Transcript and HA 2 - Elsevier

Influenza
There are three types of influenza in humans: A,B, C
1. Flu A: Found in many animal species, in addition to humans
Closely related to Type B but not Type C
Main type responsible for human epidemics
Demonstrates the greatest antigenic variability (“antigenic drift”)
Reservoir in nature is waterfowl
2. Flus B and C: Found almost exclusively in humans
Flu C can also infect swine
Flu C is morphologically and antigenically distinct from A, B
3. Flu A strains designated by host from which isolated, where isolated, year of
isolation, and type of HA and NA. An isolate (strain) number may also be
included if there are multiple isolates.
Example: A/goose/Leipzig/137/79 (H7N2)
ORTHOMYXOVIRIDAE
GENUS/
MEMBERS
VIRUS NAME
ABBREV.
USUAL
HOST(S)
TRANSMISSION
DISEASE
WORLD
DISTRIBUTION
FLUAV
Humans , birds,
swine
Airborne
Respiratory
disease
Worldwide
FLUBV
Humans
Airborne
Respiratory
disease
Worldwide
FLUCV
Humans
Airborne
Respiratory
disease
Worldwide
Mammals
Tick-borne
INFLUENZAVIRUS A
InfluenzaA
INFLUENZAVIRUS B
Influenza B
INFLUENZAVIRUS C
Influenza C
“THOGOTO-LIKE VIRUSES”
Thogoto virus
THOV
Influenza
Virus
Structure of Influenza Virus
Influenza A virus is an enveloped particle that when spherical is
about 120 nm in diameter
Many particles are not spherical but filamentous in shape
There are two glycoproteins at the surface in surface “spikes”
HA (hemagglutinin) is present as homotrimers
NA (neuraminidase) is present as homotetramers
Protein M2 forms ion channels in the lipid bilayer
The matrix protein M1 lines the inner side of the lipid bilayer
The genome consists of 8 RNA segments present in helical
nucleocapsids
Influenza Nucleocapsids
NP is the major nucleocapsid protein.
It has a major structural role
It is also required for the switch from mRNA synthesis to genome
replication.
PA, PB1, and PB2 are minor components of the nucleocapsid and form the
RNA synthesis machinery.
The function of PA is unknown but may be involved in the switch
from mRNA synthesis to genome replication
PB1 is an endonuclease that process the mRNA primer; it also is a
polymerase that catalyzes nucleotide addition
PB2 recognizes the cap of host cell mRNA required for priming mRNA
synthesis
M is the matrix protein.
It is a peripheral membrane protein that underlies the viral membrane.
It interacts with the nucleocapsid and with the tails of HA, NA, and M2
Attachment & Entry
The HA spike is a homotrimer with a molecular weight of 110 kDa.
HA is synthesized as a 549 aa precursor called HA0 which is anchored in the membrane
near the C-terminus.
HA0 is cleaved into HA1 (328 aa) and HA2 (221 aa)
At the N-terminus of HA is a 16 aa hydrophobic signal peptide for insertion into the ER.
A single Arg separates HA1 from HA2 and cleavage is by a cellular trypsin-like proteinase
HA1 and HA2 remain covalently associated after cleavage by a disulfide bridge
The C terminus of HA2 contains a 26 aa uncharged membrane-spanning domain followed
by a 10 aa hydrophilic cytoplasmic domain
The HA polypeptide is glycosylated at specific asparagine residues
HA-mediated membrane fusion
The HA trimer is stabilized by a hydrophobic core formed between the three stalk
regions. Attachment sites for the cellular receptors are located near the top of each
large globular region, which also contains neutralization epitopes.
The exact glycoprotein(s) that serve as host cell surface receptors has not been
identified, but it is known to contain sialic acid.
After binding of HA to the cell surface receptor(s) the virus is internalized by
endocytosis.
The low pH of endosomes ( pH 5.0-6.0) results in an irreversible conformational change
in HA which results in the extrusion of the highly conserved hydrophobic amino
terminus of HA2 from its position in the native protein.
This region, termed the ‘fusion peptide’, promotes membrane fusion.
The mechanism by which the ‘fusion peptide’ promotes membrane fusion is not
completely understood.
The subsequent fusion of viral and endosomal membranes allows the release of the viral
genome into the cellular cytoplasm
Activation of the HA Spike
HA0 precursor
Cleaved spike
After acid treatment
and proteolysis
Activation of Fusion Activity of Flu HA0 by Cleavage
View of One Monomeric Unit in the Spike
Structures of the Native and Fusion Active Conformations of the Influenza
Hemagglutinin
A.
S-S
S
TM
N
16aa
C
HA 1 (328aa)
B.
C.
HA 2 (221aa)
B’.
40
C’.
40
A
B
153
76
76
G
76
153
B
129
C
1
F
S
S
E
C
105
105
A
105
105
1(N)
D
38
40
153
129
175
1
E
F
S
S
153
G
H
1(N)
Change illustrated for one monomeric unit of the trimeric spike
D
Model for Fusion
Neuraminidase
NA spike consists of a tetramer. NA is a type 2 glycoprotein
with the N terminus inside and the C terminus outside.
NA removes sialic acid from oligosaccharides on cell-surface
proteins and glycolipids, thus destroying receptors for
the virus.
Also removes sialic acid from HA so that progeny influenza
virions cannot aggregate.
Separates virus particles from inhibitory
mucopolysaccharides in the respiratory tract allowing
efficient infection.
Genome Segments of Influenza viruses
Influenza A
Influenza C
RNA
Segment
Length
(nt)
1
2341
PB2
759
2
2341
PB1
3
2233
PA
4
2073
HA
566
5
1565
NP
498
N ucleocapsid
protein
6
1413
NA
454
N euraminidase
7
1027
M1
252
Matrix protein
spliced
M2
97
934
NS1
230
spliced
NS2
8
Encoded Protein
Name
(aa)
RNA
Segment
Length
(nt)
Cap recognition,
RNA synthesis
1
2365
PB2
774
757
RNA synthesis
2
2363
PB1
754
716
RNA synthesis
3
2183
PA
709
4
2073
HEF
655
5
1809
NP
565
6
1180 spliced
M
242
CM2
(139)
139
NS1
286
NS2
122
Function
121
H emagglutinin, fusion,
major surface antigen, sialic
acid binding. HEF of
FLUCV also has esterase
activity
Ion channel
??
N onstructural protein
N uclear export protein
internal
initiation
7
??
Encoded Protein
Name
(aa)
934
spliced
Synthesis of mRNAs and RNA Replication
5'
ppp-AGC
G
A
3'
CCUUGUUUCUACU
AAAGCAGG
vcRNA
15-22nt
Replication
3'
HO-UCG
5'
C
UUUCGUCC
U
"Cap-snatching"
7
m
m GpppX
Y
A
G GC AAAGCAGG
10-13 nt A
UUUUUU
mRNA synthesis
5'
GGAACAAAGAUGA
Genome
RNA
3'
AAAA AAAAAAAAA(PolyA)
mRNA
Splicing to Produce Influenza A mRNAs
M1 protein
(252aa)
Translation
5'
Poly(A)
CAP
3'
Cap-snatching, mRNA
synthesis
3'
Cap-snatching, mRNA synthesis,
splicing
5'
CAP
M1 mRNA
5'
3'
Poly(A)
Genome
Segment 7
M2 mRNA
Translation
M2 protein
(97aa)
Since influenza RNA synthesis occurs in the nucleus, the cellular splicing
machinery can be used
In Flu A two mRNAs are produced from both segments 7 and 8
One mRNA is unspliced, the second is spliced
Influenza C Has an Esterase
Flu C lacks NA and has only 7 segments
It has HEF that performs the functions of HA and NA in Flu AB
The receptor for Flu C is 9-O-acetyl-N-acetyl neuraminic acid
The Flu C esterase removes the 9-O-acetyl group to destroy the
receptor
The HEF gene is also present in some coronaviruses,
which must have obtained it by recombination with Flu
C at some time in the past
M2
M2 tetramers form ion channels in viral and cellular
membranes
Exposure to low pH is required to dissociate the nucleocapsid
from the matrix protein, allowing the nucleocapsid to be
transported to the nucleus
M2 also prevents premature activation of the fusion
activity of HA
Amantadine interferes with the function of M2 and is an
effective flu antiviral
Virus Assembly
Nucleocapsids assemble in the nucleus during genomic RNA synthesis
The encapsidation signal is at the end of the RNA and not present in mRNAs
Nucleocapsids are exported to the cytoplasm in a process that requires NS2
and M1
Glycoproteins are synthesized on the ER and transported to the
plasma membrane
Nucleocapsids bud through the plasma membrane to form virions
More than 8 segments may be packaged: Ten segments randomly selected
would result in ~3% of progeny virions having at least one each of
the 8 segments
Random selection of segments would mean efficient reassortment
during mixed infection, which is known to occur
Influenza - Some History
Oldest record of an epidemic probably caused by flu: Hippocrates, 412 BC.
Epidemics have occurred relatively frequently but at irregular intervals
Epidemics vary in severity but the very young and elderly are most at risk.
Epidemics appear to radiate from specific locations
Example: 1781 epidemic that spread across Russia from Asia.
Influenza has killed untold millions throughout the centuries
1. 1918-1919 epidemic was particularly severe
2. 20-1000 million people died, more than died in World War I.
3. 80% of US WWI deaths were due to influenza
4. A significant factor in the German loss was influenza
First human influenza virus was isolated in 1933.
Different strains cause different epidemics, but human strains can recirculate
Antigenic Shift and Drift in Flu A
HA and NA are the major surface antigens of the virus
Antigenic drift describes the selection of variants by the immune system
Relatively slow
Resistance is only partial
Antigenic shift describes the results of recombination (reassortment)
There are 15 different subtypes of HA
There are 9 different subtypes of HN
Subtypes differ by 30% or more in amino acid sequence
A reassortant with a different HA and/or HN may cause a pandemic
Only a few of the subtypes have been isolated from humans
Influenza A in Birds
The reservoir of influenza A in nature is birds
All 15 HA and 9 NA have been found in aquatic birds
In particular, migratory ducks are important in the maintenance and
spread of influenza
Influenza infection of birds is usually asymtomatic
Influenza replicates in the respiratory tract and the intestinal tract of birds
It is excreted in the feces and high concentrations have been
found in waters in which migratory ducks congregate
The virus appears to be in equilibrium in birds--little or no sequence drift
has been found in bird viruses and disease seldom results from infection
In contrast, the virus drifts rapidly in humans and vaccines
must be reformulated yearly, and serious illness is produced
Epidemic Influenza Strains
Year
Virus
Common Name
1889
H2N2
1900
H3N8
1918
H1N1
Spanish
1957
H2N2
Asian
1968
H3N2
Hong Kong
1977
H1N1
Russian
When a new strain appears the previous strain usually dies out
At present, H3N2 and H1N1 continue to cocirculate in humans
An H1N1 strains has circulated continuously in pigs in the U.S. since 1918
Sialic Acid (N-Acetyl Neuraminic Acid)
Terminal NANA is attached to galactose by a2,3 or a2,6
linkages
Different HAs prefer one or the other linkage
Avian intestine contains predominantly a2,3 linkages
Human trachea contains predominately a2,6 linkages
Pig trachea contains both linkages and serves as an
efficient intermediate host in which reassortment can
take place--pigs are often referred to as mixing chambers
Other components also contribute to host specificity, best
studied for NP
U.S. Life Expectancy
1918 Influenza Deaths
The 1918 Flu in America
Excess Mortality Caused by Influenza A and B Virus in the United
States Between 1934 and 1990.
8
Influenza affects
10-20% of U.S.
population each
year, causing up
to 70,000 deaths.
Average death
rate in people
over 65 is 1/2200
but in 1957-8 it
was 1/300.
Excess Mortality (Deaths X 10 -4)
Type H2N2 appears
Type H3N2 appears
6
4
2
0
1935 1940 1945 1950 1955 1960 1965 1970 1975 1980 1985 1990
Year
Influenza A type H1N1
Influenza B
Influenza A type H2N2
Cocirculating B and A
Influenza A type H3N2
Illness Induced by Influenza Virus
Influenza virus infects superficial cells throughout the respiratory tract.
There is little or no spread to other organs.
High temperatures often accompany the infection, 38-41 C, that last 3-6 days.
Cough and weakness can last 1-2 weeks longer.
Extensive destruction of epithelial cells of the LRT can result in primary viral
pneumonia.
Influenza infection can result in secondary bacterial infection of the LRT
resulting in bacterial pneumonia.
Death following influenza infection is usually due to pneumonia, whether viral or
bacterial or combined.
Immunity following influenza infection is incomplete and appears to fade in time.
It has been suggested that the high death in young adults in the 1918 pandemic
could have resulted from a more active immune response to the virus.
Bird Influenza
An epidemic of influenza in chickens occurred in Hong Kong in 1997
The virus was highly virulent, killing 70-100% of infected chickens
Bird viruses are not normally transmitted to humans but the 1997
Hong Kong virus resulted in 18 humans becoming infected
This virus was highly virulent in humans--6 of 18 infected people died
The virus was H5N1 and did not spread in humans--no person to
person transmission occurred
To eradicate the virus and to prevent new reassortants from
arising that might give rise to epidemic virus by direct person to
person transmission, 1.6 million chickens were slaughtered
Defenses against Influenza
Antivirals
Amantadine and Rimantadine licensed for use and ameliorate symptoms
Inhibitors of NA being developed
Vaccines
Inactivated vaccines are in widespread use
These vaccines must be reformulated every year because of shift
and drift
They are 60-80% effective
Attempts being made to develop attenuated virus vaccines that could
be reformulated yearly by reassortment
An emergency response to swine flu in 1976 demonstrates the
difficulties in preparedness decisions
Bunyaviridae
Sin Nombre Virus
La Crosse Virus
BUNYAVIRIDAE
GENUS/
MEMBERS
USUAL
HOST(S)
TRANSMISSION/
VECTOR
HUMAN
DISEASE
WORLD
DISTRIBUTION
BUNYAVIRUS ( ~150 types)
Bunyamwera
Rodents, rabbits Aedes mosquitoes
Febrile illness
Worldwide
La Crosse
Humans,rodents Aedes triseriatis
Encephalitis
Midwest US
Snowshoe hare
Lagomorphs
Mosquitoes (Culiseta Rarely infects
and Aedes)
humans
California
encephalitis
Rodents, rabbits
Aedes melanimon
A. dorsalis
Encephalitis
(rare)
Western US,
Canada
Jamestown Canyon
w hite-tailed deer
Aedes species,
C. inornata
Increasing
North America
Apodemus
agrarius
Rattus species
Feces,urine,
saliva
Feces,urine,
saliva
Hemorrhagic
fever
Hemorrhagic
fever
Worldwide
HANTAVIRUS
Hantaan
Seoul
Northern US
Eastern Asia,
Eastern Europe
Prospect Hill
Microtus
None?
pennsylvanicus
Sin Nombre
Peromyscus
maniculatus
Feces,urine,
saliva
Sheep, goats
Tick-borne
Africa
Nairobi sheep disease. Sheep, goats
Tick-borne
Africa
NAIROVIRUS
Dugbe
Crimean-Congo
hemorrhagic fever
Humans, cattle, Tick-borne
sheep, goats
United States
Pulmonary
syndrome
Western US and
Canada
Hemorrhagic
fever
Africa, Eurasia
Nonfatal febrile
illness
Hemorrhagic
fever
??
Mediterranean
PHLEBOVIRUS (~50 types)
Sandfly fever Sicilian Humans
Rift Valley fever
Sheep,humans,
cattle, goats
Uukuniemi
Birds
Phlebotomous
flies
Mosquitoes, also
contact, aerosols
Tick-borne
Plants
Thrips
Africa
Finland
TOSPOVIRUS
Tomato spotted wilt
None
Australia, Northern
hemisphere
Genome Organization of the
Minus strand
genome segments
( range of sizes in kb)
S RNA
M RNA
L RNA
(0.94 - 2.9 kb)
(3.6-4.8 kb)
(6.4-8.9kb)
3’
BUNYAVIRUS
Bunyaviridae
5’
3’
5’
3’
5’
N
(BUNV)
Post-translational cleavage
NS s
Nested reading frames
NS s
C
PHLEBOVIRUS
G2
NS m
L
G1
N
(RVFV)
Post-translational cleavage
L
N
Ambisense transcription and translation
C
TOSPOVIRUS
NS s
NS m G1/G2
G1/G2
N
C
NS m
N
(TSWV)
L
N
G2
Ambisense transcription and translation
G1
NAIROVIRUS
(C-CHFV)
N
Post-translational cleavage
G2
L
G1
HANTAVIRUS
(HTNV)
N
Post-translational cleavage
G1
G2
L
Ambisense Coding Strategy of Bunyavirus S RNA
NSs protein
3'
Translation
5'
CAP
mRNA
synthesis
5'
NS s mRNA
3'
vcRNA
Replication
3'
mRNA
synthesis
3'
Translation
N protein
Genome RNA
NmRNA
CAP
5'
5'
Phylogenetic Tree of Rodent-borne Hantaviruses
Rodent-borne Hantaviruses
76-118
cumc-b11
Hantaa
hojo
n
lee
isolates
hv114
b1
sr-11 Seoul
isolate
80-39
s
Thailand
Dobrava
*
vindeln
vranica
Puumal
cg1820 a
sotkamo isolates
90-13
Tula
Prospect Hill
Bayou
Black Creek Canal
Rodent hosts
Murinae
(Old world rats and mice,
found in Europe and Asia)
Arvicolinae
(Voles; found in Europe,
Asia, and the Americas)
Laguna Negra
Sin Nombre
New York
El Moro Canyon
Sigmodontinae
(New World rats and mice,
found only in the Americas)
Distribution of Various Hantaviruses in Eurasia
Latvi
a
R om
ani a
Ser
bi a
U kr
ai n
e
Hantaan virus
Variant Hantaan
Puumala virus
Variant Hantaan
and Puumala
H antavirus Pulmonary Syndrome in the Americas
Canada
Sin Nombre
New York
Monongahela
United States
Bayou
Black Creek Canal
Juquitiba
Brazil
Rio Mamore
Bolivia
Paraguay
Oran
Laguna Negra
Argentina
Uruguay
Number of HPS cases
1-10
Lechiguanas
Andes
11-50
51-150
>150
Chile
Arenaviridae
Budding Machupo Virion
Lassa Virions
Tacaribe Virion
Genome Organization and Replication Strategy of an Arenavirus
Lassa fever virus S RNA (3417nt)
G2 (234aa)G1 (256aa)
Translation
3' Cleavage
5'
mRNA synthesis
3'
5'
3'
5'
Replication
5'
GPC mRNA
vcRNA
Genome RNA
mRNA synthesis
Translation
3'
NmRNA
N protein (570 aa)
Z (99aa)
Translation
Lassa fever virus L RNA (7279 nt)
5
'
3
'
5
'
mRNA synthesis
Replication
mRNA synthesis
5
'
3
'
5
'
Z mRNA
vcRNA
Genome RNA
LmRNA
Translation
L protein (2218 aa)
Representative Arenaviruses
Rodent Host
Disease
Where Found
Mus musculus
Meningitis
Worldwide
Mastomys sp.
HF
West Africa
Praomys sp.
?
CAR
Sigmodon hispidus
None?
Florida
Neotoma albigula
Oryzomys albigularis
3 fatal ARDS
Western U.S.
None?
Colombia
Zygodontomys brevicauda
Venezuelan HF
Venezuela
Calomys musculinus
Argentine HF
Argentina
Calomys callosus
Bolivian HF
Bolivia
Sabia
?
3 severe cases
Brazil
Tacaribe
?
?
Trinidad
Old World
Lymphocytic
choriomeningitis
Lassa
Mobala
New World
Tamiami
Whitewater Arroyo
Pichinde
Guanarito
Junin
Machupo
Arenaviruses in the New World
Virus
Some Viruses
Causing
Disease
a
Geographic
Range
Vector transmission
Case Mortality b
%
Treatment
(Prevention)
ARENAVIRIDAE
Junin
Argentine HF
Argentine pampas
Infected field rodents,
Calomys musculinus
Antibody effective, ribavirin
probably effective; preventive
vaccine exists
Ribavirin probably effective
Machupo
Bolivian HF
Beni province,
Bolivia
Infected field rodents,
Calomys callosus
Guanarito
Venezuelan HF
Venezuela
Infected field rodents,
Zygodontomys brevicauda
No data for humans,
ribavirin probably effective
Sabiá
HF
Rural areas near
Salo, Brazil
Unidentified
infected rodents
Intravenous ribavirin
effective in one case
Lassa
Lassa fever
West Africa
Infected
Mastomys rodents
15
Ribavirin effective
Rift Valley fever
Rift Valley fever
Sub-saharan
Africa
Aedes mosquitos
50
Rapid course; ribavirin or
antibody might be effective
Crimean-Congo
HF
CrimeanCongo HF
Hantaan, Seoul,
Puumala, and
others
Sin Nombre and
others
HFRS
15-30
BUNYAVIRIDAE
Hemorrhagic fever
Africa, Middle East,
Balkans, Russia,
W. China
Worldwide
(See Fig 4.24)
Tick-borne
Each virus maintained in a
single species of infected
rodents
15-30
Variable
Ribavirin used and
probably effective
c
Ribavirin useful; supportive
therapy is mainstay
HPS, also
rare HF
Americas
(See Fig 4.25)
Filovirus HF
Africa
Unknown
Yellow fever
Yellow fever
Africa, South
America
Aedes mosquitos
20
Very effective
vaccine
Dengue
DHF,DSS
Tropics and
subtropics
worldwide
Aedes mosquitos
<1
Supportive therapy useful;
vector control
Kyasanur forest
disease
KFD
Mysore State,
India
Tick-borne
Omsk
hemorrhagic
fever
OHF
Western Siberia
Poorly understood cycle
involves ticks, voles,
muskrats??
As for viruses causing
HFRS
40-50
Rapid course makes
specific therapy difficult
FILOVIRIDAE
Marburg,
Ebola
Marburg -25
EbolaZ 30-90
No effective therapy, barrier
nursing prevents spread
of epidemics
FLAVIVIRIDAE
0.5 - 9
?
a Abbreviations used: HF hemorrhagic fever; HFRS - hemorrhagic fever with renal syndrome; HPS - hantavirus
pulmonary syndrome; DHF - dengue hemorrhagic fever; DSS - dengue shock syndrome; KFD - Kyasanur Forest disease;
OHF - Omsk hemorrhagic fever
c
b
In humans.
Hantaan is 5-15% fatal, while Puumala is <1% fatal.
This table includes data from Nathanson et al. (1996) Table 32.1 on p. 780.
????
N eeds further study
Representative Viruses Causing Encephalitis
Flaviviridae
Alphaviruses
St. Louis encephalitis
Eastern equine enceph.
Japanese encephalitis
Western equine enceph.
West Nile
Venezuelan equine enceph.
Murray Valley enceph.
Tick-borne enceph.
Bunyaviridae
Herpesviridae
Herpes simplex
Paramyxoviridae
La Crosse
Mumps
California enceph.
Measles