History of Virology. Russian Bacteriologist Dimitri Iwanowski TMD tobacco mosaic disease TMV isolated and purified

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1 Viruses & Prions

2 Viruses Virus miniscule, acellular, infectious agent having one or several pieces of either DNA or RNA No cytoplasmic membrane, cytosol, organelles Cannot carry out any metabolic pathway Neither grow nor respond to the environment Cannot reproduce independently Obligate intracellular parasites

3 Viruses Viruses contain DNA or RNA And a protein coat - capsid Some are enclosed by an envelope Some viruses have spikes Infect only specific cells in a specific host Obligate parasites need living cells

4 History of Virology 1892 viruses 1 st mentioned Russian Bacteriologist Dimitri Iwanowski TMD tobacco mosaic disease Filtered plant sap Liquid still infectious 1935 TMV isolated and purified American Chemist Wendell Stanley Virus = Latin for poison

5 Virus Sizes To Study Needed to Develop: EM (TEM & SEM) Ultracentrifuge & tissue culture Most 20nm -200nm Figure 13.1

6 Growing Viruses Viruses must be grown in living cells. Bacteriophages form plaques on a lawn of bacteria.

7 Growing Viruses Viruses 1 st cultivated in live animals or in embryonated eggs Figure 13.7

8 Growing Viruses Next Tissue culture techniques were developed. Continuous cell lines may be maintained indefinitely. How do you see growing virus in living cells? Figure 13.8

9 Virus Identification Cytopathic effects - CPE Figure 13.9

10 Cytopathic Effects TMV Figure 13.3a

11 Categorizing Viruses 1. Genome compostion RNA or DNA dsdna ssrna ssdna dsrna

12 Categorizing Viruses 2. Capsid made up of capsomeres (proteins) differentiated by symmetry Helical viruses capsomeres arranged in a tube, nucleic acid inside Polyhedral viruses Icosahedral (20 faces, 12 corners) Complex viruses capsid + tail + tail fibers + pins + face plate

13 Helical Viruses Figure 13.4a, b

14 Polyhedral Viruses Figure 13.2a, b

15 Complex Viruses Figure 13.5a

16 Categorizing Viruses 3. Presence of Envelope Is the capsid surrounded by a phospholipid bilayer derived from the hosts plasma membrane

17 Viruses and Host Range Viruses have specific hosts Plant Animal Bacteria Host Range Determined by specific receptors on the host cell surface What are these used for normally?

18 Viruses named after Viral Taxonomy the disease they cause Poliovirus, mumps virus, measles virus the river near place of isolation Ebola virus Viruses that infect bacteria Are called bacteriophages Letters and numbers are used for names T 4, λ phage How are bacteria named?

19 Viral Life Cycles Bacteriophage Lytic cycle Lysogenic cycle Viruses DNA virus RNA virus Retrovirus

20 Attachment Attachment of virus to bacterial cell Entry Synthesis Assembly Release Multiplication of Viruses in a Bacterial Host Cell Entry of nucleic acid into host cell Production of nucleic acid & proteins Nucleic acid and capsid proteins assemble Viruses burst out of the host cell

21 Lytic Replication of Bacteriophages Figure 13.8

22 Lytic Replication of Bacteriophages Figure 13.8

23 Lytic Phage Replication Cycle Figure 13.9

24 Lytic cycle Phage causes lysis and death of host cell Lysogenic cycle Prophage DNA incorporated in host DNA (Temperate phages)

25 The Lysogenic Cycle Figure 13.12

26 Prophage Specialized Transduction gal gene Galactose-positive donor cell gal gene Bacterial DNA 1 2 Prophage exists in galactose-using host (containing the gal gene). Phage genome excises, carrying with it the adjacent gal gene from the host. gal gene 3 Phage matures and cell lyses, releasing phage carrying gal gene. Galactose-negative recipient cell 4 5 Phage infects a cell that cannot utilize galactose (lacking gal gene). Along with the prophage, the bacterial gal gene becomes integrated into the new host s DNA. Galactose-positive recombinant cell 6 Lysogenic cell can now metabolize galactose. Figure 13.13

27 Viral Life Cycles Bacteriophage Lytic cycle Lysogenic cycle Viruses DNA virus RNA virus Retrovirus

28 Multiplication of Viruses in an Animal Host Cell Adsorption Virus attaches to cell membrane Penetration By endocytosis or fusion Uncoating By viral or host enzymes Replication Production of nucleic acid & proteins Assembly Release Nucleic acid and capsid proteins assemble By budding (enveloped viruses) or rupture

29 Replication of Animal Viruses Same basic replication pathway as bacteriophages Differences result from Presence of envelope around some viruses Eukaryotic nature of animal cells Lack of cell wall in animal cells

30 Adsorption of Animal Viruses Chemical attraction Animal viruses do not have tails or tail fibers Have glycoprotein spikes or other attachment molecules that mediate attachment

31 Viral Spikes Figure 13.3b

32 Penetration and Uncoating of Animal Viruses Figure 13.12ab

33 Penetration and Uncoating of Animal Viruses Figure 13.12c

34 Adsorption, Penetration, and Uncoating Figure 13.14

35 Replication of Animal Viruses Each type of animal virus requires different strategy depending on its nucleic acid Must consider How mrna is synthesized? What serves as template for nucleic acid replication?

36 Replication & Assembly of DNA Virus 7 Virions are released Papovavirus 1 Virion attaches to host cell Host cell DNA Capsid 6 Virions mature DNA Cytoplasm 2 Virion penetrates cell and its DNA is uncoated Capsid proteins mrna 5 Late translation; capsid proteins are synthesized 4 Late transcription; DNA is replicated 3 Early transcription and translation; enzymes are synthesized Figure 13.15

37 Replication & Assembly in RNA Viruses Figure 13.17

38 Replication & Assembly of a Retrovirus Reverse transcriptase Capsid Envelop Virus Two identical + stands of RNA Identical strands of RNA Viral proteins RNA 4 5 Mature retrovirus leaves host cell, acquiring an envelope as it buds out. Transcription of the provirus may also occur, producing RNA for new retrovirus genomes and RNA that codes for the retrovirus capsid and envelope proteins. Provirus Host cell Reverse transcriptase Viral RNA 3 1 DNA of one of the host cell s chromosomes Retrovirus penetrates host cell. 2 Virion penetrates cell and its DNA is uncoated The new viral DNA is tranported into the host cell s nucleus and integrated as a provirus. The provirus may divide indefinitely with the host cell DNA. Figure 13.19

39 Summary of Replication & Assembly of Animal Viruses Table 13.3

40 Release of an enveloped virus by budding Figure 13.20

41 Release of Enveloped Viruses by Budding Figure 13.13

42 Assembly and Release of Animal Viruses DNA viruses - assembly in nucleus RNA viruses develop solely in cytoplasm Number of viruses produced &released depends on type of virus Size of virus initial health of host cell Enveloped viruses cause persistent infections Naked viruses released by exocytosis or may cause lysis and death of host cell

43 Latency of Animal Viruses Some viruses can remain dormant May exist for years with no viral activity, signs, or symptoms Some latent viruses do not become incorporated into host chromosome When provirus is incorporated into host DNA, condition is permanent; becomes permanent physical part of host s chromosome

44 Summary of Bacteriophage and Animal Virus Replication Table 13.4

45 Cytopathic Effects Cytopathic effects - CPE Figure 13.9

46 Causes of Cytopathic Effects 1. Host cell DNA, RNA, and protein synthesis has been stopped by the virus 2. Fusion of the plasma membranes of many cells (Herpes viruses) 3. Inclusion bodies in host cytoplasm Rabies = Negri bodies (viral particles) 4. Toxic effect of capsid proteins Mumps virus & Influenza virus 5. Host Chromosomal Disruptions Herpes virus 6. Transformation of cells into malignant cells

47 Cancer Transformed/malignant cells = uncontrolled cell division Oncovirus has oncogenic effect on the host (causes cancer) Genetic material of oncogenic viruses becomes integrated into the host cell's DNA. Activated oncogenes transform normal cells into cancerous cells. Transformed cells have increased growth, loss of contact inhibition, and T antigens.

48 Oncogene Theory Figure 13.15

49 Factors Involved in Activation of Oncogenes Ultraviolet light Radiation Carcinogens Viruses

50 How Viruses Cause Cancer Some carry copies of oncogenes as part of their genomes Some stimulate oncogenes already present in host Some interfere with tumor repression when they insert into host s repressor gene Several DNA and RNA viruses are known to cause ~15% of human cancers Burkitt s lymphoma Hodgkin s disease Kaposi s sarcoma Cervical cancer

51 Oncogenic Viruses Oncogenic DNA Viruses Adenoviridae Adenovirus - adenocarcinomas Herpesviridae EBV Burkitt s lymphoma Poxviridae Smallpox, cowpox misc. Papovaviridae Human papilloma virus cervical cancer Hepadnaviridae HBV liver cancer Oncogenic RNA viruses Retroviridae HTLV 1 (human T- lymphotrophic virus) HTLV 2 Causes leukemia in adults

52 Types of Infections Acute Viral Infections Rapid onset, short duration Influenza Latent Viral Infections Virus remains dormant and is later induced into activity HSV1 - Cold sores, varicella-zoster virus - shingles Chronic/Persistent Viral Infections Long duration, generally fatal HCV, Subacute sclerosing panencephalitis (measles virus)

53 Cytopathic effects Serological tests Detect antibodies against viruses in a patient Use antibodies to identify viruses in neutralization tests, viral hemagglutination, and Western blot Nucleic acids RFLPs PCR Virus Identification

54 Prions 1982 purposed by American neurobiologist Stanley Prusiner Infectious proteins Transmissible by ingestion, transplant, & surgical instruments Spongiform encephalopathies: Sheep scrapie, Creutzfeldt-Jakob disease, Kuru, fatal familial insomnia, mad cow disease (bovine spongiform encephalopathy) Large vacuoles in the brain

55 Prions Prion = proteinaceous infectious particle Reduced infectivity = proteases, not radiation PrP C, normal cellular prion protein (glycoprotein), on cell surface PrP Sc, scrapie protein, accumulate in brain cells forming plaques

56 Characteristics of Prions Two stable tertiary structures of PrP Normal functional structure with α-helices called cellular PrP Disease-causing form with β-sheets called prion PrP Prion PrP converts cellular PrP into prion PrP by inducing conformational change

57 Tertiary Structures of PrP Figure 13.21

58 Characteristics of Prions Normally, nearby proteins and polysaccharides force PrP into cellular shape Mutations in PrP gene result in initial formation of prion PrP When prions present, they cause newly synthesized cellular PrP to refold into prion PrP

59 Prions PrP c PrP Sc Lysosome Endosome Figure 13.21

60 All = fatal neurological degeneration, deposition of fibrils in brain, & loss of brain matter Prion Diseases Large vacuoles form in brain = spongy appearance Spongiform encephalopathies BSE, CJD, kuru Only destroyed by incineration; not cooking or sterilization

61 Scrapie in Sheep Kuru Figure 13.22

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