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Geschichte Der Pockenkrankheit (Smallpox) Poxviruses

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Geschichte Der Pockenkrankheit (Smallpox) Poxviruses Geschichte der Pockenkrankheit (Smallpox) Poxviruses Mahalingam, S. et al., 2004, TRENDS Im., 637-639 Molekulare Virologie © 2008 by Ruth Brack-Werner Poxviridae Poxviruses Variolavirus Harrison, SC. et al., 2004, PNAS 101, 11178-11192 Molekulare Virologie © 2008 by Ruth Brack-Werner Poxviridae Poxviruses Harrison, SC.etal., 2004, PNAS 101,11178-11192 Übertragbar von Tier auf Mensch Nur beim Menschen Molekulare Virologie © 2008 by Ruth Brack-Werner Variolavirus Varianten Poxviruses Poxvirus variola maior 30 - 40% Mortalitätsrate Poxvirus variola minor ca. 1% Mortalitätrate Nur ca. 2% Unterschied zwischen beiden Genomsequenzen (ca. 185 000 Bp). Molekulare Virologie © 2008 by Ruth Brack-Werner Pockenviren Struktur: Modell Vaccinia Virus Poxviruses IMV 350 nm DNA-abhängige RNA Polymerase; RNA-Prozessierungsenzyme Harrison, SC. et al., 2004, PNAS 101, 11178-11192 250 nm Molekulare Virologie © 2008 by Ruth Brack-Werner Verschiedene Formen von Vaccinia-Viren Poxviruses IMV: Intracellular mature virion Enthalten eine Lipid- IEV: Intracellular enveloped virion Dopplemembran EEV: Extracellular eveloped virion mehr als die IMV CEV: Cell-associated enveloped Form. virion Weniger empfindlich ggü neutralisierende AK, Complement- faktoren als die IMV; EEV EEV IMV CEV Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccinia-Virus Morphogenese Poxviruses Smith, GL et al.,2002, JGenVirol. 83, 2915-2931. Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccinia-Virus DNA Poxviruses Komplette Sequenz der Kopenhagen und WR-Stämme bekannt. ITR ITR 191 kbp Virulence Proteine, Host-range Proteine, Strukturproteine, Immune Modulatoren Essentielle Enzyme Harrison, SC. et al., 2004, PNAS 101, 11178-11192 Molekulare Virologie © 2008 by Ruth Brack-Werner Replikation der Pockenviren. Poxviruses Die Replikation der Pockenviren läuft vollständig im Zytoplasma ab; Pockenvirus-Proteine werden an zelluläre Ribosomen synthetisiert; Molekulare Virologie © 2008 by Ruth Brack-Werner Proteine für die Transkription von Vaccinia Virus Genen Poxviruses Host Host Broyles S.S. 2003, Host JGenVirol 84, 2293-2303. Molekulare Virologie © 2008 by Ruth Brack-Werner Auswahl von Pockenvirus-Enzymen für die DNA Synthese Poxviruses Enzym Funktion DNA-Polymerase Synthese des viralen dopplesträngigen DNA Genoms Processivity factor Synthese des viralen dopplesträngigen DNA Genoms DNA Ligase Replikation der viralen DNA; Bildung der kovalent geschlossenen Enden des DNA Genoms; Thymidinkinase Phosphorylierung von Thymidin- Nukleosiden zu TMP Thymidilatkinase TMP -> TTP Uracil-DNA-Glycosylase Teil des Reparatursystems; Entfernt Uracilreste aus der DNA; Molekulare Virologie © 2008 by Ruth Brack-Werner Vergleich der Thymidin-Kinasen von HSV-1 und Vaccinia Virus Poxviruses HSV-1 TK Vaccinia Virus TK Typ I II Größe (kDa) 40 20 Quartär-Struktur Dimer Tetramer Feedback Nein Ja Regulation (Hemmung durch dTDP bzw. dTTP) Substratspezifität Gering: Hoch: Nur Thymidine, bzw. Phosphorylierung Bromdeoxyuridine; von Guanosine- Analoga (z.B. Acyclovir, Gancyclovir) Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccinia-Virus Replikationszyklus: Frühe Phase Poxviruses 1. Viruseintritt Fusion von vir. und zellul. Membrane Enthüllung des Virus; Freisetzung des viralen Kerns. 2. Synthese von frühen mRNA im Virus-Kern 3. Produkt. der frühen Proteine 4. Sekretion von Proteinen: Immunmodulatoren und Proteine die das Wachstum von umliegenden Zellen beeinflussen. 5. Freisetzung der viralen DNA aus dem Virus-Kern; Ende der Expression von frühen viralen Genen; Harrison, SC. et al., 2004, PNAS 101, 11178-11192 Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccinia-Virus Replikationszyklus: Mittlere („intermediate“) PhasePoxviruses 6./7. Replikation des viralen Doppelstrang DNA-Genoms 8. Transkription von “intermediate Phase” mRNAs; Relokation des Wirtsproteins Vitf2 in das Zytoplasma 9, 10. Synthese von Proteinen für die Spät- Phase: RNA polymerase; Transkriptionsfaktoren; Virus-Bestandteile; Harrison, SC. et al., 2004, PNAS 101, 11178-11192 Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccinia-Virus Replikationszyklus: Frühe Phase Poxviruses Harrison, SC. et al., 2004, PNAS 101, 11178-11192 11. Synthese von Virionproteine 12-14 Synthese des IMV 15. Freisetzung des IMV nur durch Zellyse. 16. Erwerbung einer zweiten Doppelmembran und Bildung des IEVs; 17. Transport zur Zelloberfläche; und Aktinpolymerisation: CEV; 18. Transfer des CEV zu Nachbarzelle oder Freisetzung als EEV Molekulare Virologie © 2008 by Ruth Brack-Werner Johnston JB, and McFadden G, 2003, JVi. 6093-6100 Virale Proteine für das Überleben Stützproteine für das Überleben der Infizierter Zellen Zelle als Virusproduzent Poxviruses Virorezeptoren: Imitate von Rezeptoren für Zytokine, und andere proinflam- Matorische und antivirale Moleküle Virokine: Imitate von Zytokinen, Complementfaktoren und anderen Immunmolekülen des Wirtes Virale Proteine zur Tarnung der infizierten Zellen Molekulare Virologie © 2008 by Ruth Brack-Werner Pocken als Krankheit Poxviruses Die letzten bekannten Fälle: 1949 USA 1972 Yugoslavian 1977 Somalia Ab 1979 galt die Pockenkrankheit offiziell als ausgelöscht (WHO) Verbreitung des Virus: über Tröpfchen und Kontakt; Molekulare Virologie © 2008 by Ruth Brack-Werner Progression of Smallpox Poxviruses Source: Foege, Lane, and Millar, Am J. Epi, 1969 http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Typical Progression of Smallpox Poxviruses • Incubation Period • Pre-eruptive Stage • Macules • Papules • Vesicles • Pustules •Scabs • Scars http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Smallpox Eruption Over Time Poxviruses • A major diagnostic characteristic of smallpox is that lesions in a given area are similar in appearance and feel. • Progression occurs, however, from area to area: – Pharynx, Palate. – Face. – Proximal Extremities. – Hands and Feet. http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Variolation Poxviruses Gesunde werden dem Schorf aus Pusteln erkrankter Personen ausgesetzt VORTEIL 10-fache Abnahme der Mortalität bei Pockenerkrankung ABER 2.5% der Menschen starben bei der Variolation (Cotton Mather und Zabdiel Boyleston(e) ca. 1720 in Boston: Variolation von 244 Menschen: 6 Todesfälle) Variolierte Menschen sind infektiös. TROTZDEM Befahl George Washington 1776, dass die gesamte Continental Army sich der Variolation unterzog. Molekulare Virologie © 2008 by Ruth Brack-Werner http://www.bt.cdc.gov/agent/smallpox/training/overview Vaccinia Virus Poxviruses • Origin of vaccinia virus unknown. • Genetically distinct from cowpox and variola. • May be a virus now extinct in nature. http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Smallpox Vaccine Poxviruses 1796 Edward Jenner develops vaccine 1805 Use of cows to produce vaccine 1940’s Freeze-drying technology 1965 Licensure of bifurcated needle 1971 Routine vaccination stopped in US 1975 Attenuated strains - Japan 1983 Vaccine removed from civilian market http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccine Production Poxviruses • Grown on skin of calves, sheep, water buffalo. • Lesions harvested before crusting: – Maximizes viral titer. • Pulp ground. • Mixed with 40%-60% glycerol. • Distributed in glass capillary tubes. http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner http://www.bt.cdc.gov/agent/smallpox/training/overview Dryvax Smallpox (Vaccinia) Vaccine Administration Schedule Poxviruses Schedule: • 1 Successful Dose >18 years of age. Revaccination: • 10 Years - nonhighly-attenuated vaccinia and recombinants. • 3 Years – highly attenuated orthopoxviruses. http://www.bt.cdc.gov/agent/smallpox/training/overview/ Molekulare Virologie © 2008 by Ruth Brack-Werner New Smallpox Vaccines, 2003 Poxviruses • Live vaccinia virus produced using cell culture technology. • Distributed as a freeze-dried powder. • Do not contain antibiotics. • Diluent contains glycerin and phenol. http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Smallpox Vaccine Administration Poxviruses • Vaccinia virus may be cultured from the site of a primary vaccination from 2-3 days after vaccination until the scab separates. • Care must be taken to prevent spread of the virus to other parts of the body or other persons. • Hands must be washed after every contact with the vaccination site or any materials (clothing, dressing, etc.) that has come into contact with the vaccination site. http://www.bt.cdc.gov/agent/smallpox/training/overview Molekulare Virologie © 2008 by Ruth Brack-Werner Smallpox Vaccine Adverse Reaction Rates* Poxviruses Primary Reaction Vaccination Inadvertent inoculation 25-529 Generalized vaccinia 23-242 mallpox/training/overview Eczema vaccinatum 10-39 Progressive vaccinia 0.9-1.5 Post-vaccinial encephalitis 3-12 Death 1 http://www.bt.cdc.gov/agent/s *Rates per million primary vaccinations Molekulare Virologie © 2008 by Ruth Brack-Werner Gründe für die erfolgreiche Ausrottung der Pockenkrankheit Poxviruses Mahalingam, S. et al., 2004, TRENDS Im., 637-639 Molekulare Virologie © 2008 by Ruth Brack-Werner Vaccinia Virus: Viele offene Fragen Poxviruses Was ist (sind) der Zelloberflächen- Rezeptor (e)? Wie wird die Transkription reguliert? Wir werden die mRNAs prozessiert? Wie bewegt sich das Virus? Warum kann sich das Virus in einigen Zelltypen vermehren, in anderen aber nicht? Kann man antivirale Therapien gegen Pockenviren entwickeln? Molekulare Virologie © 2008 by Ruth Brack-Werner.
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