Micro ch 13

  1. Describe viruses
    • can be viewed as genetic info - either RNA or DNA (never both) - contained within a protective coat
    • incapable of metabolism, replication, or motility
    • must have host cell
  2. How can viruses be categorized?
    • The International Committee on Viral Taxonomy:
    • Key characteristics used currently are the genome structure (type of nucleic acid and strandedness) and the hosts they infect (bacteria, archaea, animals, plants)
  3. Bacteriophages
    • Also referred to simply as phages
    • Refers to the group of viruses that infect bacteria
  4. Virion
    • viral particle
    • consists of nucleic acid surrounded by a protein coat
  5. capsid
    • protein coat of viruses
    • protects the nucleic acid from enzymes and toxic chemicals in environment
    • Also carries enzymes required to infect host
    • Composed of capsomers
  6. capsomers
    identical protein subunits arranged in precise manner to form capsid
  7. Nucleocapsid
    the capsid together with nucleic acid it encloses
  8. 2 types of viral "architecture"
    Naked & enveloped
  9. Enveloped viruses
    • Envelope composed of a lipid bilayer outside of capsid
    • Sandwiched btwn envelope is matrix protein, which is unique to enveloped viruses
  10. Naked viruses
    • Those viruses without an "envelope" (duh)
    • Nearly all phages are naked
  11. spikes
    • Protein structures on animal viruses which stick out either the lipid bilayer of enveloped viruses or the capsid of naked viruses 
    • Allow for attachment
  12. 3 general shapes of viruses
    • Icosahedral
    • Helical
    • Complex
  13. Icosahedral virus
    • type of virus shape which appear spherical when viewed with electron microscope, but surface is actually 20 flat triangles arranged in similar manner as soccer ball
    • Efficientt design for any container that uses identical sub-units and requires the least energy to assemble
  14. Helical virus
    • type of virus shape that appears cylindrical when viewed with electron microscope
    • Capsomers are arranged in helix, somewhat similar to spiral staircase.
    • Some are short and rigid, others are long and filamentous
  15. Complex virus
    • type of virus shape that has intricate structures
    • Phages are most common example
    • Have icosahedral nucleocapsid as "head" and long helical protein component as "tail"
    • Has "tail fibers" that look like legs
  16. DNA viruses that have double-stranded DNA and NAKED
    • Papillomaviridae - Human Papillo..viruses
    • Polyomaviridae - Merkel cell skin cancer
    • Adenoviridae - Human adeno respiratory inf.
  17. DNA viruses that have double-stranded DNA and ENVELOPED
    • Herpesviridae - Herpes zoster & simplex
    • Poxviridae - Small pox virus
  18. DNA viruses that have single-stranded DNA and are NAKED
    Parvoviridae - Human parvo (5th disease)
  19. RNA viruses that are double stranded and are NAKED
    Reoviridae - Human rotavirus (diarrheal disease)
  20. RNA viruses that are single-stranded (Plus strand) and NAKED
    • Picornaviridae - Polio, rhino, hep A
    • Caliciviridae - Norovirus (gastrointestinal)
  21. RNA viruses that are single-stranded (Plus strand) and ENVELOPED
    • Togaviridae - Rubella, Chikugunya
    • Flaviviridae - Yellow fever, dengue, hep c
    • Coronaviridae - Coronavirus (SARS)
  22. RNA viruses that are single-stranded (minus strand) that are ENVELOPED
    • Rhabdoviridae - Rabies
    • Filoviridae - Ebola
    • Paramyxoviridae - Mumps, measles
    • Orthomyxoviridae - Influenza
    • Bunyaviridae - Hantavirus
    • Arenaviridae - Lassa virus
  23. Reverse transcribing viruses - DNA & ENVELOPED
    Hepadnaviridae - Hep B
  24. Reverse Transcribing Viruses - RNA & enveloped
    Retroviridae - AIDS (Human immunodeficiency virus)
  25. Grouping of human viruses based on route of transmission
    • Enteric - Fecal/oral route (enteroviruses, diarrhea, polio)
    • Respiratory - Respiratory or salivary route (influenza, measles, rhino - colds)
    • Zoonotic - Vector (such as arthropods) sandfly, west nile. Also animal to human directly = rabies, cowpox
    • Sexually transmitted - duh
  26. Arboviruses
    • meaning arthropod borne
    • viruses spread by arthropods
    • Ex: mosquitos, ticks, sandflies
    • Cause disease such as west nile, La crosse enceph, yellow fever
  27. 3 types of different relationships bacteriophages can have with their hosts:
    • Lytic phage
    • temperate phage
    • filamentous phage
  28. Lytic phages
    • also called virulent phages
    • Exit the host at end of infection cycle by lysing the cell
    • result in formation of new virus particles
    • Called productive infection

    Consists of 5 steps
  29. 5 step process of lytic phage
    • Attachment
    • Genome entry
    • Synthesis
    • Assembly
    • Release
  30. Attachment step of lytic phage
    • Upon contact, phage particle attaches by means of protein on tail to receptor on cell surface or to an appendage, such as pilus
    • *cells that lack the receptor used by a given phage are resistant
  31. Genome entry of lytic phage
    • Bacteriophage injects it's genome into cell
    • Tail contracts & phage DNA injected inside cell, leaving phage coat outside
  32. Synthesis step of lytic phage
    • Phage genome is transcribed and phage proteins synthesized
    • Phage DNA is replicated, other virion components are made and host DNA is degraded
  33. Assembly step of lytic phage
    phage components are assembled into mature virons
  34. Release
    • Phage makes lysozyme, which is enzyme that digests host cell from within, causing cell to lyse, thereby releasing phage
    • Entire process takes about 30 mins
  35. Burst size
    number of phage particles released after bacterial cell is lysed
  36. Lysogeny
    characterized by integration of the bacteriophage nucleic acid into the host bacterium's genome or formations of a circular replicon in the bacterium's cytoplasm.
  37. Temperate phage
    • have the option of either directing a lytic infection (producing infection) or incorporating DNA into host cell genome (lysogenic infection)
    • Which cycle occurs is largely random
  38. Lysogenic infection
    refers to temperate phage infection in which virus incorporates its DNA into host cell genome

    Lysogen refers to the infected cell
  39. What is the most thoroughly studied temperate phage
    • Lambda ( λ )
    • Has a linear DNA chromosome that join together inside host to form circular molecule
    • From there, it will either enter lytic or lysogenic cycle
  40. Prophage
    The integrated phage DNA that gets replicated along with the host chromosome prior to cell division
  41. What happens in temperate phage infections if it goes into lysogenic cycle
    • The integration process uses enzyme called integrase to insert phage DNA into host cell chromosome at specific site (process called site-specific recombination)
    • Cell divides
    • Excision of phage DNA from host DNA
    • Goes to lytic infection
  42. In the temperate phage infections, what happens if it goes to the lytic infection...
    • Phage attaches to bacterium
    • Injected linear phage DNA circularizes and enters lytic infection
    • Replication of phage DNA and synthesis of phage-encoded proteins
    • Cells lyse, releasing new phage
  43. Repressor
    • Involved in temperate phage infection
    • A protein when prevents expression of the gene required for excision, therefore essential for maintaining the lysogenic state
  44. What happens if a lysogenic culture is treated with DNA-damaging agent like UV light
    • Destroys the repressor protein responsible for maintaining the integration of prophage
    • The prophage is excised from the chromosome, allowing the phage to enter lytic cycle, and productive infection results
  45. Phage induction
    • refers to the release of the phage from a damaged cell
    • similar to rats fleeing a sinking-ship
  46. Consequences of Lysogeny
    Immunity to superinfection (infection by same phage) - if another λ phage injects its DNA into a λ lysogen, DNA will not be expressed

    Lysogenic conversion - a change in the phenotype of a lysogen as a consequence of specific prophage it carries. Ex: only lysogenic strains of S. pyogenes and C. Botulinum produce toxins
  47. Filamentous phages
    • single-stranded DNA phages that look like long fibers
    • cause productive infections, but does not kill host
    • infected cells grow more slowly than uninfected counterparts
  48. What occurs during first half of filamentous phage infection
    • phage attaches to bacterial cell & injects it's single-stranded DNA
    • Host cell DNA polymerase synthesizes complementary strand
    • The double-stranded DNA is referred to as replicative form (RF) 
    • One strand of RF is then used as temple to make mRNA as well as multiple copies of phages single-stranded genome (ssDNA)
  49. In filamentous phage infection....
    Explain the + and - strands
    • when the RF is being replicated, the + strand DNA functions as phage genome
    • The - strand DNA is transcribed into mRNA, which mRNA is then translated into phage coat proten
  50. What happens at end of filamentous phage infection
    • Phage DNA is extruded from cell
    • Phage coat protein molecules are inserted into host cell's cytoplasm
    • At same time, other phage-encoded proteins form pores that span cytoplasmic and outer membrane
    • Phage DNA is secreted through pores, when coat it to form nucleocapsid
  51. Two types of transduction
    generalized and specialized
  52. Generalized transduction
    • results from a packaging error during phage assembly
    • Some phages degrade bacterial chromosome into many fragments during lytic infection, can be mistakenly packaged into phage head during assembly
  53. Generalized transducing particles
    • refers to a phage head that contains only bacterial genes instead of phage genes (by accident)
    • Can't direct a phage replication cycle
  54. What happens to generalized transducing particles when released from phage-infected host
    • Bind to another bacterial cell and injects it's DNA
    • DNA may integrate into cell by homologous recombination, replacing DNA of host cell
    • Any gene of the donor cell can be transferred this way, which is why this mechanism is called generalized transduction
  55. Specialized transduction
    • A temperate phage injects it's DNA into bacterial host
    • Phage DNA integrates into host cell DNA to become prophage
    • When prophage is excised from bacterial chromosome, a mistake is made; some bacterial DNA flanking the phage DNA is taken and a piece of phage DNA is left behind
    • Replication and assembly produces defective phage particles that carry certain bacterial DNA in place of some phage DNA
    • DNA of defective phage is injected into new host, but can't cause productive infection
    • Bacterial DNA integrates into host genome via homologous recombination; can now be replicated along with rest of DNA
  56. Defense mechanisms of bacteria to resist invasion by viruses
    • Altering receptor sites
    • Restriction modification systems
    • CRISPR system
  57. Explain bacteria altering receptor sites
    • recall first step in phage infection is attachment to specific receptors on cell surface
    • If bacterium alters or covers receptor, is resistant
  58. Restriction - Modification System
    • System that protects bacteria from phage infection by quickly degrading incoming foregin DNA
    • Do this by combined action of 2 types of enzymes: restriction and modification enzymes
  59. In restriction-modification system:
    What does restrictive enzyme do?
    What does modification enzyme do?
    • The restriction enzyme recognized specific short nucleotide sequences and then cuts the DNA molecule at those sequences. 
    • These enzymes explain why some bacteria can degrade foreign DNA and also play important role in biotech.

    Modification enzyme protects the host cell DNA from action of restriction enzyme by adding methyl groups to the nucleobases recognized by restriction enzyme, which makes them invisible to restriction enzyme
  60. What does CRISPR stand for
    Clusters of Regularly Interspersed Short Palindromic Repeats
  61. CRISPR
    • type of bacterial defense system against viruses
    • bacterial cells that survive some phage infections retain small segments of phage DNA, incorporating them into bacterial genome
    • Segments of phage DNA - called spacer DNA - are inserted into segment called CRISPR
    • Provides historical record of past phage infection, allowing bac cell & it's progeny to recognize & block subsequent infection by same type of phage
  62. How does CRISPR system work
    • It's not yet clear, but may function by type of RNA
    • The CRISPR array, including spacer sequence, is transcribed & cut into small RNA's called crRNA's
    • crRNA's bind to complexes called Cas (CRISPR-associated sequences)
    • When spacer RNA in Cas base pairs w nucleic acid of invading phage, that nucleic acid is targeted for destruction
    • CRISPR can also recognize & destroy other foreign DNA that bacterial strain has encountered in past
  63. plaque assays
    • routinely used to quantitate phage particles in samples such as sewage, seawater and soil
    • A double layer agar is used
    • Top layer, the soft layer, is inoculated with both the bacterial host and phage containing specimen. Is poured over surface of agar-filled petri dish
    • Bacteria present multiply rapidly, producing dense lawn of bacterial growth
    • Any phages absorb susceptible bac & lyse them, releasing progeny phage that diffuse and infect neighbor bac.
    • Plaques form due to cell lysis caused by phage
  64. Plaques
    • circular zones of clearing
    • formed in lawn of plaque assay due to cell lysis caused by phage
    • Each plaque represents a plaque-forming unit (PFU) initiated by a single phage particle infecting a cell
  65. Titer
    the concentration of infectious phage particles in the original phage suspension
  66. What is the 5 step process of animal virus replication
    • Attachment (absorption)
    • Penetration and uncoating 
    • Synthesis of viral proteins & replication of genome
    • Assembly
    • Release
  67. Animal Virus Replication:
    • Animal viruses surfaces are studded with attachment proteins/spikes
    • These proteins/spikes attach to host cell receptors on membrane 
    • *Because a virion must bind to specific receptors, particular virus may be able to infect only single or limited # of cell types, and most viruses can infect only a single species
    • *This accounts for the resistance some animals have to certain diseases
  68. Animal Virus Replication:
    Penetration and uncoating
    • This mechanism is different depending on whether virus is naked or enveloped
    • In all cases, the entire virion is taken into cell
    • *differs from most phages, where only nucleic acid enter and capsid stays outside in bacterim
  69. Animal Virus Replication:
    Penetration & uncoating: 
    Enveloped viruses
    Enter host cell by 1 of 2 mechanisms: fusion with host membrane or endocytosis
  70. Animal Virus Replication:
    Penetration & uncoating:
    Enveloped viruses - fusion with host membrane
    • Absorption: spikes of virion attach to specific host cell receptors
    • Membrane fusion: envelope of virion fuses with plasma membrane
    • Nucleocapsid released into cytoplasm: viral envelope remains part of plasma membrane
    • Uncoating: Nucleic acid separates from capsid
  71. Animal Virus Replication:
    Penetration & uncoating:
    Enveloped viruses - endocytosis
    • Viruses that enter this way exploit the process of receptor-mediated endocytosis, a normal mechanism by which cells bring certain extracellular material into the cell
    • Absorption: attachment to receptors triggers endocytosis
    • Endocytosis: Plasma membrane surrounds the virion, forming an endocytic vesicle
    • Release from vesicle: envelope of virion fuses with the endosomal membrane
    • Uncoating: nucleic acid separates from capid
    • (pg 318)
  72. Animal Virus Replication:
    Penetration and uncoating:
    Naked viruses
    • *have no lipid envelope & can't fuse to host membrane to enter. Therefore, only enter by endocytosis
    • ONce in the endosome, virus damages vesicle membrane, resulting in nucleocapsid being released into cytoplasm
  73. Animal Virus Replication:
    Refers to the nucleic acid separating from it's protein coat prior to the start of replication (after penetration & uncoating)
  74. Animal Virus Replication:
    Synthesis of viral proteins and replication of genome
    • Production of viral particles in infected cells requires 2 interrelated events:
    • expression of viral genes to produce structural and catalytic proteins
    • Synthesis of multiple copies of the viral genome
  75. Animal Virus Replication:
    Replication strategy of viruses divided into 3 general categories based on those used by:
    • DNA viruses
    • RNA viruses
    • reverse transcribing viruses
  76. Animal virus replication:
    Replication of DNA viruses
    • DNA viruses generally replicate in the nucleus and use the host cell machinery for DNA synthesis as well as gene expression, although they often encode their worn DNA polymerase
    • Replication of ssDNA viruses is similar to that of double-stranded DNA, but the complementary strand must be synthesized first
  77. What virus is the exception to the rule that DNA viruses encode in host nucleus
    • Pox viruses
    • replicate in cytoplasm, encode all the enzymes and factors necessary for DNA and RNA synthesis
  78. Animal Virus Replication:
    Replication of RNA viruses
    • vast majority of RNA viruses are single stranded and usually replicate in cytoplasm
    • Requires a virally encoded enzyme replicase to synthesize complementary RNA strand
  79. Replicase
    • Enzyme required for replication of RNA viruses
    • Lacks proofreading abilities, so makes more mistakes than DNA polymerases
    • Because of high number of mutations generated, some RNA viruses can adapt more quickly to selective pressures of DNA counterparts
  80. Antigenic drift
    • occurs as mutations accumulate in genes encoding key viral surface proteins that are recognized by the immune system
    • due to errors caused by replicase enzyme
    • because of these changes, a person whose immune response protected him/her against flu one year may not protect against the variant that cirulates the next year
  81. Antigenic shift
    happens when 2 different strains of a segmented virus infect he same cell and reassortment occurs, meaning the viral particles that exit the cell have various combinations of segments from the initial infecting strains
  82. Replication of reverse-transcribing viruses
    • Encode the enzyme reverse transcriptase
    • RNA molecule is used as template for DNA
    • Are error prone
  83. Retroviruses
    • Includes HIV
    • have the + strand RNA genome & carry reverse transcriptase within the virion
    • After entering host, reverse transcritase uses RNA as template to make 1 DNA strand
    • Complement to that strand is then used to make double stranded DNA, which integrates into host chromosome
    • Once integrates, genome can direct infection or remain latent
    • Can't be eliminated from cell once DNA copy is made
  84. Animal Virus Replication:
    • Involves same general principles as phages
    • Protein capsid must be formed, then genome & any necessary enzymes packed within
  85. Animal Virus Replication:
    • Just as entry depends on whether virion is enveloped or naked, so does it's release
    • Most enveloped are released by budding
    • Naked are released as cell dies
  86. Budding
    • Process by which enveloped viruses are released:
    • Viral proteins that will become envelope spikes insert into host plasma membrane
    • Viral matrix protein coats inside of plasma membrane
    • Nucleocapsid extrudes from the host cell, becoming coated with matrix proteins and envelope with protein spikes
    • New virus is released
    • (Pg 321)
  87. Apoptosis
    programmed cell death
  88. How are naked viruses released???
    • As the cell dies
    • Some viruses trigger apoptosis prior to release of virus particles
    • Immune response of animal can also lead to same process
  89. Categories of animal virus infections
    • relationship btwn viruses & animal hosts can be divided into 2 major categories of infections:
    • Acute and persistent
  90. Acute infections
    • characterized by the sudden onset of symptoms of a relatively short duration
    • Ex: flu, mumps, poliomylitis
  91. Persistent infections
    • infection that can continue for years, or even the life of host with or without symptoms
    • Person can transmit disease to others even without sx
    • 2 different types: chronic or latent
  92. Chronic infections
    characterized by continuous production of low levels of viral particles
  93. Latent infections
    • Viral genome remains silent within host cell, yet can reactivate to direct productive infection
    • Some do not integrate into host cell chromosome; rather they replicate independently as host genome, like plasmid
  94. Provirus
    • the silent viral genome that can replicate independently of host genome, much like plasmid
    • explains why cold sores can recur
  95. Examples of persistent, chronic infections
    • Hep B = hepatocytes (liver cells) = hepatitis, cirrhosis
    • Hep C virus = Hepatocytes = hepatitis, cirrhosis
    • HIV = Activated helper T cells, Macrophages = AIDS
  96. Examples of persistent, latent infections
    • Herpes simplex type 1 = neurons of sensory ganglia = oral herpes
    • Herpes simplex type 2 = neurons of sensory = genital herpes 
    • Varicella zoster = satellite cells of sensory ganglia = chickpox & shingles
    • Cytomegalovirus = salivary glands, kidney, epithelium, WBC = CMV pneumonia, eye inf
    • Epstein-Barr virus = B cells = Burkitts lymphoma
    • HIV = Memory helper T
  97. oncogenes
    • viruses that carry genes which are very similar to DNA sequence to proto-oncogenes (genes that stimulate cell growth)
    • their entry into cells can interfere with cell's own mechanisms, leading to tumor formation
  98. cell culture
    • also called tissue culture
    • commonly used to cultivate most animal viruses
  99. primary cultures
    • refers to cells obtained from animals by removing tissue & processing it to get individual cells
    • these cells are then grown on petri dish with liquid nutrient medium
    • One prob is that normal cells only divide limited # of times. Therefore, tumor cells are ofen used
  100. cytopathic effect
    observable change in a cell in vitro caused by a viral action such as cell lysis
  101. inclusion body
    • A distinct region in the infected cell that is the site of viral replication
    •  certain viruses cause infected cell to form it
    • position depends on type of virus
  102. Quantitating animal viruses...
    most precise method for determining concentration of animal viruses in sample
    • the plaque assay
    • a monolayer of tissue culture cells is the host
    • *clear zones surrounded by uninfected cells are counted to determine the viral titer
  103. quantal assay
    • can be used to estimate a viral titer
    • several dilutions of virus prep are administered into number of animals, cells, or chick embryos (depending on host specificity)
    • The titer of virus, or endpoint, is dilution at which 50% of inoculated hosts are infected or killed
    • Can be reported as ID50(infective dose) or LD50(lethal dose)
  104. hemagglutination
    • the phenomenon that certain viruses cause RBC to agglutinate
    • occurs when individual viral particles attach to surface molecules of multiple RBC simultaneously, connecting cells to form an aggregate
    • Visible only at high concentrations
    • Ex: Orthomyoxviruses
  105. Viroids
    • consist solely of single-stranded RNA molecule 
    • Only infect plants
    • Enter through wound sites
  106. Prions
    • composed solely of protein
    • have been linked to number of slow, always fatal, human diseases as well as animal diseases
    • Prion proteins accumulate in neural tissue
    • Neurons die & brain function deteriorates as tissues develop holes
  107. Human Prion diseases
    • Kuru = caused by cannibalism
    • Variant Creutzfeldt-Jakob dis = consuming prion-contaminated beef
    • Creutzfeldt-Jakob dis = inherited
    • Gerstmann-straussler scheinker syndrome = inherited
    • Fatal familial insomnia = inherited
  108. Animal Prion diseases
    • Scrapie - goats/sheep
    • Bovine spongiform encephalopathy - cattle
    • Chronic wasting disease - deer/elk
    • Transmissible mink encephalopathy - ranch mink
    • Feline spongiform encephalopathy - cats
  109. Transmissible spongiform encephalopathies
    • refers to sponge like appearance of brain tissue full of holes 
    • refers to all prion diseases
  110. How do prions accumulate in tissue and cause disease is they lack nucleic acid?
    • Animals synthesize a protein in neurons, especially in brain, that has nearly identical amino acid sequence to infectious prions
    • Key difference btwn normal cellular prion protein and the infectious form is the shape of the protein, which influences it stability
    • Normal form, PrPc (for prion protein, cellular) is deadily destroyed by host cell proteases, so has turnover process
    • In infectious prion proteins, PrPSC (for prion protein, scrapie) less susceptible to degradation by proteases & become insoluble
Card Set
Micro ch 13
micro ch 3