Virology from PID

• Unenveloped (naked) viruses have a protein capsid as a shell, within that capsid they have viral nucleic acid as well as a few enzymes which may be needed for their replication
• Enveloped viruses are composed of a lipid bilayer which is host derived

  Embedded in the lipid bilayer are viral spike proteins

  These proteins are important in terms of allowing the virus to enter cells

  Spike proteins are also recognised by antibodies
• Either DNA or RNA viral genome

  Both can be either ss (single stranded) or ds (double stranded)

  most are DNA ds
• Ss RNA viruses are more complicated as they can have different polarity.
• Positive sense – same sense as messenger RNA. RNA can go straight into a cell and start making viral proteins (doesn’t need to replicate before it makes its proteins)
• Negative sense – viruses do need to replicate, copy their negative sense RNA genome to make their mRNA which makes protein.
• RNA viruses

  Need an RNA polymerase to copy their RNA genome (no equivalent enzyme in the host)

  RNA dependent RNA polymerase

  RNA polymerases are error prone

  No proof-reading capability

  More mutation/changes as a result – more advantageous strain becomes dominant

  DNA viruses don’t mutate/adapt well as much
• Consequence of RNA viruses/RNA polymerase

  RNA viruses are more variable

  Within a species of virus are more subtypes/serotypes

  Can evolve rapidly if need

  If a virus jumps from one species to another, RNA viruses can more readily adapt

  Often zoonotic (jump from animals to humans)
• Capsid Structure

  Protein coat that encloses the nucleic acid

  Three Capsid types:

  Icosahedral – organised spherical structure

  Helical

  Complex – pox viruses
• Complex Capsid:

  Some of the large viruses have capsid structures that are more complex

  Poxviridae

  >100 proteins

  Neither helical or icosahedral structure

  Enveloped, brick-shaped or ovoid virion

  Surface membrane displays surface tubules or surface filaments

  Examples: Smallpox, Mouse pox
• Virus Envelope is present on:

  Few viruses with icosahedral capsid

  All viruses with helical capsid

  Complex capsid
• Enveloped viruses:

  Enveloped viruses acquire envelope in the final part of their lifecycle as they emerge from the infected cell. They bud out by exocytosis through the host cell membrane

  Viral envelope contains host cell lipid bilayer as well as viral proteins

  Viral proteins contain receptors needed for virus entry

  envelope is basically a coat of the host cell membrane

  
• Unenveloped:

  Naked viruses are released by lysis of the infected cell (cell breaking open and releasing the virus, therefore no budding, no envelope as it doesnt go through the membrane, its simply released into the extracellular space to search for another cell to hijack)

  Cause apoptosis so that viral cells can escape and hijack more cells to infect further

  Viral receptors are present on the capsid surface
• Biological properties of enveloped viruses:

  • Lipid bilayer is very fragile
  • affected by heat, dryness
  • do not survive long in environment
  • More fragile than viruses with just a capsid
  • More easily destroyed by
  • Detergents
  • Disinfectants
  • Outside environment
  • If the envelope is destroyed, then the virus is not infectious
  • Destroys the receptors needed for entry
• unenveloped viruses are very robust

  • remain viable in the environment for a long time, even months
  • e.g. norovirus
• Unenveloped Viruses

  • composed of protein
  • Environmentally stable to
  • temperature
  • pH
  • proteases
  • detergents
  • drying
  • Resistant to detergents
  • can dry out but retain infectivity
  • easily spread by aerosols
  • can survive harsh conditions e.g. gut
  • lyse cell to release virus, therefore must kill cell
  • usually cause acute infections
• Enveloped Viruses:

  • composed of lipids, proteins and glycoproteins
  • environmentally liable/destroyed by
  • acid
  • detergents
  • drying
  • heat

  Must stay wet

  not easily spread (large droplets, secretions, transfusions/transplants)

  cannot survive in GIT

  released by budding therefore doesnt kill cell in order to spread

  can cause persistent infections
• Criteria used for classifying viruses:

  • Nature of the viral genome
  • DNA or RNA; Polarity of nucleic acid
  • Structure of nucleic acid (ss or ds); linear or circular; segmentation
  • Structure and symmetry of the viral nucleocapsid
  • Icosahedral; Helical; Complex
  • Presence or absence of an envelope
  • Size and Morphology
  • Genome organisation and different coding strategies
  • Tissue and cell tropism
  • Varying pathogenicity
• Viral Proteins:

  • Structural proteins:
  • Capsid proteins
  • Envelope proteins
  • Matrix protein (layer inside the envelope and outside capsid)
  • Virion associated enzymes
  • Non-structural proteins:
  • Proteins that are not structural components of the virus
  • Often enzymes (but some enzymes can be structural)
  • Also some viruses encode regulatory proteins, oncoproteins, etc.
• Function of the Virus Capsid:

  Structural component of the virus capsid (icosahedral and helical and some complex viruses

  Protect the viral nucleic acid and deliver the viral nucleic acid to the cell

  Capsids of naked viruses contain receptors that attach to the host membrane to allow entry

  Contains sites that will induce an antibody response
• Steps in the Virus Life Cycle:

  • Attachment
  • Entry
  • Receptor mediated endocytosis
  • Cell membrane fusion
  • Uncoating
  • At the plasma membrane
  • In endosome by changes in pH
  • Viral Gene Transcription
  • Genome Replication
  • Translation
  • Assembly
  • Release
  • Non enveloped viruses by cell lysis
  • Enveloped viruses by budding from the plasma membrane
• Virus Attachment:

  Attachment to host cell is a highly specific process

  Involves complimentary receptors on the surface of a susceptible host cells- Highly specific

  Receptor can be protein, carbohydrate or lipid

  Initial binding is reversible - hydrostatic bonding

  Once bound, may cause a conformational change that then allows binding to a co-receptor
• Virus Entry:

  Viruses must cross the plasma membrane to enter the host cell

  Entry into the cell by either

  Cell membrane fusion (Non–endocytotic pathways)

  where the virus particle fuses with the membrane of the cell

  Receptor mediated endocytosis (Endocytotic pathways)

  Virus particle taken inside the cell in a vesicle
• Cell Membrane Fusion

  Virus membrane fuses with plasma membrane and nucleocapsid is released into cytoplasm

  Occurs at neutral pH (pH independent fusion)

  Examples - HIV, herpes virus

  
• Receptor Mediated Endocytosis

  • Virus particle binds to host cell receptors
  • Enters cell in an endosome
  • Virus membrane fuses with membrane of the endosome and nucleocapsid is released into cytoplasm

  
• Replication:

  • DNA viruses
  • dsDNA viruses use host machinery in the nucleus (except poxviruses) to make more ds DNA (that’s where the enzymes for replication are)
  • ss DNA converted to ds DNA then replicates like ds DNA
  • RNA viruses
  • replicate in the cytoplasm (except influenza and retroviruses)
  • Have own RNA polymerase so dont need to be in nucleus
  • All make viral mRNA which then migrates into the cytoplasm to synthesis viral proteins using the host ribosomes
• Virus Assembly:

  Translation of viral proteins in the cytoplasm

  Assembly of virus capsids from newly synthesised components (de novo assembly)

  Encapsidation of the viral nucleic acid

  viruses are made by assembly rather than division

  The virus particles make all their components, so they make more nucleic acid, they make more structural proteins, capsid, proteins, envelope proteins. And those particles are built by assembly

  
• Virus Release:

  Enveloped viruses by budding from the plasma membrane

  Acquire the envelope on the way out from plasma membrane or internal membranes (nucleus, ER)

  Non enveloped viruses released by cell lysis
• How is a virus recognised as foreign?

  • Innate Immune responses recognises Pathogen-Associated Molecular Patterns (PAMPs)
  • recognised as forgein as there are bits of nucleic acids that are just not normally found in mammalian cells or in other organisms
  • Un-methylated C-G dinucleotides (CpG motifs) on DNA viruses
  • high frequencies in viruses
  • low frequency in mammalian cells
  • Double-stranded RNA (only RNA viruses produce this)
  • Uracil-rich, single-stranded RNA
• What host cell receptors are involved

  Pattern Recognition Receptors (PRRs)

  Cytoplasmic PRRs

  Toll-like receptors (TLRs)

  Bind to viral products and activate a signalling cascade that results in the synthesis of cytokines that block virus replication in the infected host
• Toll-like receptors (TLRs)

  • Found on the surface of cells and within endosomes of phagocytic cells (Neutrophils and APCs)
  • Detect extracellular viruses
  • Can be either
  • On the surface of cells (TLR-1, 2, 4, 5 & 6)
  • In the endosome (TLR-3,7, 8 & 9)
  • TLR-3 binds ds RNA
  • TLR-7 binds Uracil-rich ss RNA (e.g. HIV)
  • TLR-8 binds ss RNA
  • TLR-9 binds CpG motifs within viral DNA
  • Sense the presence of viral nucleic acid and other conserved molecular components of invading pathogens
• Interferons

  Most important of the broad host defences against viral infection

  Protects adjacent cells from infection

  Inhibition of viral replication

  Helps activates T-cell mediated immunity

  Activation of macrophages

  Up-regulates MHC receptors on virus-infected cells
• IFNα and IFNβ (type 1)

  Produced by most cell types early in infection (Innate response) - protect cells

  Activate genes that have antiviral activities

  dsRNA dep protein kinase R

  RNase L

  destroy viral RNA in the cell

  Helps stimulate MHC class I (enhance presentation of viral peptides to T cells)

  Activates NK cells

  Induces apoptosis
• IFNγ (type 2, proinflammatory)

  Involved in the regulation of nearly all phases of immune and inflammatory responses (both innate and adaptive)

  Produced by NK cells and T lymphocytes

  Enhances MHC expression on APCs

  More important as an immunoregulator than as an antiviral agent

  Enhances the cytotoxic activity of T cells, macrophages and NK cells
• Components of the Adaptive Immune Response

  • Humoral Immune response (B lymphocytes)
  • Days
  • Antibody production
  • Cellular Immune Response (T lymphocytes)
  • Days
  • Cytotoxic T cells (CTLs) CD8+
  • Kill virus-infected cells
  • Cytokines that eliminates viral RNA
  • T Helper cells (TH) CD4+
  • Activate macrophages
  • Cytokines
• Components of the Adaptive Immune Response - B and T cells

  Elicit several kinds of response

  Eliminate virus

  Destroy virus-infected cells

  Prevent re-infection

  Virus-specific immune response

  Recognise viral proteins and carbohydrates (antigens)

  So the antibody response (HUMORAL) gets rid of virus that circulating and outside cells, whereas a cellular response will eliminate virus in the infected cells.
• Humoral Immune Response

  B lymphocytes

  Resident in the lymphatic tissue

  Respond to antigenic stimulus by producing and secreting antibodies

  Carry highly specific receptors that recognise viral epitopes

  Recognise Ag in their native form (no processing)

  
• B cell activation

  • get clonal amplification of that B cell with that antibody binding site, those B cells will then go on to produce that particular antibody.
  • Some of those antibodies will become memory cells and give the host lifelong immunity

  
• Production of antibodies

  IgM is the first antibody that is produced then class switch to IgG and gain memory, massive secondary response

  IgG and IgA predominate in the secondary response
• IgG = Main circulating antibody - important in longterm immunity

  IgM = circulating antibody produced early in infection

  IgA = secretory antibody - primary defense at mucosal surface