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Cloning of Trypanosoma brucei and Leishmania major genes encoding the GlcNAc-phosphatidylinositol de-N-acetylase of glycosylphosphatidylinositol biosynthesis that is essential to the African sleeping sickness parasite.

The second step of glycosylphosphatidylinositol anchor biosynthesis in all eukaryotes is the conversion of D-GlcNAcalpha1-6-d-myo-inositol-1-HPO(4)-sn-1,2-diacylglycerol (GlcNAc-PI) to d-GlcNalpha1-6-d-myo-inositol-1-HPO(4)-sn-1,2-diacylglycerol by GlcNAc-PI de-N-acetylase. The genes encoding this activity are PIG-L and GPI12 in mammals and yeast, respectively. Fragments of putative GlcNAc-PI de-N-acetylase genes from Trypanosoma brucei and Leishmania major were identified in the respective genome project data bases. The full-length genes TbGPI12 and LmGPI12 were subsequently cloned, sequenced

Expression and localization of serum resistance associated protein in Trypanosoma brucei rhodesiense.

The trypanosome lytic factor (TLF) is a primate specific innate defense mechanism that restricts the host range of African trypanosomes. Trypanosoma brucei rhodesiense, the causative agent of the acute form of human sleeping sickness, is resistant to the cytolytic action of TLF. By differential display PCR we have identified a gene in T. b. rhodesiense that is preferentially expressed in cell lines resistant to TLF. The protein sequence predicted from the gene shows homology to the trypanosome variable surface glycoprotein (VSG) gene family and in particular, to the previously reported human s

Clinical and veterinary trypanocidal benzoxaboroles target CPSF3.

African trypanosomes cause lethal and neglected tropical diseases, known as sleeping sickness in humans and nagana in animals. Current therapies are limited, but fortunately, promising therapies are in advanced clinical and veterinary development, including acoziborole (AN5568 or SCYX-7158) and AN11736, respectively. These benzoxaboroles will likely be key to the World Health Organization's target of disease control by 2030. Their mode of action was previously unknown. We have developed a high-coverage overexpression library and use it here to explore drug mode of action in Trypanosoma brucei

Performance of parasitological and molecular techniques for the diagnosis and surveillance of gambiense sleeping sickness.

OBJECTIVES: Recently, improvements have been made to diagnostics for gambiense sleeping sickness control but their performance remains poorly documented and may depend on specimen processing prior to examination. In a prospective study in the Democratic Republic of the Congo, we compared the diagnostic performance of several parasite detection techniques, immune trypanolysis and of m18S PCR on whole blood stored in a stabilisation buffer or dried on filter paper. METHODS: Individuals with CATT whole blood (WB) titer ≥1∶4 or with clinical signs indicative for sleeping sickness were examined fo

A trypanosome metacyclic VSG gene promoter with two functionally distinct, life cycle stage-specific activities.

In the mammalian bloodstream, African trypanosomes express the variant surface glycoprotein (VSG), continual switching of which allows evasion of the host immune response. Bloodstream VSG genes are transcribed from polycistronic bloodstream expression sites with promoters which are located 45-60 kb upstream. These promoters are not exclusively stage-regulated, being active in the insect midgut stage where VSG is not expressed. However, the metacyclic VSG (M-VSG) genes, a small subset activated when VSG synthesis begins in the metacyclic stage in the tsetse fly salivary glands, are transcriptio

Medical Management of the Kidney Transplant Recipient : Infections and Malignant Neoplasms

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… .* *Geographically focused infections will need to be considered in certain cases, such as malaria, leishmaniasis, trypanosomiasis, and strongyloidiasis. 1 Sources of infections specific to recipients of renal transplant: perinephric fluid collections (e.g., lymphoceles, wound hematomas, urine leaks) … Figure 101.1 Timetable of infections.* *Geographically focused infections will need to be considered in certain cases, such as malaria, leishmaniasis, trypanosomiasis, and strongyloidiasis. 1 Sources of infections specific to recipients of renal transplant: perinephric fluid collections (e.g., lymphoceles …

A post-transcriptional respiratome regulon in trypanosomes.

Post-transcriptional regulons coordinate the expression of groups of genes in eukaryotic cells, yet relatively few have been characterized. Parasitic trypanosomatids are particularly good models for studies on such mechanisms because they exhibit almost exclusive polycistronic, and unregulated, transcription. Here, we identify the Trypanosoma brucei ZC3H39/40 RNA-binding proteins as regulators of the respiratome; the mitochondrial electron transport chain (complexes I-IV) and the FoF1-ATP synthase (complex V). A high-throughput RNAi screen initially implicated both ZC3H proteins in variant sur

Disclosing the essentiality of ribose-5-phosphate isomerase B in Trypanosomatids.

Ribose-5-phosphate isomerase (RPI) belongs to the non-oxidative branch of the pentose phosphate pathway, catalysing the inter-conversion of D-ribose-5-phosphate and D-ribulose-5-phosphate. Trypanosomatids encode a type B RPI, whereas humans have a structurally unrelated type A, making RPIB worthy of exploration as a potential drug target. Null mutant generation in Leishmania infantum was only possible when an episomal copy of RPIB gene was provided, and the latter was retained both in vitro and in vivo in the absence of drug pressure. This suggests the gene is essential for parasite survival.

Trypanosoma brucei mitochondrial respiratome: composition and organization in procyclic form.

The mitochondrial respiratory chain is comprised of four different protein complexes (I-IV), which are responsible for electron transport and generation of proton gradient in the mitochondrial intermembrane space. This proton gradient is then used by F₀F₁-ATP synthase (complex V) to produce ATP by oxidative phosphorylation. In this study, the respiratory complexes I, II, and III were affinity purified from Trypanosoma brucei procyclic form cells and their composition was determined by mass spectrometry. The results along with those that we previously reported for complexes IV and V showed that

Co-infection with Trypanosoma brucei brucei prevents experimental autoimmune encephalomyelitis in DBA/1 mice through induction of suppressor APCs.

The immune system has co-evolved with the infectious agents that challenge it, and in response pathogens have developed different mechanisms to subvert host immunity. A wealth of evidence suggests that infections are important components in the development of a functional immune system, and understanding the modulation of the host immune system by pathogens may offer new therapeutic strategies in a non-infectious setting. We investigated how infection with the protozoan parasite Trypanosoma brucei brucei (Tbb) modulates the autoimmune response to recombinant myelin oligodendrocyte glycoprotein

Bovine stem cell factor: production of a biologically active protein and mRNA analysis in cattle infected with Trypanosoma congolense.

The cDNA coding for the soluble form of bovine stem cell factor (boSCFAla165) was cloned and recombinant protein was produced in bacteria as a histidine tagged-protein. The protein was purified from the inclusion bodies in one step by metal chelation chromatography under denaturing conditions. Recombinant bovine SCF was shown to act synergistically with interleukin 3 (IL-3) and erythropoietin (EPO) in stimulating the growth of bone marrow progenitor cells such as colony forming units-granulocyte macrophage (CFU-GM) and burst forming units-erythroid (BFU-E). Analysis of SCF mRNA expression by r

A 64-year-old woman with rapid neurologic decline diagnosed with Toxoplasma encephalitis after presumed metastatic cancer.

In an era of fragmented medical care, concurrent clinical features that ultimately lead to a unified diagnosis may not be prioritized appropriately. We present a case of a 64-year-old woman referred to hematology clinic for anemia, with recent memory loss and gait disturbance. Two months later, she developed pneumonia; imaging workup showed a left renal mass. Neurologic function continued to decline precluding definitive nephrectomy. She then presented with new-onset seizure and initial neuro-imaging was reported as unremarkable. One month later, outpatient neurologic workup demonstrated new l

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… tuberculous abscess Fungal Aspergillosis Cryptococcosis Rhodococcosis Zygomycosis Histoplasmosis Coccidioidomycosis Nocardiosis Mucormycosis Paracoccidioidomycosis Actinomycosis Parasitic Toxoplasmosis (often HIV-associated) Neurocysticercosis Amebic brain abscess Echinococcosis Cerebral sparganosis Trypanosomiasis …

Characterization, localization, essentiality, and high-resolution crystal structure of glucosamine 6-phosphate N-acetyltransferase from Trypanosoma brucei.

A gene predicted to encode Trypanosoma brucei glucosamine 6-phosphate N-acetyltransferase (TbGNA1; EC 2.3.1.4) was cloned and expressed in Escherichia coli. The recombinant protein was enzymatically active, and its high-resolution crystal structure was obtained at 1.86 Å. Endogenous TbGNA1 protein was localized to the peroxisome-like microbody, the glycosome. A bloodstream-form T. brucei GNA1 conditional null mutant was constructed and shown to be unable to sustain growth in vitro under nonpermissive conditions, demonstrating that there are no metabolic or nutritional routes to UDP-GlcNAc othe

Cell-mediated immunity in nutritional deficiency.

Dietary deficiencies of specific nutrients profoundly alter cell-mediated immune responses in man and experimental animals. Both moderate and severe deficiencies are associated with significant changes in immunocompetence. Diets with inadequate levels of protein, calories, vitamin A, pyridoxine, biotin and zinc result in loss of thymic cellularity. Secondary to thymic atrophy, the production of thymic hormones critical for the differentiation of T lymphocytes is reduced, especially in protein-calorie malnutrition and zinc deficiency. Confirmation of a T cell maturational defect in nutritional

Coxiella burnetii associated with BVDV (Bovine Viral Diarrhea Virus), BoHV (Bovine Herpesvirus), Leptospira spp., Neospora caninum, Toxoplasma gondii and Trypanosoma vivax in reproductive disorders in cattle.

This is a cross-sectional study to assess the presence of antibodies in ruminants against selected pathogens associated with reproductive disorders in cattle in four Brazilian states, including the zoonotic agent Coxiella burnetii. The used tests were Virus Neutralization Assay for IBR and BVD, Microscopic Agglutination Test for Leptospira spp., Indirect Fluorescent Antibody Test (IFAT) for C. burnetii and Toxoplasma gondii, and Enzyme-Linked Immunosorbent Assay for Neospora caninum and Trypanosoma vivax. Seropositivity for C. burnetii was 13.7% with titers from 128 to 131,072; 57.8% for BoHV-

Regulated processive transcription of chromatin by T7 RNA polymerase in Trypanosoma brucei.

Inability of T7 RNA polymerase to processively transcribe higher eukaryotic chromatin is interpreted as a correlate of its reported inhibition by nucleosomes on reconstituted templates in vitro . We used chromosomally integrated reporter cassettes to examine features of T7 transcription in a lower eukaryotic system. Luciferase reporters were targeted to rDNA in transgenic Trypanosoma brucei stably expressing the phage polymerase. Because trypanosome mRNAs are capped by RNA splicing in trans , T7 transcription could be gauged by luciferase activity. In contrast to findings from higher eukaryote

Ly6C- Monocytes Regulate Parasite-Induced Liver Inflammation by Inducing the Differentiation of Pathogenic Ly6C+ Monocytes into Macrophages.

Monocytes consist of two well-defined subsets, the Ly6C+ and Ly6C- monocytes. Both CD11b+ myeloid cells populations have been proposed to infiltrate tissues during inflammation. While infiltration of Ly6C+ monocytes is an established pathogenic factor during hepatic inflammation, the role of Ly6C- monocytes remains elusive. Mice suffering experimental African trypanosome infection die from systemic inflammatory response syndrome (SIRS) that is initiated by phagocytosis of parasites by liver myeloid cells and culminates in apoptosis/necrosis of liver myeloid and parenchymal cells that reduces h