VIRAL AGENTS CAUSINGVIRAL AGENTS CAUSING
GASTROENTERITISGASTROENTERITIS
1. Rotavirus
2. Enteric adenoviruses
3. Noroviruses :
a. Norwalk-like viruses
b. Calicivirus
c. Astrovirus
Major Viruses
Viruses associated with gastroenteritis (cont) :
Other viruses (minor):
• Coronaviruses
• Parvoviruses
• Pestiviruses
• Toroviruses
ROTAVIRUSROTAVIRUS
• First isolated in 1973 from children with
diarrhea
• EM identification from duodenal biopsies
• Human and animal strains
ROTAVIRUSROTAVIRUS
• 60-80nm in size
• Non-enveloped virus
• Double capsid
• EM appearance of a wheel with radiating spokes
• Icosahedral symmetry
• double stranded (ds) RNA in 11 segments
(double – double)
Viral Structural Proteins (VP)Viral Structural Proteins (VP)
• Outer structural proteins - VP7 and VP4
VP7VP7=glycoprotein
VP4VP4=protease-cleaved, P protein, viral
hemagglutinin, and forms spikes from the surface
• Inner core structural proteins VP 1, 2, 3, 6
• VP6 is an important antigenic determinant
• Genome is composed of 11 segments of double-stranded RNA,
six structural
coding for proteins
five nonstructural
• Seven serological groups have been identified (A-G), three of
which (groups A, B, and C) infect humans .
ClassificationClassification
•Groups, subgoups, serotypes based on viral capsid
proteins
•7 Groups (A through G)
•Group A is the most common and has 2 subgroups
•10 human serotypes based on G protein (VP 7)
•8 P protein serotypes
ClassificationClassification (contd.)
•Electropherotypes
mobility of RNA segments by PAGE
Used in epidemiologic studies
Rotavirus - PropertiesRotavirus - Properties
• Virus is stable in the environment
• Relatively resistant to handwashing agents
• Susceptible to disinfection with 95% ethanol,
‘Lysol’, formalin
PathogenesisPathogenesis
• Targeted host cells- mature enterocytes lining
the tips of intestinal villi
• Intermediate/infective sub-viral particle (ISVP)
produced through proteolysis
• Enter host cell by endocytosis
• Virus replicates in the host cell cytoplasm
ReplicationReplication
• mRNA transcription with viral RNA polymerase
• Capsid proteins formed
• mRNA segments formed, assembled into immature
capsid
• mRNA replicated to form double stranded RNA
genome
HistopathologyHistopathology
• Mature enterocytes lining the tips of intestinal
villi are affected
• Villous atrophy and blunting
• Death of the mature enterocytes
Epidemiology - WorldwideEpidemiology - Worldwide
• Millions are affected
• 600,000-850,000 deaths/year
• A major cause of diarrhea-associated
hospitalizations
• Seroprevalence studies show that antibody is
present in most by age 3y.
Rotavirus-Rotavirus-Worldwide distributionWorldwide distribution
(source- centers for disease control and prevention)
Epidemiology : U.S.Epidemiology : U.S.
•No. of children under 5y. affected ~ 2.7 million
•Physician visits per year ~ 500,000
•Hospitalizations per year ~ 50,000
•Deaths per year ~ 20 - 40
•% cases w/ dehydration ~ 1-2.5%
EpidemiologyEpidemiology
• Age- 4mo - 2 years
Protection of younger infants through transplacental
antibody transfer
• Asymptomatic infections are common, especially in
adults
• Nosocomial infections
• Outbreaks
EpidemiologyEpidemiology (contd.)
• Seasonality
Winter months (Nov. through May in US)
Gradual spread W to E
Year-round in the tropics
• Incubation period - thought to be <4 days
EpidemiologyEpidemiology (transmission)
• Mainly person to person via fecal-oral route
• Fomites
• Food and water-borne spread is possible
• Spread via respiratory route is speculated
EpidemiologyEpidemiology (spread)
• Contagious from before onset of diarrhea to a few
days after end of diarrhea
• Large amounts of viral particles are shed in diarrheal stools
• Infective dose is only 10-100 pfu
EPIDEMIOLOGYEPIDEMIOLOGYDifferences in Groups
• Group A infections are most common
• Group B has been associated with outbreaks in
adults in China
• Group C is responsible for sporadic cases of
diarrhea in infants around the world
Clinical FeaturesClinical Features
• Incubation period - thought to be <4 days
• Fever- can be high grade (>102F in 30%)
• Vomiting, nausea precede diarrhea
• Diarrhea
- usually watery (no blood or leukocytes)
- lasts 3-9 days
- longer in malnourished and immune deficient indiv.
- NEC and hemorrhagic GE seen in neonates
Mechanism of diarrheaMechanism of diarrhea
• Watery diarrhea due to net secretion of intestinal fluid
• Activation of the enteric nervous system -possible role of enterotoxin
Clinical FeaturesClinical Features (contd.)(contd.)
• Dehydration is the main contributor to mortality.
• Secondary malabsorption of lactose and fat, and chronic diarrhea are possible
• Recovery is usually complete.
• However, severe diarrhea without fluid and electrolyte
replacement may result in dehydration and death .
Immunological Aspects
• Immunoglobulin (Ig) A , in the lumen of the gut immunity
to infection .• Actively or passively acquired antibodies (including antibodies
in
colostrum and mothers milk) lessen the severity of disease
but does not consistently prevent reinfection .• Absence of antibody small amounts of virus infection
and diarrhea .
DiagnosisDiagnosis
• Antigen detection in stool by ELISA, Latex
Agglutination (for Group A rotavirus)
• EM- non-Group A viruses also
• Culture- Group A rotaviruses can be cultured in
monkey kidney cells
• Serology for epidemiologic studies
Treatment and PreventionTreatment and Prevention
• Treatment- Treatment-
Supportive - oral, IV rehydration
• PreventionPrevention-
Handwashing and disinfection of surfaces
VaccineVaccine• Live tetravalent rhesus-human reassortant vaccine
(Rotashield)
• Licensed for use in August 1998
• Removed from the market in October 1999 due to
risk of intussusception
• Cases were seen 3-20 days after vaccination
• Approx. 15 cases/1.5 million doses
• New vaccine from bovine rotavirus under trial
Production of rhesus rotavirus (RRV), human rotavirus (HRV) x rhesus rotavirus (RRV)reassortant quadrivalent vaccine wit VP7 serotype 1, 2, 3, and 4 specificity
GASTROENTERITIS DUE GASTROENTERITIS DUE TO ADENOVIRUSTO ADENOVIRUS
• Types 40, 41
• Belong to serogroup F
• Some cases due to types 31, 3, 7
Diarrhea due to Diarrhea due to Enteric AdenovirusEnteric Adenovirus
• Age <4 years
• Year round
• Spread via fecal-oral route
Clinical features of Enteric Clinical features of Enteric Adenovirus gastroenteritisAdenovirus gastroenteritis
• Incubation period 3 -10 days
• Diarrhea lasts for 10 -14 days
• Can also cause intussusception, mesenteric
adenitis, appendicitis
Diagnosis- Enteric adenovirusesDiagnosis- Enteric adenoviruses
• Isolation requires special media-Graham 293
• ELISA for rapid detection is available
HUMAN CALICIVIRUSESHUMAN CALICIVIRUSES(HuCV)(HuCV)
•Belong to Family Caliciviridae
•Non-enveloped RNA viruses
with ss RNA
•27-35 nm in size
•Contain a single capsid protein
HUMAN CALICIVIRUSESHUMAN CALICIVIRUSES
•Genomic analysis divides it into 4 groups
•Human caliciviruses belong to 2 genera
CLASSIFICATION OF HuCVCLASSIFICATION OF HuCV
NLV (Norovirus)Norwalk virus
Hawaii virus
Snow Mountain virus
Montgomery county virus
Taunton (England)
SLV (Sapovirus)
Sapporo virus
Manchester virus
Houston/86
London/92
Morphology of HuCV- typicalMorphology of HuCV- typical
• Typical morphology
• 32 cup-like depressions
• EM appearance of “Star of David”
E.g.- Sapporo-like viruses
Morphology of HuCV- atypicalMorphology of HuCV- atypical
• Atypical morphology
• Smooth surface
• Small Round Structured viruses
E.g.- Norwalk-like viruses
CLINICAL FEATURESCLINICAL FEATURES
• Adults and Children
• Usual incubation Period is <24 hours
(ranges from 12hrs. to 4 days)
• Short duration of illness <3 days
• Nausea, vomiting, fever, headache
• Abdominal cramping
• Watery diarrhea
Epidemiology-NorovirusesEpidemiology-Noroviruses
• Worldwide distribution
• >23 million cases/year in the U.S.
• Major cause of foodborne outbreaks of GE
• Most people have had infections by age 4
years (by seroprevalence studies)
Spread of Norwalk virusSpread of Norwalk virus
A. Person-to-person Fecal-oral spread (stool/vomitus)
B. Fecal contamination of food or water
C. Spread through fomites?
Epidemiology-NorovirusesEpidemiology-Noroviruses
•Asymptomatic infections- seroconversion but asymptomatic shedding of virus
•Low infective dose •Viral excretion during convalesence
(up to 2 weeks)•Ability to survive in water chlorination at
routine levels
Epidemiology of OutbreaksEpidemiology of Outbreaks
• Cruise ships, schools, nursing homes, etc. • Can involve infants and school-age children
• Source usually is contaminated food and water (seafood-oyster and shellfish etc.)
Diagnosis- Human Diagnosis- Human CalicivirusesCaliciviruses
• Specimen- stool , vomitus, environmental swabs,
[not yet on foods]
• Immune EM
• RT-PCR in state public health labs.
• Serology for epidemiologic purposes
ASTROVIRUSASTROVIRUS
• Described in relation to an outbreak of
gastroenteritis in 1975
• Detected by EM
• Immunologically distinct from Human Caliciviruses
• Belong to family Astroviridae
• 8 human serotypes are known
ASTROVIRUS- structureASTROVIRUS- structure
• Small ss RNA virus
• Non-enveloped
• 27-32nm in size
• Round with an unbroken, smooth surface
• EM appearance of a 5 or 6 pointed star within
smooth edge
• Contain 3 structural proteins
ASTROVIRUSASTROVIRUS - Epidemiology
• Worldwide
• Mainly in children <7 years of age.
• Transmission person-to-person via
fecal-oral route
• Outbreaks due to fecal contamination of
sea-food or water
ASTROVIRUSASTROVIRUS - Clinical Features
• Infants and children are most often affected
• Short incubation period 1-4 days
• Nausea, vomiting, abdominal cramping and watery
diarrhea
• Constitutional symptoms-fever, malaise, headache
• Twenty-five years ago, little was known about the causes of diarrhea, which kills an estimated 3 million infants and children worldwide every year.
• Scientists knew that bacteria and parasites were implicated in only approximately 10 to 20 % of all cases of diarrhea..
• In 1973,, researchers in Australia discovered a virus in infants with severe diarrhea and named it “rotavirus” for its wheel-like shape.
• One year later, NIAID researchers were the first to identify rotavirus in the United States.
Rotavirus Biology
•Rotaviruses belong to the family Reoviridae ,
genus Rotavirus.
• They have a characteristic wheel-like appearance when viewed by electron microscopy.
•Nonenveloped, double-shelled
Group A rotaviruses
• Endemic worldwide (represents >
95% of
currently identified strains in humans
)
• The leading cause of severe diarrhea among infants and children . • Accounts for about half of the cases requiring hospitalization.
Group B rotavirus,
• Also called adult diarrhea rotavirus or
ADRV
• Has caused major epidemics of severe
diarrhea affecting thousands of persons
of all ages in China.
Group C rotavirus•
• Has been associated with rare and sporadic
cases of diarrhea in children in many
countries.
• First outbreaks were reported from Japan
and England
• Subgroups classification based upon
neutralization epitopes of the outer capsid proteins,
VP4 and VP7
Antigenic specificity of VP7 G serotypes
Antigenic specificity of VP4 P serotypes
Fourteen G serotypes and twenty one P serotypes have been detected in humans.
Neutralization assays measure reactivity
predominantly to VP7 proteins.
Transmission
• Rotavirus infection is very
contagious .
• Viral particles pass in the stool of infected persons before and after they have symptoms of the illness .
• Spread is by the oral-fecal route
• The virus can survive for long periods on hard surfaces (e.g.
toys and tables) and is not killed by standard
disinfectants... children forget to wash their hands often
enough, especially before eating and after using the toilet.
toilet Get infected
• Infected food handlers may contaminate foods that require
handling and no further cooking, such as salads, fruits.
• The infective dose is presumed to be 10-100 infectious viral
particles.
Because a person with rotavirus diarrhea often excretes large
numbers of the virus (108-1010 infectious particles/ml of
feces), infection doses can be readily acquired .
• Asymptomatic rotavirus excretion has been well
documented may play a role in perpetuating
endemic disease .
N.B some have reported low titers of virus in respiratory
tract secretions and other body fluids..
• Rotavirus is the single most important cause of life-threatening diarrhea in children younger than 2 years.
• Affects approximately 130 million infants and children worldwide.
• In the United States alone, rotavirus causes more than 3 million cases of childhood diarrhea each year, leading to an estimated 55,000 to 100,000 hospitalizations and 20 to 100 deaths.
• Humans of all ages are susceptible to rotavirus infection,
although Children , premature infants, the elderly,
and the immunocompromised are prone to more severe
symptoms caused by infection with group A rotavirus.
Who catch the infection ?
• An infant’s first bout of diarrhea from rotavirus is the most severe , subsequent reinfections decrease in severity. These findings indicated that infants gradually develop partial immunity to the virus and that a vaccine might prevent the disease.
• Temporary lactose intolerance may occur.
• NSP4 protein may act in a toxin-like manner
Calcium ion influx into enterocytes
Release of neuronal activators
Neuronal alteration in
water absorption
• Loss of the ability to absorb water net secretion
of water and loss of ions watery diarrhea
• The incubation period ranges from 1-3 days .
• Symptoms often start with vomiting followed by 4-8 days of
diarrhea.
Symptoms and Signs
• Some may have a slight rise in temperature .
Immunological Aspects
• Immunoglobulin (Ig) A , in the lumen of the gut immunity
to infection .• Actively or passively acquired antibodies (including antibodies
in
colostrum and mothers milk) lessen the severity of disease
but does not consistently prevent reinfection .• Absence of antibody small amounts of virus infection
and diarrhea .
• Infection in infants and small children is generally symptomatic .
• In adults infection is usually asymptomatic .
• Asymptomatic rotavirus infections are common in neonates
because of passively acquired maternal immunity , breast
feeding , and possible infection with less virulent strains
(EIA) 1) Identification of the viral antigen in the patient's stool latex
agglutination most widely used screening test for clinical specimens .
2) Electron microscopy (EM)
3) polyacrylamide gel electrophoresis (PAGE) is used in some laboratories
for RNA typing.
4) A reverse transcription-polymerase chain reaction (RT-PCR) has been
developed to detect and identify all three groups of human rotaviruses.
Diagnosis
• Serotypes can be identified using monoclonal antibodies
against VP7 and VP4.
• Neutralization is detected on tissue cultures as a CPE
Antigenic specificity of VP7 G serotypes
Antigenic specificity of VP4 P serotypes
• Treatment is nonspecific and consists of : 1) Oral rehydration therapy to prevent dehydration.
2) About one in 40 children with rotavirus gastroenteritis will
require hospitalization for intravenous fluids .
• For persons with healthy immune systems, rotavirus gastroenteritis is a self-limited illness, lasting for only a few days.
Treatment
• Total prevention of the spread of rotavirus is virtually impossible.
prevention
• In hospitals health officials control rotavirus outbreaks by isolating
infected patients and by ordering strict hand-washing procedures.
• Even in the cleanest environments with the best hygiene, most children
still become infected with rotavirus before age 4 or 5.
Rotavirus vaccine
• Scientists knew that although many strains of rotavirus exist, only four
cause the majority of diarrhea cases in young children in the United
States.
• Aiming for prevention : NIAID researchers developed a vaccine
(RRV-TV) designed to protect against the four strains of rotavirus .
• During the 1970s, NIAID scientists analyzed the genetic material of
rotavirus, Identified two important proteins , VP4 and VP7 produced
by the genes , and determined the function of these proteins.
• Proteins on the surface of the virus were found to be critical for triggering
an immune response in the body against rotavirus.
• NIAID researchers focused on these proteins to develop a vaccine.
Tetravalent Oral Live-Attenuated Vaccine
• The oral vaccine contains four different, live attenuated viral strains,
serotypes ( 1,2,3,4) .
• One strain (serotype 3) is an unmodified rhesus monkey rotavirus
(RRV) which does not cause disease in humans
• The other three are made by reassortment (genetic recombination)
of that monkey RRV with three human rotaviruses of serotypes
1,2,4.
• Each reassortant vaccine strain contains 10 monkey RRV genes and
the VP7 gene for one serotype of the human rotavirus envelope
proteins:
VP-7 (serotype 1) , VP-7 (serotype 2) , VP-7 (serotype 4)
• The combined vaccine provided comprehensive protection
against the four serotypes (1,2,3,4)
• Studies showed that high doses of the RRV-TV vaccine, designed to protect against four strains of rotavirus, were very effective in
preventing severe, dehydrating rotavirus disease.
• Breast-feeding did not interfere with the effectiveness of the rotavirus vaccine ensuring good nutrition in infants
• In August 1998, the first live attenuated rotavirus vaccine
(Rotashield{registered} {Wyeth Lederle Vaccines and
Pediatrics}) was approved for use in infants by the Food and
Drug Administration. The Advisory Committee on Immunization
Practices has recommended that this vaccine be given as a
three-dose schedule to infants aged 2, 4, and 6 months.
•However, on July 15, 1999, the US Centers for Disease Control and
Prevention (CDC) recommended that doctors stop giving the
rotavirus vaccine to infants.
• On October 22, 1999, the Advisory Committee on Immunization
Practices voted to stop recommending the vaccine
Why?
• Centers for Disease Control and Prevention (CDC) advisory committee
received an overwhelming amount of data all indicating a strong
association between ( rotavirus vaccine ) and bowel obstruction among
some infants during the first one to two weeks following vaccination.
• Apparently, many infants who received the rotavirus vaccine developed
Intussception of the bowel within one to three weeks after receiving
a dose or two of the vaccine.
• The risk of intussusception was increased 19-fold in the first 3 to 7
days after vaccination and almost fourfold (3.6) in the 8 to 14 days
after vaccination (P<0.0002).
• Children who have already received the vaccine and have not had
problems do not appear to be at risk now.
• In the meantime, research on better vaccines for rotavirus continues.
1293
585 females ( 45.24%)
All were tested using enzyme linked immunosorbent
assay (ELISA).
708 males (54.76%)
Number of infected patients 136
73 males (53.68%)
63 females (46.32%)
% of infected patients 10.52
5.65% males
4.87% females
AGE
adultsschoolpreschooltoddlersinfantsneonates
Count
40
30
20
10
0
SEX
male
female3
4
7
17
21
6
3
6
22
30
7
Numbers of Infected Males to Females in Different Age Groups
0-28 28d-1y 1-3 3-6 6-18 >18
27
24
1210
15
12
13
10
malefemale
sex
Bars show counts
Infections in( Males : Females ) in Different Seasons
DURATION
10.08.06.04.02.00.0
80
60
40
20
0
Std. Dev = 1.77
Mean = 3.3
N = 118.0077
76
22
4
Duration of Illness
(days)
SEASON
automnsummerspringwinter
Count
30
20
10
0
SEX
male
female
10
12
10
24
13
15
12
27
Number of Patients in Different Seasons (Males : Females )
SEASON
automnsummerspringwinter
Count
30
20
10
0
SEX
male
female
10
12
10
24
13
15
12
27
Number of Patients in Different Seasons (Males : Females )
ALL NATIONALITIES
Count
50
40
30
20
10
0
SEX
male
female
Number of Infected Males & Females in Each Nationality
ALL NATIONALITIES
Count
40
30
20
10
0
SEASON
winter
spring
summer
automn
Infections Among Different Nationalities in Each Season