VIRAL AGENTS CAUSING GASTROENTERITIS

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VIRAL AGENTS CAUSINGGASTROENTERITIS
Helmut Albrecht, MDID Division Chief, USC
DOM(based on lecture by Nandini Narayan, MD)
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Viruses found in the gut (1)

• A. Associated with gastroenteritis
• Rotaviruses
• Adenoviruses 40/41
• Caliciviruses
• Norwalk like viruses or SRSV (Small Round
  Structured Viruses)
• Astroviruses
• SRV (Small Round Viruses)
• Coronaviruses
• Toroviruses

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Viruses found in the gut (2)

• B. Found in the gut, not normally associated with
  gastroenteritis
• Polio
• Coxsackie A
• Coxsackie B
• Echo
• Enteroviruses 68-71
• Hepatitis A
• Hepatitis E
• Adenoviruses 1-39
• Reoviruses
• C. Found in the gut as opportunistic infection
• CMV
• HSV
• VZV
• HIV

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VIRAL AGENTS CAUSINGGASTROENTERITIS

• Pathogens discussed in todays lecture
• Rotavirus
• Enteric adenoviruses
• Caliciviruses
• Astroviruses
• Toroviruses

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ROTAVIRUS

• Family Reoviridae
• Genus Rotavirus
• Other genera Orthreovirus, Coltivirus, orbivirus
  (sheep)

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ROTAVIRUS- discovery

• First isolated in 1973 in Australia by Ruth
  Bishop at the Royal Children's Hospital in
  Melbourne.
• EM identification from duodenal biopsies from
  children with diarrhea.
• "Virus particles in epithelial cells of duodenal
  mucosa from children with acute non-bacterial
  gastroenteritis," Lancet, 11281-3, 1973.
• Described in stool samples from children by
  Albert Z. Kapikian, in the US
• Human and animal strains are recognized

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Rotavirus
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ROTAVIRUS STRUCTURE
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STRUCTURAL FEATURES OF ROTAVIRUS

• 60-80nm in size
• Non-enveloped virus
• EM appearance of a wheel with radiating spokes
• Icosahedral symmetry
• Double capsid
• Double stranded (ds) RNA in 11 segments

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STRUCTURAL FEATURES OF ROTAVIRUS (contd.)

• Double capsid (outer and inner capsid)
• Core with genome
• Capsid is cleaved by trypsin to form ISVP
• intermediate/infective sub-viral particle

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Rotavirus structure
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ROTAVIRUS- ultrastructure
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ROTAVIRUSEM STRUCTURE
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VIRAL STRUCTURAL PROTEINS (VP)

• Outer structural proteins - VP7 and VP4 VP7
  - Glycoprotein
• VP4 - protease-cleaved, P protein, viral
  hemagglutinin forms spikes from the surface
• Inner core structural proteins VP 1, 2, 3, 6
• VP6 is an important antigenic determinant

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CLASSIFICATION- Groups

• 7 Groups (A through G) and 2 subgroups (Iand II)
  based on VP6 differences
• Group A is the most common
• Group B (outbreaks in China)
• Group C (worldwide)

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CLASSIFICATION - Serotypes

• Serotypes based on viral capsid proteins inducing
  neutralizing Ab
• 14 G serotypes based on G protein (VP 7)
  differences
• 5 predominant strains in U.S. (G1-G4, G9) account
  for 90 of isolates
• Strain G1 accounts for 73 of infections
• 20 P serotypes based on P protein (VP4) with
  P4/P8 predominance
• Common PG combinations are P8G1, P8G2, P4G2, P8G4

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CLASSIFICATION (contd.)

• Electropherotypes are based on the mobility of
• RNA segments by PAGE
• Useful in epidemiologic studies

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ROTAVIRUS- PROPERTIES

• Virus is stable in the environment (months)
• Relatively resistant to handwashing agents
• Susceptible to disinfection with 95 ethanol,
  Lysol, formalin

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PATHOGENESIS

• 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

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REPLICATION

• mRNA transcription with viral RNA polymerase
• Capsid proteins formed, assembled into immature
  capsid
• RNA replicated to form double stranded RNA genome

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HISTOPATHOLOGY

• Mature enterocytes lining the tips of intestinal
  villi are affected
• Villous atrophy and blunting
• Death of the mature enterocytes

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HISTOPATHOLOGY

• Infiltration of lamina propria with mononuclear
  cells
• Repopulation of the villous tips with immature
  secretory cells crypt hyperplasia

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HISTOPATHOLOGY
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EPIDEMIOLOGY

• A major cause of diarrhea-associated
  hospitalizations and deaths
• Sero-prevalence studies show that antibody is
  present in most by age 3y

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ROTAVIRAL DISEASE BURDENWorldwide
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WORLDWIDE DISTRIBUTION OF ROTAVIRUS(Source-
Centers for Disease Control and Prevention)
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DISEASE BURDENUSA

• 20-40 deaths/year
• Cases with dehydration 1-2.5
• 50,000-70,000 hospitalizations/year
• Physician visits 500,000/year
• 2.7- 3.5 million affected each year
• Economic impact

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ROTAVIRAL DISEASE BURDEN- U.S.A.Source- MMWR
August 11, 2006 / 55(RR12)1-13
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EPIDEMIOLOGY

• Age- children 4mo - 2 years are most
  affectedProtection of younger infants through
  through transplacental antibody transfer
• Asymptomatic infections are common, especially in
  adults
• Nosocomial infections
• Outbreaks
• Severe Disease young, immunocompromised

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Epidemiology (contd.)

• Seasonality
• Winter months (Nov. through May in US)
• Gradual spread W to E
• Year-round in the tropics
• Incubation period - thought to be lt4 days

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EPIDEMIOLOGY(Source- Centers for Disease Control
and Prevention)
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SEASONAL SPREAD - U.S.(Source- CDC)
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TRANSMISSION

• Mainly person to person via fecal-oral route
• Food and water-borne spread is possible
• Fomites
• Spread via respiratory route is speculated

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EPIDEMIOLOGY - 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 10-100 pfu

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EPIDEMIOLOGYDifferences 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

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Rotavirus Immunity

• Antibody against VP7 and VP4 partially protective
• First infection usually does not lead to
  permanent immunity
• Reinfection can occur at any age
• Subsequent infections generally less severe

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CLINICAL CASE

• A 22 month old female is admitted to the
  pediatric ward for cough and fever up to 103F.
• Chest X ray shows left lower lobe pneumonia.
• She is being treated with intravenous Ceftriaxone
  and her fever is gradually improving.
• On hospital day 5, she develops diarrhea with 4
  watery stools and a fever of 102F.
• Stool studies showed no traces of blood and no
  fecal leukocytes.
• Further studies are pending.

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CLINICAL FEATURES

• Incubation period - thought to be lt4 days
• Fever- can be high grade (gt102?F in 30)
• Vomiting, nausea precede diarrhea
• Diarrhea
• usually watery (no blood or leukocytes)
• lasts 3-9 days
• longer in malnourished and immune deficient
  individuals.
• NEC and hemorrhagic GE seen in neonates

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MECHANISM OF DIARRHEA

• Watery diarrhea due to net secretion of
  intestinal fluid and loss of absorptive surface
• Activation of the enteric nervous system
• Role of NSP4 peptide regions as an enterotoxin

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CLINICAL FEATURES (contd.)

• Dehydration is the main contributor to mortality
• Secondary malabsorption of lactose and fat, and
  chronic diarrhea are possible

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Clinical appearance of dehydrationPhoto Credit
Dr. D. Mahalanabis, World Health Organization
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DIAGNOSIS

• Antigen detection in stool
• ELISA, LA (Group A rotavirus),
  immunochromatographic assay
• EM- non-Group A viruses also
• Culture- Group A rotaviruses can be cultured in
  monkey kidney cells
• Serology for epidemiologic studies

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TREATMENT AND PREVENTION

• Treatment
• Supportive- oral, IV rehydration
• Prevention
• Hand hygiene and disinfection of surfaces
• Vaccine

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Rotavirus Vaccine (Rota)

• Created by genetic reassortment
• Causes nonhuman rotavirus strains to express
  human rotavirus
• antigens on their surface
• Nonhuman rotaviruses have low pathogenicity for
  humans
• Replicate but do not cause disease

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ROTAVIRAL VACCINE Historical lessonsPhoto
source- PATH vaccine resource library

• 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,
  especially the first dose
• Approx. 15 cases/1.5 million doses given
• Attributable risk 110,000 (1st dose)

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RotaTeq (Merck)

• Live oral vaccine licensed 2006 in US
• Contains 5 reassortants (WC3 bovine strain with
  viral surface proteins of human serotypes G1-4
  and P1A)
• Contains no preservatives or thimerosal
• 3-dose schedule age 2,4,6 month
• Minimum age of first doses is 6 weeks
• First dose should be administered between 6 and
  12 weeks of age (until age 13 weeks)
• Do not initiate series after 12 weeks of age

MMWR 200655(RR-12)1-13.
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Rotavirus Efficacy and Safety Trial (REST)

• 70,000 enrollees in 11 countries
• 34,000 enrolled in the US
• Conducted from 2001 to 2004
• Double-blind (with sponsor blinding),
  placebo-controlled, randomized
• Data published in Jan. 2006
• Vesikari T, Matson DO, Dennehy P, et al. Safety
  and efficacy of a pentavalent human-bovine (WC3)
  reassortant rotavirus vaccine. N Engl J Med
  200635423--33.

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Efficacy of RotaTeq

• 98 reduction in severe RV GE within first year
  of vaccination
• 96 reduction in hospitalization
• 74 reduction of RV GE (any grade of severity)
  within first year of vaccination
• 71 reduction of RV GE in first 2 years of
  vaccination

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Incidence of intussuception-RotaTeq New Eng J
Med 2006 35423-33
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Rotarix?

• Live attenuated monovalent vaccine
• Manufactured by Avant Immunotherapeutics
• Contains the G1P8 human rotavirus strain
• Studied in Latin America in large trials
• 2 dose schedule
• Safety established- intussusception not
  increased, efficacious
• Licensed in other countries

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GASTROENTERITIS DUE TO ENTERIC ADENOVIRUS
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Adenovirus Gastroenteritis

• Isolated 1953 by Rowe from adenoidal tissue
• Linear double-stranded DNA
• 70-75nm in diameter
• Genus Mammalian AV (mastadenoviruses)
• 6 human subgenera (A-F)
• 51 human serotypes (1-51)
• Known oncogenic potential of some serotypes
• Commonly used as vectors in
• gene therapy and vaccine delivery

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Adenoviral core/capsid proteins
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Adenovirus Gastroenteritis

• Types 40, 41
• Belong to serogroup F
• Isolation requires special media-Graham 29
• ELISA for rapid detection is available

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Diarrhea due to Enteric Adenovirus

• Age lt4 years
• Ubiquitous
• Year round
• Spread via fecal-oral route

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CLINICAL FEATURES- Adenovirus gastroenteritis

• Incubation period 3 -10 days
• Diarrhea lasts for 10 -14 days
• Can also cause intussusception, mesenteric
  adenitis, appendicitis
• Treatment with cidofovir promising for severe
  disease in immunocompromised

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HUMAN CALICIVIRUSES
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HUMAN CALICIVIRUSES(HuCV)

• Family Caliciviridae
• Non-enveloped RNA viruses
• with ss sense RNA
• 27-35 nm in size
• Contains single capsid protein

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Calicivirus

• Norwalk virus and Norwalk-like
• Sapporo-like viruses
• Vesivirus
• Lagovirus

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CLASSIFICATION 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

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MORPHOLOGY- typical

• 32 cup-like depressions
• EM appearance of Star of David
• 31-35 nm size
• E.g.- Sapporo-like viruses

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HUMAN CALICIVIRUSES - SLV
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Morphology of HuCV- atypical

• Smaller size- approx. 27 nm
• Rough, feathery surface but no internal pattern
• Small Round Structured viruses
• E.g.- Norwalk-like viruses

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NOROVIRUS

• Photo source- http//www.kyobiken.or.jp/norovirus.
  JPG

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Norwalk virus

• winter vomiting disease 1968, Norwalk OH
• Cause 40 of nonbacterial epidemics
• 45 foodborne, 52 shell fish associated
  outbreaks
• Explosive epidemics
• camps, cruise ships, nursing homes
• Food borne illness
• raw shellfish

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EPIDEMIOLOGY - Noroviruses

• Worldwide distribution
• gt23 million cases/year in the U.S.
• Major cause of food-borne outbreaks of GE (gt50)
• Most people have had infections by age 4 years
  (by sero-prevalence studies)

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Norwalk virus Clinical Features

• 24 hour (range 12-96) incubation period
• Vomiting prominent
• Headache, myalgia, fever
• Diarrhea 1-3 days, less severe than rotavirus
• Affects all ages
• Treatment symptomatic
• rehydration, antidiarrheals
• Complications rare
• immunocompromised

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SPREAD

• Person-to-person fecal-oral spread
  (stool/vomitus)
• Fecal contamination of food or water
• Fomites (stool/vomitus)
• Ingestion of aerosolized particles

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SPREAD OF NOROVIRUS

• A study has shown that where fingers come into
  contact with virus-contaminated material,
  Norovirus can be consistently transferred via the
  fingers to melamine surfaces and from there to
  other typical hand-contact surfaces, such as
  taps, door handles and telephone receivers..It
  was found that contaminated fingers could,
  sequentially transfer virus to up to seven clean
  surfaces. Norovirus can survive for up to
  several days in the environment and still cause
  infection.
• ENVIRON. HEALTH JOURNAL March 2005, pages 22-24

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SPREAD- Viability of Caliciviruses

• Survive on surfaces for several days
• Survive in water chlorinated at routine levels
  (up to 10 ppm)
• Survive freezing, heating up to 60C
• Evidently survive in steamed shellfish

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EPIDEMIOLOGY

• Asymptomatic infections- seroconversion but
• asymptomatic shedding of virus
• Low infective dose (10 pfu)
• Viral excretion during convalesence ( 2 wk.)
• Protective immunity short-lived
• NLV cross protection?
• Blood group antigens predilection?

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EPIDEMIOLOGY Outbreaks

• Cruise ships, schools, nursing homes,
• Can involve infants and school-age children
• Source usually is contaminated food and water
  (E.g. seafood-oyster and shellfish, salads, cake
  icing, raw fruit etc.)
• Rapid secondary spread

• Photo source- http//www.seattlest.com

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DIAGNOSIS

• Specimen- stool, vomitus, food, environmental
  swabs (during outbreak investigations)
• RT-PCR in state public health labs.
• Serology for epidemiologic purposes
• Immune EM is less used

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Control of spread of CV GE

• Unclear as to which is the best method of
  disinfection of surfaces
• CDC recommends environmental disinfection of
  non-porous surfaces with 1000 ppm bleach solution
  (1 part bleach 50 parts water)

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HUMAN ASTROVIRUS
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ASTROVIRUS

• Described in relation to an outbreak of
  gastroenteritis in 1975
• Detected by EM
• Immunologically distinct from Hu CV
• Animal strains are known

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Classification of Astrovirus

• Genus Astrovirus
• Family Astroviridae
• Human serotypes HuAstV 1-8

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ASTROVIRUS- 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
• Genome has been sequenced

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ASTROVIRUS- STRUCTUREPhoto source-
http//www.ncbi.nlm.nih.gov/ICTVdb/Images/Cornelia
/astro3.htm
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ASTROVIRUS - Clinical Features

• Infants and children are most often affected
• Elderly and immune compromised persons also
• Short incubation period 1-4 days
• Nausea, vomiting, abdominal cramping and watery
  diarrhea
• Constitutional symptoms-fever, malaise, headache

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ASTROVIRUS - Epidemiology

• Endemic worldwide
• True disease burden is not yet clear
• Mainly in children lt7 years of age
• Transmission- person-to-person fecal-oral
• Outbreaks due to fecal contamination of
• sea-food or water

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Outbreaks of AstrovirusSource-
http//www.ozestuaries.org/indicators/shellfish_cl
osures.jsp
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ASTROVIRUS - Diagnosis

• EM (virus shed in stool in great numbers)
• EIA
• RT-PCR

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Toroviruses

• Family Coronaviridae
• Genus Torovirus
• Human and animal pathogen
• Pleomorphic, coated ss () RNA virus
• Core doughnut-shaped (torus)
• Watery diarrhea in 2 12 months old
• Diagnosis EM

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Cytomegalovirus

• Herpesvirus
• Immune competent
• Mononucleosis-type syndrome
• Immunocompromised
• retinitis
• hepatitis
• gastrointestinal involvement (colon)

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Cytomegalovirus colitis
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Request Stool for viruses

• Hospital 1 Enterovirus, adenovirus,
  herpesviruses (CMV, HSV)
• Hospital 2 CMV
• Hospital 3 Enterovirus
• Hospital 4 Stop sending us this!
• Hospital 5 Please resubmit in viral cx
  medium! gt HSV, CMV