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ARBOROVIRUSES
Mohammed El-Khateeb
2nd April 2015
Overview
 Etiology
 Epidemiology and history
 Pathogenesis and Pathology
 Clinical Manifestation
 Diagnosis
 Treatment
 Prevention and Control
2
Arthropod-Borne Viruses
arbov
Arthropod-borne viruses (
iruses) are More Than
600 Different viruses that can be transmitted to man by
arthropod vectors.
The WHO definition is as follows:
“Viruses maintained in nature principally, or to an
important extent, through biological transmission
between susceptible vertebrate hosts by
hematophagus arthropods or through transovarial
and possibly venereal transmission in
arthropods.”
Arthropod-Borne Viruses
Arboviruses belong to three families
1. Togaviruses e.g. EEE, WEE, and VEE
2. Bunyaviruses e.g. Sand fly Fever, Rift
Valley Fever, Crimean- Congo
Hemorrhagic Fever
3. Flaviviruses e.g. Yellow Fever, dengue,
Japanese Encephalitis
Arthropod-Borne Viruses
• Togaviridae
• Flaviviridae
a) Sindbis
b) Semliki Forest
c) Venezuelan equine
encephalitis
d) Eastern equine
encephalitis
e) Western equine
encephalitis
f) Chikungunya
a) Dengue
b) Yellow fever
c) Japanese encephalitis
d) West Nile encephalitis
e) St. Louis encephalitis
f) Russian spring-summer
encephalitis
g) Powassan encephalitis
Negatively Stained Virions
of Semliki Forest Virus
Togaviridae, genus Alphavirus
Arthropod-Borne Viruses
FAMILY
ENVELOPE
SYMMETRY
GENOME
Icosahedral
40-75 nm
40-45 nm
ssRNA (+ve)
Yes, Heat,
detergent
labile
Helical
ssRNA (-ve)
segmented
no
Icosahedral
dsRNA,
segmented
Yes. Heat,
detergent
labile
VIRAL STRUCTURE
It has three proteins
• Envelope protein
• Core protein
• Membrane protein
General Antigenic Properties of
Flaviviral Proteins - 1
E-glycoprotein
 Most important flaviviral antigen
 Very immunogenic
 Most serological assays detect reactivity
with this protein
Capsid
 Elicits primarily group-reactive antibody
M-protein
 Very small (75 a.a.)
 Embedded in the virion envelope
membrane and not highly immunogenic
Replication
10
Arthropod Vectors
Mosquitoes
Japanese encephalitis, dengue, yellow fever, St.
Louis encephalitis, EEE, WEE, VEE etc.
Ticks
Crimean-Congo haemorrhagic fever, various tickborne encephalitides etc.
Sandflies
Sicilian sandfly fever, Rift valley fever.
Examples of Arthropod Vectors
Aedes Aegyti
Culex Mosquito
Assorted Ticks
Phlebotmine Sandfly
Animal Reservoirs
In many cases, the actual reservoir is not known.
The following animals are implicated as reservoirs
Birds
Japanese encephalitis, St Louis
encephalitis, EEE, WEE
Pigs
Japanese encephalitis
Monkeys Yellow Fever
Rodents VEE, Russian Spring-Summer
encephalitis
Transmission
• The most common route of infection is
•
bite of infectious mosquito
Other transmission modes where
revealed in 2002 such as
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
Blood Transfusion
Organ Transplantation
Intrauterine
Percutaneous exposure (occ. exposure)
Breastmilk (probable)
Transmission Cycles
 Man - arthropod -man
 e.g. dengue, urban yellow fever.
 Reservoir may be in either man or arthropod vector.
 In the latter transovarial transmission may take place.
 Animal - arthropod vector - man
 e.g. Japanese encephalitis, EEE, WEE, jungle yellow fever.
 The reservoir is in an animal.
 The virus is maintained in nature in a transmission cycle
involving the arthropod vector and animal. Man becomes
infected incidentally.
 Both cycles may be seen with some arboviruses such as
yellow fever.
Man-Arthropod-Man Cycle
Man - arthropod -man
e.g. dengue, urban yellow fever.
Reservoir may be in either man or arthropod vector.
In the latter transovarial transmission may take place.
Animal-Arthropod-Man Cycle
Animal - arthropod vector - man
e.g. Japanese encephalitis, EEE, WEE, jungle yellow fever.
The reservoir is in an animal.
The virus is maintained in nature in a transmission cycle involving
the arthropod vector and animal. Man becomes infected incidentally.
Epidemiologic Feature
 The major outbreaks coincided with the
heavy rainfall or floods.
 Seasonal: more common in summer, July
to October
 Infection provides life long immunity.
 Worldwide distribution
 More than 530 species, 150 pathogen to
man
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Pathogenesis
The nature of flavivirus disease is
determined primarily by
• The specific tropisms of the individual
virus type
• The concentration of infecting virus
• Individual host response to the infection
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Disease Syndromes of the Alphaviruses and Flaviviruses
Pathogenesis
Virus
Mononuclear Phagocyte
Blood Circulation
Viremia
Adequate
Immunological
Response
Subclinical or mild
Systemic disease
Weak
Immunological
Response
Invades the CNS
induce mortality
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Four stages
•
•
•
•
A Prodromal Stage
An Acute encephalitic Stage
The Convalescence Stage
A Sequela Stage
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Diseases Caused
• Fever and rash - this is usually a non-specific illness
resembling a number of other viral illnesses such as
influenza, rubella, and enterovirus infections. The
patients may go on to develop encephalitis or
haemorrhagic fever.
• Encephalitis - e.g. EEE, WEE, St Louis encephalitis,
Japanese encephalitis.
• Haemorrhagic fever - e.g. yellow fever, dengue,
Crimean-Congo haemorrhagic fever.
Pathogenesis and immunity
 Besides viral receptor, virus may attach to
Fc receptor (macrophages and monocytes)
via Ab to result in an increase of virus
infection.
 Antibody is produced to block infection.
However, non-neutralizing Ab may have
antibody dependent enhancement (ADE)
effect to enhance virus replication by
hundred folds.
Arthropod-Borne Viruses
a) Dengue
b) Yellow fever
c) Chikungunya
Fever/Rash/Arthritis
1. Triad of fever/rash/arthritis is characteristic of
Chikungunya, o’nyong-nyong, Ross River, Mayaro,
and Sindbis viruses
2. Symptoms generally appear after 2-3 days incubation
a) fever, chills, myalgia
b) polyarthralgia mainly affecting small joints
c) maculopapular rash
3. Arthritis generally resolves in a few weeks, but may
persist for months, or years in some cases.
Arthropod-Borne Viruses
Encephalitis
•
•
•
•
•
•
•
•
West Nile encephalitis
Japanese encephalitis
St. Louis encephalitis
Russian spring-summer encephalitis
Powassan encephalitis
Venezuelan equine encephalitis
Eastern equine encephalitis
Western equine encephalitis
Transmission of six encephalitis arboviruses
Humans
can be infected
via mosquito bites.
Small mammals
are hosts for VEE and
California viruses only.
Mosquitoes
are vectors.
Encephalitis arboviruses
can overwinter inside
mosquito eggs.
Wild birds
Horses,
and rarely other
domestic mammals
are hosts for equine viruses.
Domestic fowls
Birds are hosts for
all six encephalitis
arboviruses.
West Nile Virus
• Flavivirus
• Primary host – wild birds
• Principal arthropod vector – mosquitoes
• Geographic distribution:
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Africa,
Middle East,
Western Asia,
Europe,
Australia,
North America,
Central America
Encephalitis
1. Small proportion of individuals infected (a few days
after the onset of fever) may develop drowsiness,
neck rigidity, progressing to confusion, paralysis,
convulsions and coma.
1. Case-fatality rates average 10 – 20 % (higher in
elderly).
3.
Survivors may be left with permanent neurologic
sequelae such as mental retardation, epilepsy,
paralysis, deafness, and blindness.
Diagnosis
 Materials of epidemiology
 Clinical
 Laboratory Tests
– Tentative diagnosis
 Antibody titer: HI, IF, CF, ELISA
 JE-specific IgM in serum or CSF
– Definitive diagnosis
 Virus isolation: Blood, CSF sample, brain
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Diagnosis
 Serology - usually used to make a diagnosis of
arbovirus infections. Antibody titer: HI, IF, CF, ELISA
• Culture - a number of cell lines may be used,
including mosquito cell lines. However, it is rarely
carried out since many of the pathogens are group 3
or 4 pathogens. (Blood, CSF sample, brain)
• Direct detection tests - e.g detection of antigen and
nucleic acids are available but again there are safety
issues.
Prevention
• Surveillance - of disease and vector populations
• Control of vector - pesticides, elimination of
breeding grounds
• Personal protection - screening of houses, bed
nets, insect repellants
• Vaccination - available for a number of arboviral
infections e.g. Yellow fever, Japanese encephalitis,
Russian tick-borne encephalitis
Treatment/Vaccines/Control measures
A. Encephalitis
1. Vaccines exist for a number of these viruses, but
are used mainly for horses, at risk lab workers, and
some fowl known to be intermediate hosts
2. Control of mosquitoes is major countermeasure.
B. Yellow Fever
1. Live attenuated virus vaccine. Used when going to
endemic areas
Epidemiological Triangle
The Host
Interaction
The Virus
The Vector
Prevention
Vector (Mosquito) control
– Eliminate mosquito breeding areas: Chemical
larvicides, Biolarvicides, Environmental
management
– Adult and larval control: Anti-larval treatment
Vaccination
Personal protective measures
– Avoid prime mosquito hours: from dusk to dawn
– Indoor spray and fogging: Use of Insecticide
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Repellent Guidance
• Skin
 DEET still “gold standard”
• Both new additions good for shorter term protection
 Picaridin
• Roughly equivalent to DEET at same concentration
• Only a 7% product currently sold in US
 Oil of lemon eucalyptus
• Plant based
• 30% product similar to low concentration DEET
• Not for kids <3 years old
• Clothing
 Permethrin
Arboviruses
Structure
Positive sense ssRNA genome, icosahedral nucleocapsid, enveloped
•
Pathogenesis
• Transmitted by bite of insect from host species; sylvan and urban
cycles
• Replication in cytoplasm; budding
• Viremia to target tissue
• Influenza-like initial symptoms; different viruses cause encephalitis,
hemorrhagic fever, hepatitis, rash, arthritis
Diagnosis
• Serology and nucleic acid
Treatment/prevention
• No human vaccines except for Yellow Fever live attenuated vaccine,
control of insect population
The nomenclature of arboviruses are mostly based on endemic areas and
symptoms induced by viruses, including fever, encephalitis and hemorrhagic
Disease
Vector
Host
Distribution
Disease
fever
Alphaviruses
Sindbis*
Aedes and other
mosquitoes
Aedes and other
mosquitoes
Aedes, Culex
Birds
Africa, Australia, India
Subclinical
Birds
East and West Africa
Subclinical
Rodents, horses
Aedes, Culiseta
Birds
Culex, Culiseta
Birds
Aedes
Humans, monkeys
North, South, and
Central America
North and South
America, Caribbean
North and South
America
Africa, Asia
Mild systemic; severe
encephalitis
Mild systemic;
encephalitis
Mild systemic;
encephalitis
Fever, arthralgia,
arthritis
Dengue*
Aedes
Humans, monkeys
Worldwide, especially
tropics
Mild systemic; break-bone
fever, dengue
hemorrhagic fever, and
dengue shock syndrome
Yellow fever*
Aedes
Humans, monkeys
Africa, South America
Japanese encephalitis
Culex
Pigs, birds
Asia
Hepatitis, hemorrhagic
fever
Encephalitis
West Nile encephalitis
Culex
Birds
St. Louis encephalitis
Culex
Birds
Africa, Europe, central
Asia, North America
North America
Fever, encephalitis,
T2
hepatitis
Encephalitis
Russian spring-summer
encephalitis
Powassan encephalitis
lxodes and
Dermacentor ticks
lxodes ticks
Birds
Russia
Encephalitis
Small mammals
North America
Encephalitis
Semliki Forest*
Venezuelan equine
encephalitis
Eastern equine
encephalitis
Western equine
encephalitis
Chikungunya
Flaviviruses
RETROVIRUSES
RetroViruses
• RNA Viruses
• DNA From RNA by Reverse Transcriptase
• Insertion of new DNA into cellular DNA
• Hijacks the cell machinery to make VIRUS
• The virus only grows on T4 cells that are proliferating
in response to an immune stimulus
• Difficult to grow in culture
• Robert Gallo : HTLV-3
• Luc Montagnier: LAV
• Human Immunodeficiency Virus (HIV)
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Introduction to Retroviruses
I. Overview of retroviruses
A. History
B. Shared characteristics
C. Classification
II. Function of different regions of the retroviral genome
A. Cis acting elements
B. Gag proteins
C. Pol proteins
D. Env proteins
III. Details of life cycle:
A. Early stage
B. Late stage
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General Introduction to Retroviruses
Retroviruses
 Ubiquitous; found in all vertebrates
 Large, diverse family
 Includes HIV, FIV and FeLV
Definition and classification of retroviruses
 Common features- structure, composition and replication
 Distinctive life cycle: RNA-DNA-RNA
 Nucleic acid is RNA in virus, and DNA in infected cell
Transmission may be either:
 Horizontal - by infectious virus (exogenous virus) or vertical- by proviruses
integrated in germ cells (endogenous virus)
 Can transmit either as free viral particle or (for some retroviruses) through cellcell contact
44
A Little Retrovirus History
1960s: Howard Temin: suggested DNA “provirus” was
– part ofreplication cycle: RNA DNA  RNA  Protein
Won Nobel prize (with Baltimore) in 1970 after
they independently discovered RT activity in
infected cells
1980: Human T-cell leukemia virus discovered,
the first pathogenic human retrovirus.
1982: Human immunodeficiency virus discovered.
1990: First gene therapy trial involving the use of retroviral-based vectors
in patient with a deficiency in adenosine deaminase (ADA).
2006: Xenotropic murine leukemia-related virus discovered.
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Retroviruses
• Strange Viruses ?
At time-“central dogma of molecular biology”:DNARNAProtein
So.. RNA couldn’t be template for DNA
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
Unique replication cycle based on reverse transcription. Flow of
information from RNA to DNA. (1971 Nobel Prize Temin / Baltimore)
Retroviruses have been isolated from numerous species including
chickens (RSV), mice (MLV), monkeys (SIV), and humans (HIV, HTLV)

“Simple Retroviruses” encode only the genes gag, pol, and env
(RSV)

“Complex Retroviruses” encode in addition regulatory genes (HIV)
•
Retroviruses are single-stranded RNA viruses that replicate through a double-stranded DNA intermediate.
• They are association with the development of
tumors in their host organisms.
•
Study of these viruses eventually led to the
discovery and development of the oncogene
theory of tumorgenesis
• Some of the viruses actually contained
oncogenes within their genomes, while others
interacted with oncogenes in either a direct or
indirect way to contribute to tumor formation.
• Historically, because of their pattern of pathogenicity,
these viruses were grouped into three subfamilies:
1. The acutely oncogenic retroviruses, or
oncoretroviruses (such as those described above)
2. The lentiviruses (associated with “slow” diseases or
those with long latent periods)
3. The spumaviruses (“foamy” viruses, named because
of the pathogenic changes observed in infected
cells).
THE OLD NOMENCLATURE
Members:
1. Oncogenic viruses (Oncoviruses) (endogenous)
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Avian oncoviruses: RSV, AMV, AEV, RAVs [ RAV- 0 ; RAV-1 ].
Murine oncoviruses: e.g., MoMLV, A-MuLV.
Mammalian oncoviruses: e.g., FeLV, HaMSV, SSV .
Mouse Mammary Tumor Virus (MMTV); [ the only B- type particle ].
Mason-Pfizer Monkey Virus (MPMV); [ one of the few D-type particles ].
Human T-Cell Lymphotropic Virus ( HTLV-1 & 2 ).
2. Lentiviruses - HIV-1, HIV-2, SIV, FIV, EIAV, CAEV.
3. Spumaviruses – HFVs, SFV.
2 and 3 non endogenous
Retrovirus Classification
Genus
Example
Genome
Alpharetrovirus
Avian leukemia virus
Simple
Betaretrovirus
Mouse mammary tumor virus
Simple
Gammaretrovirus
Murine leukemia virus
Simple
Feline leukemia virus
Xenotropic murine leukemia-related virus
Deltaretrovirus
Human T-cell leukemia virus
Complex
Epsilonretrovirus
Wall-eyed sarcoma virus
Complex
Lentivirus
HIV, SIV, FIV
Complex
Spumavirus
Human foamy virus
Complex
Metavirus
Yeast TY-3
Errantvirus
Drosophila melanogaster Gypsy
Retrovirus Structural Overview
Enveloped virus with lipid bilayer and viral spike glycoproteins.
Have outer matrix protein and inner core capsid containing viral genome.
Genome: Two copies of single stranded positive-stranded RNA (8-10kb).
All retroviruses contain gag, pol and env genes.
Simple - only gag, pol, env
Complex - additional genes involved in replication.
Reverse transcriptase to generate DNA
Viral genes are integrated into host genome.
Progeny virus produced using host cell transcriptional and translational machinery.
51
Retroviruses
Transmission EM
matrix
Env
capsid
RNA
Scanning EM
3 D representation of HIV virion:
52
Retrovirus Genome (Diploid)
u
Retrovirus genome is +RNA
u
Ranges from 7-10 kb in size (1 copy)
u
Diploid: 2 copies/virion
u
Important in high recombination rate
From Flint et al. Principles of Virology (2000), 53
ASM Press
y ( Packaging Signal)
5’m7GpppG R U5
gag
pol
PPT
env
U3 R AAAA 3’
PBS- primer binding site
PPT- polypurine tract
R - repeat sequence
U3 - promoter/enhancer
PBS
U5 - reverse transcription/
integration.
CA
MA
CA
SU
NC
TM
PRO RT IN
MA-Matrix
CA- Capsid
NC- Nucleocapsid
PRO- Protease
RT- Reverse transcriptase
IN- Integrase
SU- surface envelope protein
TM- transmembrane envelope protein.
54
Gene Proteins
Retroviral Structural
genes
Function
gag = group specific antigen (internal structural proteins)
Matrix (MA),
Capsid (CA),
Nucleocapsid (NC)
binds envelope, organization
protects genome and enzymes
chaperones RNA, buds
pol = polymerase enzymes
Reverse transcriptase +
RNAase H (RT)
Protease (PR)
Integrase (IN)
RNA to DNA
degrades template RNA
maturation of precursors
provirus integration
env = envelope proteins
Surface glycoprotein (SU) receptor binding
Transmembrane protein (TM) virus-cell fusion
55
Viral life cycles
 Virulent – these viruses lyse (kill) their
host cell after infection.
 Temperate – these viruses can
replicate their genome along with the
host cell genome without killing the host
cell.
 These viruses are also capable of lyzing
the host cell
Distinct Steps in the Retroviral
Life Cycle
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Attachment, Fusion, and Entry
Reverse Transcription
Integration
Transcription
Translation
Assembly, Budding, and Maturation
Multiplication of a Retrovirus
Capsid
Reverse
transcriptase
DNA
Virus
Two identical + stands of RNA
1 Retrovirus penetrates host
cell.
Host
cell
DNA of one of the host
cell’s chromosomes
5 Mature
retrovirus leaves
host cell,
acquiring an
envelope as it
buds out.
Reverse
transcriptase
Viral RNA
Identical
strands of
RNA
2
Virion penetrates cell
and its DNA is
uncoated
4 Transcription of the provirus
Viral proteins
RNA
may also occur, producing
RNA for new retrovirus
genomes and RNA that codes
for the retrovirus capsid and
envelope proteins.
3 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.
Provirus
Retrovirus budding from a cell
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After Budding, Virus Goes from Immature to
Mature Form
Mature Form (after budding):
-Core becomes more dense
-Different retroviruses have different morphology in mature form
60
Retroviruses May Transduce Cellular Sequences
 Transforming retroviruses are generated by a recombination event in which a
cellular RNA sequence replaces part of the retroviral RNA.
 Transducing virus : carries part of the host genome in place of part
of its own sequence. The best known examples are retroviruses in
eukaryotes and DNA phages in E. coli.
 Replication-defective virus : cannot sustain an infective cycle by
itself, because some of the necessary genes are absent (replaced by
host DNA in a transducing virus) or mutated. It can, however, be
perpetuated in the company of a helper virus.
 Helper virus : provides missing viral functions to a defective virus,
enabling to complete the infective cycle during a mixed infection.
 Transformation (oncogenesis) : the ability to transform cultured
cells so that the usual regulation of growth is released to allow
unrestricted division.
Transmission of Pathogenic Retroviruses
Virus
Primary modes
of transmission
Range
Preventative
measures
ALV
feces, saliva, skin,
contact; mother to
offspring via egg
worldwide; common removal of infected
in untreated
dams; breeding
commercial flocks
resistant strains
lacking
endogenous
viruses that limit
infection
REV
feces, saliva, skin,
contact; mother to
offspring via egg
worldwide; common none taken due to
in commercial
low incidence of
flocks; contaminant disease
in Marek's disease
vaccine
MLV
mother to offspring
via milk
rare; Lake Casitas,
CA; La Puente, CA
none
MMTV
mother to offspring
via milk
most inbred strains
none