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Title: RNA VIRUSES


1
RNA VIRUSES
2
Picornaviridae
3
  • The smallest RNA-containing viruses known.
  • Comprise one of the largest (230 members) and
    most important families of human and agricultural
    pathogens.
  • The family is currently divided into five genera
    Rhinoviruses, Enteroviruses, Aphthoviruses,
    Cardioviruses, and Hepatoviruses (REACH).

4
  • Aphtovirus (foot-and-mouth disease virus)-
    Infects cloven-hoofed (footed) animals and
    occasionally humans.
  • Cardiovirus- Infects rodents
  • Three genera Rhinovirus, Enterovirus, and
    Hepatovirus include primary human pathogens with
    numerous serotypes.

5
Genus Virus Serotypes
Rhinoviruses gt 100
Enteroviruses
Polio viruses 3
Coxsackie viruses A B 23 6 1-22, 24 (23 echorviruses 9) 1-6
Echoviruses 28 1-7,9,11-21, 24-27,29-33 Echo 8 Echo 1 Echo 10 Reovirus 1 Echo 28 Rhino 1A Echo 34 Coxsackie A 24 Echo 22,23 Parechovirus
Parechoviruses 2 Previously Echo 22 and 23
Enteroviruses 68-71 4 New naming system since 1967
Hepatovirus Hepatitis A virus 1 Previously enteroviruses 72
Total 67
6
  • Enteroviruses and Hepatovirus differ from
    Rhinoviruses in
  • - Stability at pH 3
  • - Optimum T º of growth
  • - Mode of transmission
  • - Diseases caused

7
Enterovirus Particles
8
  • The virion is roughly spherical, naked, and range
    in diameter from 24 to 30 nm.
  • 60 subunits make up the icosahedral capsid each
    of which is composed of four polypeptide chains,
    VP1-VP4
  • VP1, 2 and 3 are exposed at the virion surface,
    whereas VP4 lies buried in close association with
    the RNA core.

9
  • Of the four proteins, VP1 exhibits the greatest
    sequence variability and VP4 the least.
  • VP1 is also the dominant protein, playing key
    roles in surface topography, antigenicity,
    receptor binding, and probably viral uncoating.
  • Genome is a ss RNA ( 7500 nt) of positive
    polarity, polyadenylated at 3- and has a protein
    of 22 to 24 amino acids (VPg) at the 5- end.

10
Receptors
  • The receptors for polioviruses, Coxsackie
    viruses, echoviruses, and the major serogroup of
    rhinoviruses have all been mapped to human
    chromosome 19.
  • The receptors for polioviruses and human
    rhinoviruses have been identified as members of
    the Ig superfamily, whereas the receptor for
    echoviruses has been identified as a member of
    the integrin family.

11
  • Virus Receptor
  • - Rhinovirus (major) ICAM-1
  • - Rhinovirus (minor) LDL-R
  • - Polioviruses PVR (similar
    to

  • ICAM-1) CD155
  • - Coxsackie A ICAM-1
  • - Coxsackie B Unknown
  • - Echovirus DAF, VLA-2

12
Enteroviruses
  • Arildone (pleconaril) contains a 3-methyl-
    isoxazole group that binds to the floor of the
    VP1 canyon and alters its conformation to prevent
    the uncoating of the virus.
  • Poliovirus produces a protease that degrades the
    200.000 Dalton cap-binding protein of eukaryotic
    ribosomes, thereby blocking the translation of
    cellular mRNA.

13
Orthomyxoviridae
14
INFLUENZA VIRUS
15
ORTHOMYXOVIRUSES
type A, B, C NP, M1 protein sub-types HA or
NA protein
16
  • Among the RNA viruses, influenza is very special
    in that all of its RNA synthesis take place in
    the nucleus.
  • Short capped primers are generated from host cell
    RNAs by an influenza virus-encoded cap-dependent
    endonuclease.
  • Influenza virus mRNAs undergo splicing in the
    nucleus.

17
Classification
  • The family contains two genera Influenza A and B
    viruses, and influenza C virus.
  • Genera are distinguished on the basis of
    antigenicity of nucleoprotein (NP) and matrix (M)
    proteins.
  • Influenza A viruses are divided into subtypes
    based on the antigenicity of HA and NA
    glycoproteins.

18
Distinguishing Characteristics
  • Influenza A viruses naturally infect humans,
    several other mammalian species and a wide
    variety of avian species whereas Influenza B and
    C are human pathogens.
  • The surface glycoproteins of influenza A virus
    exhibit much greater amino acid sequence
    variability than their counterparts in influenza
    B virus. Influenza C has a single multifunctional
    glycoprotein.
  • Influenza A and B viruses contain 8 RNA segments,
    whereas influenza C contains 7 segments

19
Virion Structure
  • Influenza A and B are morphologically
    indistinguishable but there are morphological
    features that distinguish influenza A and B
    viruses from influenza C virus.
  • The RNPs consist of four protein species and the
    RNA genome which occurs in eight separate
    segments containing 10 genes.
  • The segments are complexed with nucleoprotein to
    from a nucleocapsid with helical symmetry.
  • Genomic segments range from 890 to 2340 bases.

20
Replication
  • Unlike replication of other RNA viruses,
    replication of orthomyxovirus depends on the
    presence of active host cell DNA synthesis.
  • Replication in the nucleus is necessary because
    the virus lacks capping and methylating enzymes
    activities.
  • The virus scavenges cap sequences from the
    nascent mRNA generated in the nucleus and
    attaches it to its own mRNA.

21
Genome Organization
  • RNA segment 1 codes for PB2
  • RNA segment 2 for codes PB1
  • RNA segment 3 for codes PA
  • RNA segment 4 for codes HA
  • RNA segment 5 for codes NP
  • RNA segment 6 for codes NA
  • RNA segment 7 for codes M1 and M2
  • RNA segment 8 for codes NS1, and NS2

22
Virion Proteins
  • PB2, PB1 (Basic), PA (acidic)
  • NP
  • HA (16 subtypes)
  • HEF (Influenza C)
  • NA (9 subtypes)
  • M1 and M2
  • NS1 and NS2

23
Genetics
  • RNA segment Reassortment (Antigenic shifts)
  • RNA Mutations (antigenic drifts)
  • RNA Recombination
  • Nomenclature
  • - A/Swine / lowa/15/30/H1N1
  • - A/ Bangkok/1/79/H3N2

24
Paramyxoviridae
25
  • Enveloped viruses with a negative single stranded
    nonsegmented RNA genome.
  • They have special relationships with
    orthomyxoviruses and rhabdoviruses.
  • They encode and package their own RNA
    transcriptase.
  • They range in size from 150 350 nm

26
Classification
  • The Paramyxovirinae
  • Paramyxovirus Parainfluenza virus
    types
  • 1 and 3.
  • Rubulavirus mumps virus,

  • parainfluenza virus types
  • 2, 4a and
    4b.
  • Morbillivirus measles virus.
  • The pneumovirinae
  • Pneumovirus Respiratory Syncytial
  • Virus (RSV)

27
  • Nucleoprotein (NP)
  • Phosphoprotein (P Protein)
  • large (L) protein
  • The matrix (M) protein
  • Envelope Glycoproteins
  • - Attachment protein (HN,H, G)
  • - Fusion Protein (F)
  • Other proteins
  • - SH, C, V, W, I, D, NS1 and NS2.

28
Parainfluenza Virus
  • ssRNA virus
  • enveloped, pleomorphic morphology
  • 5 serotypes 1, 2, 3, 4a and 4b
  • No common group antigen
  • Closely related to Mumps virus

29
Parainfluenza Viruses
  • Important respiratory tract pathogens of infants
    and children causing 30-40 of such infections.
  • They are second only to RSV as a cause of serious
    respiratory tract disease in infants and children
    (HPIV 1-3).
  • Pleomorphic, 150-200 nm in diameter, enveloped
    with HN and F envelope glycproteins.

30
Respiratory Syncytial Virus (RSV)
  • RSV is the most important cause of viral lower
    respiratory tract disease in infants and children
    worlwide.
  • RSV infection is an important agent of disease in
    immunosuppressed adults and the elderly.
  • Ranges in diameter from 150-300 nm

31
  • Research on RSV has been impeded because
  • - It grows poorly in tissue culture and most
  • exprerimental animals
  • - it does not shut off host macromolecular
  • synthesis
  • - The virion is unstable.

32
  • RSV survives on surfaces for up to 6 hours and on
    gloves for less than 2 hours.
  • The virus loses viability with freeze-thaw
    cycles, in acidic conditions and with treatment
    by disinfectants.
  • It encodes a larger number of mRNAs than do the
    paramyxoviruses (10 compared with 6 or 7)
  • Additional genes are SH, M2, NS1, and NS2

33
  • Although six proteins appear to correspond (N, P,
    M, G/H/HN F and L) only F and L exhibit
    unambiguous sequence relatedness between the two
    subfamilies.
  • Variation in the G glycoprotein (RSV-A and B)
  • RSV utilizes ICAM-1 as its receptor.

34
Mumps Virus
  • to mump means to grimace or grin.
  • The virion is 120 200 nm in diameter
  • Contains in addition to the six major proteins V
    (viral) protein and S (soluble) protein.
  • One serotype.

35
  • MEASLES (RUBEOLA)
  • Measles is a relatively new disease of humans.
  • Probably it has evolved from an animal
    morbillivirus (rinderpest).
  • It is related to canine distemper virus.
  • Abu- Becr Al- Razi of 10th century is credited
    with distinguishing smallpox from measles.
  • He referred to measles as hasbah eruption in
    Arabic and regarded it as a modification of
    smallpox.

36
  • It is highly infectious and almost always
    produces clinical disease in those infected.
  • Virion is similar to other members of the
    paramyxoviridae but it lacks the neuraminidase.
  • Membrane cofactor protein (MCP) or CD46 is the
    receptor for the virus.
  • Measles virus is a stable monotypic virus with
    some degree of variability (strains).

37
Human Metapneumovirus
  • In 2001, van den Hoogen and colleagues reported
  • that they had isolated a paramyxovirus from
    28
  • young children in the Netherlands identified
    as a new
  • member of the metapneumovirus genus by
  • - Virological data
  • - Sequence homology
  • - Gene constellation
  • Previously, avian pneumovirus was the sole member
  • of this recently assigned genus, hence the
    provisional
  • name for the newly discovered virus human
  • metapneumovirus.

38
hMPV Features
  • Negative stranded RNA virus
  • Paramyxoviridae family
  • Related to avian pneumovirus and turkey
    rhinotracheitis virus
  • Causative agent of respiratory tract disease in
    humans
  • Most children are seropositive by the age of 5
    years
  • 2 genetic clusters of hMPV that may represent
    different serotypes

39
Rubella virus
  • Rubella virus is a member of the togaviridae but
    unlike most other togaviruses, rubella virus has
    no known invertebrate host, and the only known
    natural reservoir for rubella virus is man.
  • Rubella virus is a spherical, icosahedral,
    enveloped particle that measures 60-70 nm in
    diameter.
  • It has a ss RNA genome of about 10.000 nt that
    is encased by multiple copies of the capsid
    protein (C). Two glycoproteins, E1 and E2, are
    embedded in the envelope

40
Rhabdoviridae
41
  • A large number of member viruses that are
    serologically unrelated.
  • Rabies belongs to the genus lyssa virus (rabies
    in Greek means mad or frenzy).
  • It is bullet shaped, enveloped and has a diameter
    of 75X180 nm.

42
Rabies Virus
G, M, L, N, and NS Proteins
43
  • The genome is helical and is associated with N
    protein.
  • Virions bud from the endoplasmic reticulum
  • Replication of rhabdoviruses is followed by cell
    death except for rabies virus which is nonlytic
    causing no discernable damage

44
CORONAVIRUSES
45
HISTORY AND CLASSIFICATION
  • Avian Infectious Bronchitis (IBV)
  • (Schalk and Hawn, 1931).
  • Recovery of virus in the Laboratory (Beaudette
    and Hudson 1937).
  • Discovery of human coronaviruses
  • (Tyrrell and Bynoe, 1965).
  • Distinctive morphology.

46
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49
Genome Structure
50
Human Coronaviruses
  • Genus Coronavirus
  • Species HCoV-229E
  • HCoV-OC43
  • SARS- CoV
  • HCoV-NL63
  • HCoV-HKU1
  • HCoV-EMC
  • Responsible for about 10-20 of common colds
  • re-infection is common
  • infections year-round, most prevalent in fall and
    spring
  • incubation period about 2 to 5 days

51
VIRION STRUCTURE
  • There is considerable diversity in both the
    lengths and nucleotide sequences of the S1
    glycoproteins of different coronaviruses and even
    of different strains of a single coronavirus.
  • This diversity in S1 probably results from
    mutation and recombination between coronaviruses
    and strong positive selection in vivo.

52
Functions of Coronavirus Proteins
  •  Membrane (M) glycoprotein
  • May determine budding site on intracellular
    membranes
  • Essential for envelope formation
  • May interact with viral nucleocapsid
  • May induce alpha interferon

53
Functions of Coronavirus Proteins
  • Spike (S) glycoprotein
  • Binds to specific host cell receptor glycoprotein
  • May induce fusion of viral envelope with cell
  • membrane
  • Induces cell fusion
  • Binds immunoglobulin at Fc receptor site
  • Binds to 9-O-acetylated neuraminic acid
  • Induces neutralizing antibody
  • Elicits cell-mediated immunity

54
Coronavirus Stability
  • Stable
  • In body fluids (e.g. urine and faeces) for up to
    4 days.
  • For ? 21 days at cold temperature (4 and -80ºC).
  • At a pH of 6.
  • Inactivated rapidly
  • At a mild alkaline pH.
  • By disinfectants
  • By heating to 56 ºC

55
REPLICATION OF CORONAVIRUSES
  • Primary translation.
  • Transcription of viral RNA.
  • Replication of viral RNA.
  • Processing and intracellular transport of viral
    proteins (S glycoprotein).
  • Assembly and release of virions.

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57
Coronavirus Genetics
  • Variation is due to Mutation and
  • Recombination
  • Mutation
  • High frequency (several point mutations during
    each round of replication).
  • Analysis has shown extensive sequence
    variability in S and N genes especially due to
    deletion mutations.

58
  • The most striking example of the biological
    importance of deletion mutations is the emergence
    of porcine respiratory coronavirus (PRCV) from
    transmissible gastroenteritis virus (TGEV) which
    causes epizootic enteric infection of pigs.
  • In the early 1980s, PRCV emerged in Europe as a
    new virus that causes widespread, devastating
    epizootics of respiratory disease in pigs.

59
  • Recombination
  • High frequency (up to 25). Mechanism is by
    discontinuous transcription and polymerase
    jumping (Copy-choice). Example is acquisition of
    HE glycoprotein from influenza C
  • The capacity of coronaviruses both to recombine
    and to mutate suggests that diversity will also
    be a feature of human coronaviruses and that
    changes in pathogenicity may occur over time
    (Kenneth McIntosh, 1996).

60
Propagation and Assay in Cell Culture
  • In tissue culture, coronaviruses have a latent
    period of about 5 to 7 hours.
  • Infectivity of virions is fairly stable at pH
    6.0, but rapidly inactivated at mildly alkaline
    pH.
  • Coronaviruses can cause either cytocidal or
    persistent infections of cells in vitro and in
    vivo, depending on the virus strain and the host
    cell.

61
  • None of the human coronaviruses, except HCoV-EMC,
    grows well in cell culture without extensive
    adaptation by passage.
  • They have been propagated in human embryonic
    tracheal organ culture, in primary or secondary
    human embryonic kidney cell lines, in many
    diploid human fibroblast cell lines, and in few
    heteroploid lines.
  • The most sensitive cell line for isolation of
    virus from clinical specimens appears to be the
    diploid intestinal cell line MA 177.
  • The highest titers of both 229E and OC43 are
    obtained by growth in human rhabdomyosarcoma
    cells.

62
Reoviridae
63
  • Respiratory Enteric Orphan viruses (Albert Sabin,
    1959)
  • Non enveloped with double-layered protein capsid,
    containing 10-12 segments of the double-stranded
    RNA genomes (double double).
  • Stable over wide PH and temperature ranges and in
    air-borne aerosols.

64
  • Human Pathogens
  • - Orthoreoviruses
  • - Rotaviruses
  • - Orbiviruses
  • - Coltiviruses

65
  • Rotaviruses cause human infantile
    gastroenteritis.
  • They account for approximately 50 of all cases
    of diarrhea in children requiring hospitalization
    because of dehydration.
  • In underdeveloped countries, rotaviruses may be
    responsible for causing as many as 1 million
    deaths each year from uncontrolled viral diarrhea.

66
Rotavirus Particle
67
  • Proteolytic cleavage of the outer capsid
    activates the virus and produces an
    intermediate/infectious subviral particle (ISVP).
  • Rotaviruses resemble enveloped viruses and they
    acquire an envelope and loose it during
    replication.
  • Reassortment of gene segments can occur and thus
    create hybrid viruses.

68
  • Gene segment Protein location
    Function
  • VP1
    (inner capsid)
    Polymerase
  • VP2
    (inner capsid)
    Transcriptase
  • VP3
    (inner capsid) mRNA
    capping
  • VP4
    (outer capsid

  • spike at vertice)
    Activation by protease to VP5 and


  • VP8 in ISVP, HA and VAP
  • NS53

    RNA binding
  • VP6
    (inner capsid)

  • Groups (A-E) and

  • Subgroups (I,II) Major
    structural protein binds to NS28 at


  • ER and promote outer capsid assembly
  • NS34
  • NS35
  • VP7
    (Outer capsid)

  • Serotypes ( 1-7)
    Type-specific antigen major, outer capsid


  • component

69
  • Rotaviruses are found in many different mammals
    and birds.
  • Rotavirus is stable at room temperature and to
    treatment with detergents, pH extremes of 3.5 to
    10 or even repeated freezing and thawing.
  • Infectivity is enhanced by proteolytic enzymes
    such as trypsin.

70
  • Human and animal rotaviruses are divided into
  • 7 serotypes on the basis of antigenicity of VP7
    and VP4
  • 5 groups on the basis of electrophoretic mobility
    of VP6 DNA
  • 2 subgroups on the basis of antigenicity of the
    inner capsid protein VP6.

71
Caliciviruses and Related Agents
  • Caliciviruses are 27-38 nm, non enveloped,
    icosahedral viruses with a () ss RNA genome
    (7500 bases).

72
  • Norovirus (previously called Norwalk agent) was
    discovered by EM in stool from adults during and
    epidemic of an acute gastroenteritis in 1968 in
    Norwalk, Ohio.

73
Noroviruses
  • Noroviruses (genus Norovirus, family
    Caliciviridae) are a group of related,
    single-stranded RNA, nonenveloped viruses that
    cause acute gastroenteritis in humans.
  • Norovirus was recently approved as the official
    genus name for the group of viruses provisionally
    described as Norwalk-like viruses (NLV).
  • Currently, there are at least five norovirus
    genogroups (GI, GII, GIII, GIV and GV), which in
    turn are divided into at least 31 genetic
    clusters.

74
  • Astroviruses have a five or six-pointed star
    shape of 28-30 nm in diameter with an icosahedral
    symmetry.

75
The Retroviridae
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  • Genome
  • - 2 identical molecules of ss RNA
  • - Gene order is invariably gag- pro/ pol - env.
  • - other genes are present in some viruses.
  • Mode of Replication
  • Classification
  • - 6 genera, and human pathogens belong to 2
    genera HTLV-BLV and Lentiviruses.
  • HTLV-1 (1981), HTLV-2 (1982)

78
Human Immunodeficiency Viruses (HIV)
  • Pneumocystis carinii and Kaposis sarcoma among
    initial 4 H club of AIDS.
  • Isolation of LAV by Montagnier In April 1983.
  • A year later, Galo at NIH, isolated HTLV-3.
  • HTLV-3 and LAV showed 98-99 identity.
  • LAV-2 and HTLV-4 were isolated In 1986

79
  • In 1986, the ICVT renamed the viruses HIV-1 and
    HIV-2.
  • HIV-1 and HIV-2 are lentiviruses (lenti slow).
  • HIV-1 and HIV-2 share about 40 of their genome
    sequence.
  • Similarity is remarkable between HIV-1 and
    SIVcmp, and between HIV-2 and SIVsmm.

80
  • Greatest sequence variation exists in the env.
    gene
  • The genome is composed of 2 identical copies of
    9.749 kb ss RNA of positive polarity.
  • A tRNA molecule is positioned near the 5- end of
    each strand, with 10-50 copies of reverse
    transcriptase. The tRNA is used as a primer for
    DNA synthesis.
  • At each end, there are LTR sequences which
    contain promoters, enhancers, and other gene
    sequences for binding different cellular
    transcriptional factors.
  • Although of positive polarity, HIV genome is not
    infectious

81
  • cleavage
  • P55 ? P17(MA), P24(CA), P9(NC), and
  • P7 (?).
  • P100 ? P10 (pro) P51/P66 (RT/ RNAse H),
  • P32(1N).
  • P160 ? gP120 (SU) and gP 41(TM).

82
HIV Genome
83
Other Genes of HIV
  • tat Positive regulator of transcription
  • rev Regulator of viral expression
  • vif Affects viral infectivity
  • vpr Positive regulator of transcription,
  • augments virion production.
  • vpu Down regulates CD4
  • nef So-called negative-regulation factor. It
  • augments viral replication and
    down
  • regulates CD4

84
Antigenic Variation
  • The reverse transcriptase is very error prone and
    lacks proof reading which contribute to HIV
    diversity.
  • The immune response of the host is unable to
    completely curtail viral replication.
  • Virus gene products may be relatively invisible
    to the immune response and the virus may be able
    to mask or change its antigenic specificity.

85
  • The envelope gene displays frequent mutations.
  • HIV envelope glycoproteins have two unusual
    features.
  • - They are extensively glycosylated
  • - They contain hypervariable regions that permit
    the virus to present new antigenic configurations
    to the host.

86
  • HIV can constantly vary its surface antigenic
    composition which may allow it to avoid
    inactivation.
  • Such a mechanism hinders the development of an
    effective vaccine containing the surface
    glycoproteins.
  • Major sequence differences exist between the two
    HIV types antibodies against the surface
    glycoprotein of HIV-1 only partially cross react
    with HV-2.

87
Arboviruses
88
  • Epidemiologic classification, but taxonomically
    diverse.
  • More than 400, all of which are RNA viruses and
    about 100 of them infect humans.
  • They establish life-long infections of arthropods
    and humans are accidentally infected.
  • They have strong dependence on climatic
    conditions.

89
Families
  • Togaviridae VEE, EEE, WEE
  • Falviviridae Dengue, YF, West Nile fever,
  • JE, St. Louis
    Encephalitis, Kyasanur
  • Forest Disease, Omsk
    H.F.
  • Bunyaviridae CE, Rift valley fever, Crimean-
    Congo
  • virus, Sand fly
    fever, Hantaan virus
  • Arenaviridae Junin, Machupo, Sabia, Guanarito,
  • Lassa
  • Filoviridae Marburg and Ebola viruses

90
The Bunyviridae
  • A supergroup of at least 300 viruses.
  • Spherical, enveloped, 90-120 nm, with 3 segments
    of ambisense ssRNA.
  • Four genera Bunyavirus, Phlebovirus, Niarovirus
    and Hantavirus.
  • All, except Hantaviruses, are arthropod borne.
    Hantaviruses are rodent borne.

91
Arenaviruses
  • Pleomorphic, enveloped viruses of 120 nm in
    diameter
  • 2 circles of ambisense ss RNA
  • A sandy appearance (arenosa sandy in Greek)
    because of the ribosomes in the virion.

92
Filoviruses
  • Filamentous, enveloped, with 80nm in diameter.
  • - ss RNA of helical symmetry
  • They vary in length from 800 to 14.000nm.

93
Hepatitis Viruses
94
  • A large number of viruses can cause hepatitis
    (EBV, CMV, VZV, HSV, YF, Lassa virus etc).
  • There are other viruses, however, that only cause
    hepatitis.
  • At least six viruses, A through E and a newly
    discovered virus GB, are considered hepatitis
    viruses.
  • Hepatitis viruses differ greatly in their
    taxonomy, structure, mode of replication and mode
    of transmission as well as in the course of the
    disease they cause.

95
Hepatitis A Virus
  • It is the cause of infectious hepatitis, a term
    that was coined in 1912 to describe the epidemic
    form of the disease.
  • The virus was first isolated in 1973 by Feinstone
    et al using IEM.
  • Previously classified as enterovirus 72, HAV has
    been put in a separate genus hepatovirus

96
Differences Between HAV and Enteroviruses
  • HAV nucleotide and amino acid sequences are
    dissimilar to enteroviruses.
  • HAV is difficult to grow in cell culture and it
    usually replicates very slowly causing no CPE
    (nonlytic).
  • HAV is stable at a PH of 1
  • HAV has only one serotype and one neutralization
    site is dominant.
  • Enterovirus -specific monoclonal antibody does
    not react with HAV.

97
Virus Structure
  • HAV is a naked 27-32 nm icosahedral virus with a
    ssRNA genome of positive polarity.
  • Its genome is 7.5 kb in length, polyadenylated at
    the 3- end and carries a protein (VPg) at the 5-
    end.

98
Hepatitis A Virus
99
Hepatitis B Virus
100
Hepatitis B Virus
  • Hepatitis B virus is unusual among animal viruses
    in that
  • Infected cells produce multiple types of
    virus-related particles.
  • - 42 nm double-shelled particles (Dane
    particles)
  • - 20 nm spheres, usually present in 104-106 fold
    excess over Dane particles.
  • - Smaller quantities of filaments of 20 nm
    diameter and variable length.

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102
Hepatitis B Virus
103
  • The genome is partially double stranded.
  • It replicates utilizing an RNA intermediate and
    has a reverse transcriptase.
  • It is unusually stable for an enveloped virus.
  • The Dane particle is the only infectious form.
  • Envelope HBsAg.
  • Core HBcAg.

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105
  • With less than 3200 nucleotides, HBV has the
    smallest genome of any human virus.
  • HBV genome is relaxed circular partially duplex
    DNA species, whose circularity is maintained by
    5- cohesive ends.
  • The genome has a coding organization that is
    highly compact and over half of the sequence is
    translated in more than one frame.

106
  • HBV uses its genome economically by encoding
    different proteins within the same region of DNA
    in different reading frames.
  • About half of the genomes nucleotides are used
    to code simultaneously for different proteins,
    and all code for at least one protein.
  • The regulatory signals overlap with coding
    sequences and are not separate regions.

107
  • Four ORFs are present in the DNA.
  • - ORF P encodes the viral polymerase and the
    terminal
  • protein found on minus strand
    DNA.
  • - ORF C encodes the core protein (C antigen)
  • - ORF S/pre-S encodes the HBsAg.
  • - ORF X encodes a protein that enhances the
  • expression of heterologous and
    homologous
  • genes.

108
  • The reading frame for HBsAg was shown to have two
    in-frame initiation codons which result in three
    products L, M, and S.
  • The bulk of HBsAg is the S protein, M protein
    accounts for 5-15 of it and L for 1-2.
  • The S (gp 27, 24-27 kD) glycoprotein is
    completely contained in the M (gp36 33-36 kD)
    glycoprotein which is contained in the L (gp42
    39-42 kD) glycoprotein.

109
  • The coding organization of the core protein is
    similar, two in-frame AUGs were found in the core
    ORF.
  • Initiation at the upstream AUG gives rise to a
    C-related protein that is secreted from infected
    cells into circulation (HBeAg).
  • Three major mRNAs are produced
  • 1) 2100 b mRNA HBsAg (S, M)
  • 2) 2400 b mRNA HBsAg (L)
  • 3) 3500b mRNA HBcAg, HBeAg, polymerase and a
    protein
  • primer for DNA
    replication and it acts as a
  • template for
    genome replication

110
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111
  • In addition to a minor mRNA (700 b) which codes
    for X protein (a transactivator of transcription
    and a protein kinase).
  • HBsAg contains the group specific antigenic
    determinant termed a and type specific
    determinants termed d or y and w or r
  • Type specific determinants behave like mutual
    alleles. Combinations result in four possible
    antigenic subtypes (adw, adr, ayw, ayr).

112
HBV Variation
  • Eight genetic groups (A-H Genotypes) and a
    possible 9th type (I?) and Within genotypes 24
    subtypes have been described which differ by 4-8
    of the genome.
  • HBV Antibody Escape Mutants
  • Substitution of arginine for glycine at aa 145.
  • Immunity to vaccine does not neutralize mutant.
  • Failure to detect HBsAg in donated blood
  • HBV Precore Mutants
  • Detected in patients with severe chronic liver
    disease and who may have failed to respond to
    interferon therapy (increased pathogenicity)
  • Polymerase Variants ( drug resistance)

113
Replication
114
  • Virus particles acquire an envelope in the
    endoplasmic reticulum or proximal Golgi
  • The surface antigen is glycosylated in the Golgi
    apparatus.
  • Virions are then secreted via the constitutive
    pathway of vesicular transport.

115
Hepatitis D virus (Delta virus)
  • HDV is not a true virus but a defective virus or
    a natural satellite of HBV
  • Delta antigen is present in two forms, small
    (short) with 24 kd and large (long) with 27kd.
  • The short form which is more abundant is required
    for RNA replication whereas the long form
    suppresses viral RNA replication and is required
    for packaging of the HDV genome by HBsAg.

116
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117
  • HDV replicates only in HBV-infected cells and
    direct pathologic changes are limited to the
    liver, the only organ in which HDV has been shown
    to replicate.
  • HDV itself seems to be cytopathic and HDV antigen
    (delta) may be directly cytotoxic.

118
Hepatitis C virus
119
  • Was discovered in 1989 (post transfusion
    hepatitis).
  • Flaviviridae, genus Hepacivirus (HCV and GBV-C).
  • Enveloped, 55-65 nm.
  • Six genotypes with several subtypes for each
    genotype
  • ss RNA genome
  • - 9.4 kb
  • - over 98 contains protein coding sequence.
  • - a single large ORF

120

Hepatitis C Virus
capsid
Envelope protein
protease/ helicase
RNA polymerase
RNA dependent
c22
c 33
c-100
5
3
core
E1
E2
NS2
NS3
NS4
NS5
hypervariable region
121
  • The genome codes for nine proteins 3 structural
    and 6 nonstructural.
  • The structural proteins are core protein (P22),
    E1(gp76), and E2 (gp35) which are envelope
    glycoproteins.
  • The nonstructural proteins are NS2, NS3, NS4A,
    NS4B, NS5A, and NS5B.
  • HCV has circumvented the cap requirement by
    evolving an Internal Ribosome Entry Site (IRES)
    at its 5- end.

122
Hepatitis E Virus

123
  • Recognized as a distinct disease in 1980.
  • Virion is 32-34 nm in diameter, nonenveloped,
    with an icosahedral symmetry.
  • Although it was originally classified in
    the Caliciviridae family, the virus has since
    been classified into the genus Hepevirus, but was
    not assigned to a viral family.
  • The genome is approximately 7200 bases in length,
    is a polyadenylated single-strand RNA molecule
    that contains three discontinuous and partially
    overlapping ORFs

124
  • ORF1 encode methyltransferase, protease,
    helicase, and replicase
  • ORF2 encode the capsid protein 
  • ORF3 encodes a protein of undefined function.
  •  An in vitro culture system is not yet available.
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