamily that encodes adhesins and virulence factors that facilitate infection in susceptible hosts. We generated an integrated bioinformatic resource to predict which genes from within this 182-gene superfamily of adhesin-encoding genes play an essential role in the host-pathogen interaction. Targeted gene deletion experiments with predicted candidate surface antigens identified SRS29C as an important negative regulator of acute virulence in murine models of Toxoplasma infection. Our integrated computational and experimental approach provides a comprehensive framework, or road map, for the assembly and discovery of additional key pathogenesis genes contained within other large surface coat gene superfamilies from a broad array of eukaryotic pathogens.

PMID: 23149485 [PubMed - as supplied by publisher]

Distinct signalling pathways control Toxoplasma egress and host-cell invasion

EMBO J. 2012 Nov 13. doi: 10.1038/emboj.2012.299. [Epub ahead of print]

Distinct signalling pathways control Toxoplasma egress and host-cell invasion

Lourido S, Tang K, Sibley LD.

Department of Molecular Microbiology, Washington University School of Medicine, St Louis, MO, USA.

Calcium signalling coordinates motility, cell invasion, and egress by apicomplexan parasites, yet the key mediators that transduce these signals remain largely unknown. One underlying assumption is that invasion into and egress from the host cell depend on highly similar systems to initiate motility. Using a chemical-genetic approach to specifically inhibit select calcium-dependent kinases (CDPKs), we instead demonstrate that these pathways are controlled by different kinases: both TgCDPK1 and TgCDPK3 were required during ionophore-induced egress, but only TgCDPK1 was required during invasion. Similarly, microneme secretion, which is necessary for motility during both invasion and egress, universally depended on TgCDPK1, but only exhibited TgCDPK3 dependence when triggered by certain stimuli. We also demonstrate that egress likely comes under a further level of control by cyclic GMP-dependent protein kinase and that its activation can induce egress and partially compensate for the inhibition of TgCDPK3. These results demonstrate that separate signalling pathways are integrated to regulate motility in response to the different signals that promote invasion or egress during infection by Toxoplasma gondii.

PMID: 23149386 [PubMed - as supplied by publisher]

Tuesday, November 13, 2012

The Polymorphic Pseudokinase ROP5 Controls Virulence in Toxoplasma gondii by Regulating the Active Kinase ROP18

PLoS Pathog. 2012 Nov;8(11):e1002992. Epub 2012 Nov 8.

The Polymorphic Pseudokinase ROP5 Controls Virulence in Toxoplasma gondii by Regulating the Active Kinase ROP18

Behnke MS, Fentress SJ, Mashayekhi M, Li LX, Taylor GA, Sibley LD.

Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri, United States of America.

Secretory polymorphic serine/threonine kinases control pathogenesis of Toxoplasma gondii in the mouse. Genetic studies show that the pseudokinase ROP5 is essential for acute virulence, but do not reveal its mechanism of action. Here we demonstrate that ROP5 controls virulence by blocking IFN-γ mediated clearance in activated macrophages. ROP5 was required for the catalytic activity of the active S/T kinase ROP18, which phosphorylates host immunity related GTPases (IRGs) and protects the parasite from clearance. ROP5 directly regulated activity of ROP18 in vitro, and both proteins were necessary to avoid IRG recruitment and clearance in macrophages. Clearance of both the Δrop5 and Δrop18 mutants was reversed in macrophages lacking Irgm3, which is required for IRG function, and the virulence defect was fully restored in Irgm3(-/-) mice. Our findings establish that the pseudokinase ROP5 controls the activity of ROP18, thereby blocking IRG mediated clearance in macrophages. Additionally, ROP5 has other functions that are also Irgm3 and IFN-γ dependent, indicting it plays a general role in governing virulence factors that block immunity.

PMID: 23144612 [PubMed - as supplied by publisher]

Cluster analysis identifies aminoacid compositional features that indicate Toxoplasma gondii adhesin proteins

Bioinformation. 2012;8(19):916-23. doi: 10.6026/97320630008916. Epub 2012 Oct 1.

Cluster analysis identifies aminoacid compositional features that indicate Toxoplasma gondii adhesin proteins

Arenas AF, Salcedo GE, Moncada DM, Erazo DA, Osorio JF, Gomez-Marin JE.

Grupo de Parasitología Molecular (GEPAMOL), Centro de Investigaciones Biomédicas, Universidad del Quindío, Armenia, Colombia.

Toxoplasma gondii invade host cells using a multi-step process that depends on the regulated secretion of adhesions. To identify key primary sequence features of adhesins in this parasite, we analyze the relative frequency of individual amino acids, their dipeptide frequencies, and the polarity, polarizability and Van der Waals volume of the individual amino acids by using cluster analysis. This method identified cysteine as a key amino acid in the Toxoplasma adhesin group. The best vector algorithm of non-concatenated features was for 2 attributes: the single amino acid relative frequency and the dipeptide frequency. Polarity, polarizability and Van der Waals volume were not good classificatory attributes. Single amino acid attributes clustered unambiguously 67 apicomplexan hypothetical adhesins. This algorithm was also useful for clustering hypothetical Toxoplasma target host receptors. All of the cluster performances had over 70% sensitivity and 80% specificity. Compositional aminoacid data can be useful for improving machine learning-based prediction software when homology and structural data are not sufficient.

PMID: 23144551 [PubMed - in process]

The Toxoplasma MAG1 peptides induce sex-based humoral immune response in mice and distinguish active from chronic human infection

Microbes Infect. 2012 Nov 6. pii: S1286-4579(12)00269-9. doi: 10.1016/j.micinf.2012.10.016. [Epub ahead of print]

The Toxoplasma MAG1 peptides induce sex-based humoral immune response in mice and distinguish active from chronic human infection.

Xiao J, Viscidi RP, Kannan G, Pletnikov MV, Li Y, Severance EG, Yolken RH, Delhaes L.

Stanley Division of Developmental Neurovirology, Department of Pediatrics, Johns Hopkins School of Medicine, Baltimore, MD 21287, USA. Electronic address: jxiao4@jhmi.edu.

To distinguish active from inactive/chronic infection in Toxoplasma gondii-seropositive individuals, we have developed an enzyme-linked immunosorbent assay (ELISA) using specific peptides derived from Toxoplasma matrix antigen MAG1. We used this assay to measure matrix specific antibodies and pilot studies with infected mice established the validity of two peptides. The immune response against MAG1 occurs in about 12 days post infection and displays a sex difference later on in mouse model, with males producing higher antibody titers than females. Serum samples from 22 patients with clinical toxoplasmosis and from 26 patients with serological evidence of past exposure to Toxoplasma (more than one year infection history) were analyzed. Both MAG1 peptides detected antibodies significant frequently and robustly from active stage than from the chronic stage of toxoplasmosis. The results indicate that both MAG1 peptides may be used as a tool to differentiate active from inactive infection. It also may be considered in the design of potential vaccines in humans.

PMID: 23142034 [PubMed - as supplied by publisher]