Avian Influenza

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Avian Influenza

• Saad Gharaibeh DVM, PhD, Dip ACPV
• Dept. of Pathology and Animal Health
• Faculty of Veterinary Medicine
• Jordan University of Science and Technology
• Irbid 22110, Jordan
• saadgh_at_just.edu.jo
• 02/720-1000 ext 22059

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Avian InfluenzaHistory

1. 1878 Fowl plaque was described (Italy)
2. 1901 Fowl plaque is caused by a virus
3. 1955 It is type A influenza virus
4. 1970 AGP test introduced
5. 1972 Waterfowl is a reservoir
6. 1979 Virulence and hemagglutinin cleavability was
   established
7. 1997 Direct transmission of H5 AIV from birds to
   humans

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Avian Influenza Virus

1. Orthomyxoviridae
2. Pleomorphic RNA viruses, single stranded,
   negative sense genome.
3. Has glycoprotein projections HA, NA
4. Three antigenic types A, B, C (Avian influenzas
   are all type A)
5. 8 gene segments code for 10 proteins
6. Vary in pathogenicity

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NI test
HI test
AGP test
ELISA test
Jong et al., 2000, Journal of Infection
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HI test
ELISA test
NI test
AGP test
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Nomenclature

• A/chicken/Hong Kong/220/97 (H5N1)
• A Type of virus A, B, C
• Chicken Host of origin
• Hong Kong Geographic origin
• 220 Strain Number (Case number)
• 97 Year of isolation
• (H5N1) H N subtype

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Infectious Virus

• Needs HA0 cleaved into HA1 HA2
• Intracytoplasmic
• Furin-like enzyme (ubiquitous proteases) HP
• Trypsine-like enzyme All AIV

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Cell Types for Replication

• All AIV (trypsine-like enzymes)
• Respiratory epithelium
• GI epithelium
• HP AIV (Furin-like enzymes) Variety of cells
  resulting in a systemic infection.

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Approaches Used to Characterize AIV Pathogenicity

• In vivo methods
• Laboratory Inoculation of chickens
• Chicken embryo lethality
• In vitro methods (evaluation of HA cleavability)
• Plaques or CPE assays (CEF does not have trypsin)
• Direct detection of cleaved HA
• Nucleotide sequence of HA cleavage site
• Direct measure of pathogenicity potential
• Identify pathogenicity increases during virus
  passage in chickens under controlled conditions
• Virulence in mice and ability to infect other
  mammals

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Criteria for HP

1. AIV lethal for 6,7, or 8 / 8 four-to-six-week-old
   susceptible chickens within 10 days following IV
   inoculation with 0.2 ml of 110 dilution of a
   bacteria free, infectious allantoic fluid.
2. H5 or H7 has amino acid sequence at the
   hemagglutinin cleavage site compatible with HPAIV
3. Non-H5 or H7 that kills 1-5 chickens and grows in
   cell culture w/o added trypsin

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Signalment Clinical Signs(Low pathogenic AI
disease)

• Respiratory signs
• Diarrhea
• Drop in egg production 7-10 days 5-30
• Mild increase in Mortality (2o bacterial
  infection will increase mortality)

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Gross Lesions (Low pathogenic AI disease)

1. Catarrhal rhinitis / tracheitis
2. Ocular discharge
3. Airsaculitis
4. Ovarian involution and hemorrhage
5. Yolk peritonitis
6. Swollen kidney and urates

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Signalment Clinical Signs(Highly pathogenic AI
disease)

• Sudden onset of high mortality (up to 100)
• Depression
• /- nervous signs

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Gross Lesions (Highly pathogenic AI disease)

1. Edematous to necrotic comb and wattles
2. Edema, necrosis, and hemorrhages in different
   organs

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High Path Avian Influenza Diagnosis

• Clinical features in commercial poultry give a
  tentative diagnostic
• Sudden death and high mortality rate
• RT-PCR and sequencing
• Virus isolation and identification is the gold
  standard but very few laboratories in the world
  can handle such a virus capable of infecting
  humans.
• Commercial antigen capture ELISA (lack
  sensitivity and will cross react with other
  endemic subtypes)
• Serology AGP, ELISA, HI, NI

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RT-PCR

1. Testing can be performed in one day for multiple
   agents.
2. Sensitivity is very high and comparable to virus
   isolation.
3. Can be applied on samples from any species.
4. Decrease the chance of contamination with live
   virus.

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RT-PCR Diagnostics in JUSTSafety Considerations
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RT-PCR Diagnostics in JUSTChicken Respiratory
Disease viruses
APV
IB
ND
AI
MWM
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Serologic Testing and Surveillance

1. AGPT Type specific (available at JUST)
2. ELISA Type specific (available at JUST)
3. HI Subtype specific (available at JUST)
4. NI Subtype specific
5. Antigen capture ELISA (available at JUST)
6. RT-PCR Surveillance and diagnosis (available at
   JUST)

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Agar Gel Precipitation
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ELISA Readings
Positive Flock
Negative Flock
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Hemagglutination Inhibition
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Control Prevention

• Biosecurity
• Stamping out infected flocks
• Vaccination of flocks at high risk
• Killed vaccines
• Viral vector vaccines
• Live attenuated vaccines are not licensed for
  poultry

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Drastic measures in some Asian countries
www.animalactivism.org/ documents
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AI crossed Species Barrier into
Humans A/chicken/Hong Kong/220/97 (H5N1)
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What conditions favor AI spread?
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Densely populated countries
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Very popular Live-bird markets
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Free Range Poultry
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Is H5N1 AI bad for economies?
Disease Country Year Impact Cost (US)
mad cow UK 1990-98 Beef export 9 billion
Cholera Peru 1991 Seafood export 770 million
Plague India 1995 Tourism, trade 2 billion
H5N1 flu Hong Kong 1997 Loss of poultry (1.5 million birds) 22 million
Cholera Tanzania 1998 Seafood export 36 million
Nipah Malaysia 1999 Loss of swine (0.9 million pigs) 540 million
West Nile US 1999 400 million
SARS Worldwide 2003 Tourism, trade 80 billion
H5N1 flu SE Asia 2003-05 Loss of poultry (140 million birds so far) 10 billion? (and growing)
Sources WHO, Institute of Medicine, FAO, OIE,
Asian Development Bank, World Bank
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High Risk Areas in Jordan
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Should we start vaccinating poultry against H5N1?
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Killed Vaccines

1. Will result in only humoral antibody response
   against all viral proteins except NS1.
2. Will significantly reduce shedding of the
   challenge virus.
3. Will interfere with AGP, ELISA, HI, and NI if
   (homologous).
4. If sequence of the HA gene is identical to the
   challenge virus it may eliminate shedding
   completely.

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Swayne et al., 2000, Veterinary Microbiology
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Advantages of Vaccines

• Reduces the number of chickens from which AI
  challenge virus could be reisolated.
• Decreased the titers of virus detected in the
  cloaca and oropharynx (up to 99.99)
• Reduced environmental contamination and prevented
  subsequent bird to bird transmission.
• The use of killed H5N2 vaccine in the face of
  HPAI H5N1 virus challenge was able to protect
  chickens from disease and interrupt virus
  transmission.

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DIVADifferentiating Infected from Vaccinated
Individuals

1. The use of killed vaccine and unvaccinated
   sentinels leaving 0.5-1 of the flock
   unvaccinated and marked (wing band) and these
   individuals will be subjected to serological
   monitoring.
2. Heterogonous killed vaccine Screen for field
   infection using NI.
3. Measuring serological response to NS1 by ELISA or
   western blot.

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Vaccine / Industry / Politics

1. The use of vaccine to aid in the control of AI is
   a political issue and different people have a
   different say on this.
2. In some countries financial constrains preclude
   stamping out policy.
3. In some countries, export markets are not an
   issue to prevent vaccination.
4. In some countries, stamping out attempt may be
   unsuccessful.
5. With the ubiquitous nature of AI in wild birds
   it may be vaccination the most feasible tool to
   soften the sting of AI Beard 1981

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Vaccine / Industry / Politics

1. Field results have not shown vaccine to increase
   the risk of undetected infection in fact, field
   experience has shown that vaccination greatly
   enhances a control program. Halvorson, 2002,
   Avian Pathology
2. There is no way a vaccinated flock can be a
   greater threat to disease control than a
   non-vaccinated flock that breaks with AI.
   Halvorson, 2002, Avian Pathology
3. Epidemiological observations have shown that
   serologically positive birds are not associated
   with AI transmission. (Kradel, 1992)
4. Should the government set the rules when no
   indemnity (compensation) is paid?

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