Avian Influenza

1. Avian Influenza Saad GharaibehDVM, PhD, Dip ACPV Dept. of Pathology and Animal Health Faculty of Veterinary Medicine Jordan University of Science and Technology Irbid 22110, Jordan saadgh@just.edu.jo 02/720-1000 ext 22059

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

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

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

5. HI test ELISA test NI test AGP test

6. 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

7. Infectious Virus • Needs HA0 cleaved into HA1 & HA2 • Intracytoplasmic: • Furin-like enzyme (ubiquitous proteases): HP • Trypsine-like enzyme: All AIV

8. 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.

9. 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

10. Criteria for HP • 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 1:10 dilution of a bacteria free, infectious allantoic fluid. • H5 or H7 has amino acid sequence at the hemagglutinin cleavage site compatible with HPAIV • Non-H5 or H7 that kills 1-5 chickens and grows in cell culture w/o added trypsin

11. 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)

12. Gross Lesions (Low pathogenic AI disease) • Catarrhal rhinitis / tracheitis • Ocular discharge • Airsaculitis • Ovarian involution and hemorrhage • Yolk peritonitis • Swollen kidney and urates

15. Signalment & Clinical Signs(Highly pathogenic AI disease) • Sudden onset of high mortality (up to 100%) • Depression • +/- nervous signs

19. Gross Lesions(Highly pathogenic AI disease) • Edematous to necrotic comb and wattles • Edema, necrosis, and hemorrhages in different organs

22. 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

23. RT-PCR • Testing can be performed in one day for multiple agents. • Sensitivity is very high and comparable to virus isolation. • Can be applied on samples from any species. • Decrease the chance of contamination with live virus.

24. RT-PCR Diagnostics in JUSTSafety Considerations

25. RT-PCR Diagnostics in JUSTChicken Respiratory Disease viruses APV IB ND AI MWM

26. Serologic Testing and Surveillance • AGPT: Type specific (available at JUST) • ELISA: Type specific (available at JUST) • HI: Subtype specific (available at JUST) • NI: Subtype specific • Antigen capture ELISA (available at JUST) • RT-PCR: Surveillance and diagnosis (available at JUST)

27. Agar Gel Precipitation

28. ELISA Readings Positive Flock Negative Flock

29. Hemagglutination Inhibition

30. 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

33. Drastic measures in some Asian countries www.animalactivism.org/ documents

37. AI crossed Species Barrier into Humans A/chicken/Hong Kong/220/97 (H5N1)

38. What conditions favor AI spread?

39. Densely populated countries

41. Very popular Live-bird markets

43. Free Range Poultry

44. Is H5N1 AI bad for economies? Sources: WHO, Institute of Medicine, FAO, OIE, Asian Development Bank, World Bank

46. High Risk Areas in Jordan

47. Should we start vaccinating poultry against H5N1?

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

49. Swayne et al., 2000, Veterinary Microbiology

50. 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.