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11 Cards in this Set
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Entamoeba Histolytica
Disease caused? Transmission? Highest risk people? |
-Causes Amoebiasis; transmitted by ingestion of CYST; Fecal-Oral Route; Highest risk – Children living in areas of poor sanitation, contaminated water supplies
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Entamoeba Histolytica
Describe the lifecycle |
Cysts are ingested, trophozoite emerges in small intestine and multiples and invades the colon. Some trophozoites encyst in the colon and pass in the feces to continue the cycle. In a small percentage of infections, the trophozoite also invades other organs, such as the liver and brain, and causes abscesses.
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Entamoeba Histolytica
Why the name? How is movement accomplished? Food? |
• Entamoeba histolytica got its name because it’s an Enteric amoeba that lyses tissues (histolytica)
Characterized by: • Movement in host by extending pseudopodia; Engulfs and digests bacteria and red blood cells (RBCs); Excretes toxic products (virulence factors) that lyse host cells; Lyses host cells forming ulcers in the large intestines amoebic colitis |
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Entamoeba Histolytica
What are virulence factors? And give examples of types... [GENERAL] |
Virulence factors are molecules produced by the parasite that help:
(1) Establish infection in host (2) Cause disease or pathogenesis (3) Allow transmission from host to host (4) Evade host immune defenses General Types of Virulence Factors: (1) Adherence factors - extracellular parasites (2) Invasion factors - intracellular parasites (3) Endotoxins - capsules, LPS, LPG that coat surface of pathogen - released upon lysis of pathogen, not actively secreted (4) Exotoxins -toxins actively secreted by viable pathogens (5) Siderophores - substances produced to extract iron from host |
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Entamoeba Histolytica
What are some examples of amoebic factors implicated in pathogenesis? |
Gal/GalNac lectin: Adherence to mucin/cells, serum resistance
Fibronectin/collagen: Adherence to extracellular matrix receptors Cysteine proteinases: Invasion through the extracellular matrix Amoebapore: Lysis of target cells Phospholipases: Lysis of target cells Cytoskeleton: Adhesion plates, endocytosis, motility |
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Entamoeba Histolytica
Describe the Adherence/Killing of Host Cells by E. histolytica |
• Killing of human cells requires adherence via an ameobic cell surface lectin
- a parasite protein that specifically binds to carbohydrates on host cell PM • Carbohydrate-protein interactions are responsible for adherence • Parasite has a surface lectin that binds host cell proteins containing Galacatose and N-acetyl-D-galactosamine - called the Gal/GalNac lectin. How was this discovered?? 1. Adherence and cytolysis of human colonic epithelia cells, neutrophils, macrophages and T cells is blocked in vitro by 50 mM galactose (Gal) or N- -acetyl-D-galactosamine (GalNAc) 2. Host cell lines that are mutant in glycosylation enzymes that add terminal Gal/GalNAc oligosaccharides to plasma membrane surface proteins are resistant to killing by E. histolytica -Used wild-type(parental) and carbohydrate-deficient mutant mammalian cells lines to test parasite adherence and cytotoxicity [see slide] - Alternative assay for studying adherence: Modify the lectin on the surface of the parasite and test the effect on adherence to the mammalian target cell. - Purification of the parasite lectin using galactose affinity chromatography revealed a heterodimer composed of a 170 kDa heavy chain and a 35 kDa light chain that at held together by disulphide bonding. |
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Entamoeba Histolytica
Using the purified Gal/GalNac proteins the genes encoding this dimeric virulence factor were cloned: |
(a) Peptide sequence & PCR
(b) Antibody production and screening of cDNA expression libraries With gene in hand, recombinant E. histolytica can be made: - over-express the gene; silence the gene - what are the effects on pathogenesis in an animal model system? |
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Entamoeba Histolytica
Describe E. histolytica Gal/GalNAc lectin - a versatile virulence factor |
• Mediates adherence to mammalian host cells and induces apoptosis of host cell; E. histolytica strains with reduced ability to kill host cells have decreased levels of lgl1 mRNA; Mediates E. coli uptake by amoeba; abundance of intracellular bacteria in E. histolytica results in down-regulation of lgl1 gene expression; Induces an antibody response in infected children that plays a significant role in acquired immunity; Recent data show that the Gal/GalNAc lectin plays a role in encystation, via galactose-binding in host intestines
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Entamoeba Histolytica
Describe Ameobapore - A novel virulence factor |
• 5 kDa proteins, 77 amino acids with 6 Cysteines; Family of 3 proteins with ~50% similarity (2005 – genome sequence publication reveals 6 genes in 3 families)
• Proteins form pores in membranes - forming β-barrels – allows this pathogen to lyse host cells after Gal/GalNac mediated binding • One amoebapore (AP-A) specifically lyses eukaryotic cell membranes • Another ameobapore has specificity for bacterial membranes: used to lyse endocytosed or phagocytosed bacteria • Amoebopores do NOT bind to or form pores in E. histolytica membranes, indicating the membrane lipids can protect the parasite*** -Used RNA silencing to reduce mRNA for amoebaporeA (specific for eukaryotic (host cell) membranes) to undetectable levels -Introduced AP-A minus trophozoites into a SCID mouse model for studying liver abscess: AP-A minus parasites cause significantly smaller amebic liver abscesses compared to parental strain |
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Entamoeba Histolytica
Describe the Reclassification of non-pathogenic E. histolytica as E. dispar |
Break-through: pathogenic and non-pathogenic isolates can be distinguished
• isoenzyme studies • gene specific differences detected by PCR • comparative genomics Current Knowledge: Most isolates are not E. histolytica, but non-pathogenic E. dispar Only isolates associated with disease are E. histolytica - At least 6 cysteine proteases are expressed by E. histolytica; some are secreted and play a role in disrupting host cells [Another pathogenicity factor…] -There is a lack of cysteine protease 5 (CP5) RNA expression in Entamoeba dispar |
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Entamoeba Histolytica
Summary of E. histolytica |
Amoebiasis is a major cause of death from parasitic disease worldwide. The causative protozoan parasite, Entamoeba histolytica, is a potent pathogen that secretes proteinases that dissolve host tissues, kills host cells on contact, and engulfs red blood cells. E. histolytica trophozoites also invade the intestinal mucosa, causing amoebic colitis. Amoebas can breach the mucosal barrier and travel through the portal circulation to the liver, where they cause abscesses containing a few E. histolytica trophozoites, dead and dying hepatocytes and liquefied cellular debris. Amoebic liver abscesses grow inexorably and, at one time, were almost always fatal, but now even large abscesses can be cured by one dose of antibiotic. A variety of virulence factor are currently being investigated to better define pathogenic mechanisms. Evidence for a single, pathogenic/non-pathogenic species based on morphology is now known to be two genetically distinct species-termed Entamoeba histolytica (the pathogen) and Entamoeba dispar (a commensal).
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