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Gastroenteritis Viruses
Prof. Mary K. Estes
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Gastroenteritis VirusesGastroenteritis VirusesGastroenteritis VirusesGastroenteritis Viruses
Mary K. Estes, Ph.D.Professor of Molecular Virology and Microbiology
and Medicine-GI, BCM
Director, Texas Medical Center DDC
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Outline
• Gastrointestinal viruses
–Rotaviruses
–Noroviruses
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• The amount of diarrheal water
equals the amount of water
over Victoria Falls in 1 min
• Diarrhea is caused
by many infectious agents,
including viruses
• It has been estimated
that in any given 24 hr period,
200 million people on Earth
have gastroenteritis
Diarrhea is a health problem worldwide
Gastroenteritis Viruses
Prof. Mary K. Estes
The screen versions of these slides have full details of copyright and acknowledgements 2
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Viruses causing diarrheamajor pathogens in humans
• Rotaviruses
• Norwalk virus/Norovirus (Calicivirus)
• Sapovirus (Calicivirus)
• Astroviruses
• Adenoviruses (serotypes 40 and 41)
• Aichi virus
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Rotavirus
Dehydrating diarrhea
in young children
Viral gastroenteritis
Almost every child
(< 5 years) gets infected
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Epidemiology
World wide infection and disease
111 million (M) episodes of gastroenteritis
25 M clinic visits, 2 M hospitalizations
~ 440,000 deaths in children < 5 years
(Parashar et al., 2003, Emerg Infect Dis 9: 565-72)
Gastroenteritis Viruses
Prof. Mary K. Estes
The screen versions of these slides have full details of copyright and acknowledgements 3
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Rotavirus pathogenesis
• Disease is age-restricted:
– Virus replication in all ages
– Generally asymptomatic infection in adults
– Symptomatic infection occurs
in the young children 3 months to 2 years of age;
< 2 week old mice and rats
• Main tissue tropism – enterocytes
• Diarrhea caused by multiple factors:
– Malabsorption
– NSP4 enterotoxin
– Enteric nervous system
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Rotaviruses infect mature enterocytes of villus epithelium of the small intestine
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RV-gnotiobiotic piglets
Slide 7
mke1 mestes, 5/22/2007
Gastroenteritis Viruses
Prof. Mary K. Estes
The screen versions of these slides have full details of copyright and acknowledgements 4
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Rotavirus infection in calvesVillus blunting & loss of absorptive capacity
4 dpiUninfected
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Rotavirus infection
Time:
Virus replication:
Diarrhea:
0
0
0
Histopathology: None
7 dpi
0
0
None
Rotavirus pathogenesis
24 – 72 hpi
High titersduo<j ej = ileum
Villus atrophy and blunting,inflammation, mononuclearinfiltration of lamina propria,
v acuolation of epithelial cells
4+
ENS
12 hpi
Patchy v iral antigenduo<j ej <ileum
None
4+
Enterotoxin
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dsRNA
segment
1
23
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Protein
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6
789
10
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VP1VP2VP3
VP4
NSP1
VP6
NSP3
NSP2VP7
NSP4 - enterotoxi n
NSP5, NSP6
antigen
VP2
VP6Subgroup
SubcoreVP1, VP3
VP4
antigen
VP7
antigen
Neutralization
Neutralization
Rotavirus genes and proteins
Gastroenteritis Viruses
Prof. Mary K. Estes
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Rotavirus replication
• Occurs in the cell cytoplasm
• Viral RNA always is inside a protein shell
• Many aspects not well understood
• A unique process occurs
during the assembly of newly made particles
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αααα 2 ββββ 1 αααα v ββββ 3αααα x ββββ 2
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Participating proteins:
VP4, VP7, and NSP4
Transient lipid
bilayer formation
Following dissolution,
mature triple layered
particles are formed
Unique particle assembly by budding into the ER
• Is Ca2+-dependent ( requires ↑[Ca2+]i )
• Is NSP4-dependent
Gastroenteritis Viruses
Prof. Mary K. Estes
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Lumen
Cytoplasm
COOH
NH2
CHO
CHO
L25
ER membrane
175
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67 85
140 ?
112AAH
Toxin
93
Cleavage133
NSP4: an intracellular receptorand enterotoxin
H2
H1
H3
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NSP4-inoculated mouse
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Gastroenteritis Viruses
Prof. Mary K. Estes
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Properties of Rotavirus enterotoxin
• First described viral enterotoxin (NSP4)
• A novel toxin;
No sequence similarity to other know n toxins
• Induces age-dependent diarrhea
• Mobilizes intracellular Ca2+ – not age-dependent
• Does not induce histologic changes in the intestine
• Induces age-dependent chloride secretion
in intestinal mucosa of mice
• May be a target for treatments or vaccines
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How does the Rotavirus enterotoxin NSP4 act?
• NSP4 is released from virus-infected cells
• Cells have a receptor for NSP4
• Binding to this receptor causes cell signaling
that leads to chloride secretion and diarrhea
• Antibody to NSP4 will partially protect against disease
• Some avirulent rotaviruses have mutated NSP4 genes
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Effects of Rotavirus infection of polarized epithelial cells
• Disrupts
– Tight junctions
– Microvillar, microfilament network
• Perturbs protein targeting
– Sucrase-isomaltase
• Induces chemokine responses
• Evokes intestinal secretion
– Enterotoxin
– Enteric nervous system
Gastroenteritis Viruses
Prof. Mary K. Estes
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Enterocytes
Nucleus
NSP4
Cl -
Ca 2+
Cl -
Cl -
?
Courtesy of A. Einerhand
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New unexpected information about Rotavirus replication
Rotavirus infections are not limited to intestinal enterocytes
Rotavirus can be found in the serum of infected animals
and children (viremia)
Extraintestinal rotavirus can be seen
in the absence of diarrhea
Does extraintestinal Rotavirus infect other tissues
and cause non-diarrheal disease?
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Treatment and vaccines
Gastroenteritis Viruses
Prof. Mary K. Estes
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Treatment?
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Oral rehydration therapy
Per liter of clean water
• 90 mEq sodium
• 20 mEq potassium
• 80 mEq of chloride
• 30 mEq of citrate
• 20 g glucose
• ¾ T. table salt
• 1 T. baking soda
• 1 c. orange juice or 2 bananas
• 4 T. sugarOR
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Gastroenteritis Viruses
Prof. Mary K. Estes
The screen versions of these slides have full details of copyright and acknowledgements 10
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Vaccines?
• Live-attenuated reassortant vaccines
– Three currently approved and others being tested
– Not yet clear if these vaccines will protect
in developing countries where needed most
• Inactivated virus
• Virus-like particles
Need to be tested
as backup plan
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Conclusions
• Rotaviruses cause diarrhea by several mechanisms -
Enterotoxin, Malabsorption, ENS
• Why is disease age-restricted?
– Age-dependent chloride channel, Immunity
• Rotaviruses cause extraintestinal infections-
do they cause other diseases?
• Live, attenuated vaccines are safe and their effectiveness
in developing countries needs to be established
• New, effective vaccines and therapies may target
the rotavirus enterotoxin
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First identified human Calicivirus
Prototype strain of genetically related Noroviruses
23 million cases of gastroenteritis per year in USA
• Cause illness in babies, children, adults and elderly
• Cause outbreaks in daycare centers, schools,
nursing homes, hospitals, military
• Cause >96% of all outbreaks of nonbacterial gastroenteritis
Norwalk virus (NV)
Noroviruses
Gastroenteritis Viruses
Prof. Mary K. Estes
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• Highly stable
• Highly infectious (1-10 virions)
Norovirus gastroenteritis
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• Many asymptomatic infections
and prolonged post-symptomatic shedding
• Some people resistant to infection (20%)
– Possibly due to genetic factor?
• Persistent infections
– Transplant patients, immunosuppressive therapy
• Unusual clinical presentation
– Disseminated intravascular coagulation
• Spread by contaminated food,
water and environmental surfaces
Norovirus background
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Norovirus pathogenesis
• Incubation period 1 – 2 days
• Produces shortening of microvilli in small intestine
with infiltration of mononuclear cells
• Transient malabsorption
and delayed gastric emptying may occur
• Cause of vomiting unknown
• Immune responses poorly characterized
Gastroenteritis Viruses
Prof. Mary K. Estes
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Clinical responses of 2 adult volunteers given NV
Dolin et al., JID 123: 307-312 (1971)
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1970s NV studies
12 volunteers6 ill
NV
Immunity due to previous exposure?
Repeated susceptibilityRepeated resistance
6 not ill
6 ill
6 not ill
NV> 2 yrs
Genetic factor may determine susceptibility or resistance to NV
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Challenges studying Norwalk virus (NV)
• Infection and disease only in humans-
no small animal model
• No cell culture –
no in vitro system for infection and replication
• Low concentration of virions in stool
Gastroenteritis Viruses
Prof. Mary K. Estes
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Virus-like particles (VLPs)
• Expressed capsid proteins VLPs
- Virus surrogate
- Structurally and antigenically similar
to Norw alk virion capsids
- Model for Norw alk virus-cell binding
50 nm
(Prasad et al., Science 1999)
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Norwalk VLPs hemagglutinate
human red blood cells
Discovered the Norwalk VLPhemagglutination receptor is the H antigen
Model for Norwalk virus-host cell binding:
Hutson et al., J Virol (2003)
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What is the H antigen?
• Carbohydrate histo-blood group antigenon the cell surface and on secreted mucins
• Presence controlled by genetically determined
expression of a carbohydrate modifying enzyme, a fucosyltransferase, FUT2
FUT2-/- = 20% of population
FUT2+/+ = secretor
Henry et al., Vox Sang (1995)
Gastroenteritis Viruses
Prof. Mary K. Estes
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The H antigen is a host susceptibility factorfor Norwalk virus infection
Non-infected Infected
FUT2-/- 8 0
FUT2+/-, FUT2+/+ 1 42
n = 51
• FUT2-/- individuals are resistant to NV infection(0/8 infected, p < 0.0001)
Hutson et al. (J. Med. Virol. 77: 116; 2005)
FUT2 genotype versus NV challenge outcome
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Is host susceptibility similar for all Noroviruses ?
NO
• Hemagglutination patterns and CHO binding patterns
are strain specific
• Secretor status does not influence susceptibility
to Snow Mountain virus infection
• All people will be susceptible to some norov irus!
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Summary: Noroviruses
Clinical significance
is increasing –
numbers and settings
H histo-blood group
CHO antigen is host
susceptibility factor
Knowledge of NV-CHO
binding site (P domain)
or the structure of other
viral proteins may lead
to antivirals to help
prevent transmission
NN
CC
H 329H 329
W 375W 375N 331N 331
NN
CC
H 329H 329
W 375W 375N 331N 331
Gastroenteritis Viruses
Prof. Mary K. Estes
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