Laboratory Methods for Assessing and Licensing Influenza Vaccines for Poultry.

Avian influenza (AI) vaccines for poultry are based on hemagglutinin (HA) proteins, and protection is specific to the subtype. An estimated 313 billion doses have been used between 2002 and 2018 for high pathogenicity AI control. No universal vaccines are currently available. The majority of AI vaccines are inactivated whole influenza viruses that are grown in embryonating chicken eggs, emulsified in oil adjuvant systems, and injected subcutaneously or intramuscularly. Live virus-vectored vaccines such as recombinant viruses of fowl pox, Newcastle disease, and herpesvirus of turkeys containing inserts of AI virus HA genes have been used on a more limited basis. Also, vaccines have been licensed or registered based on baculovirus and defective replicating alphavirus (RNA particles) expressing HA protein or DNA vaccine with HA gene insert. In studies to evaluate vaccine efficacy and potency, the protocol design and its implementation should address the biosafety level needed for the work, provide information required for approval by Institutional Biosafety and Animal Care Committees, contain information on seed strain selection, provide needed information on animal subjects and their relevant parameters, and address the selection and use of challenge viruses. Various metrics have been used to directly measure vaccine-induced protection, including prevention of death, clinical signs, and lesions; prevention of decreases in egg production and alterations in egg quality; quantification of the reduction in virus replication and shedding from the respiratory tract and gastrointestinal tracts; and prevention of contact transmission in in vivo poultry experiments. In addition, indirect measures of vaccine potency and protection have been developed and validated against the direct measures and include serological assays in vaccinated poultry and the assessment of the content of HA antigen in the vaccine. These indirect assessments of protection are useful in determining if vaccine batches have a consistent ability to protect. For adequate potency, vaccines should contain 50 mean protective doses of antigen per dose, which corresponds to 0.3-7.8 μg of HA protein in inactivated vaccines, depending on immunogenicity and antigenic relatedness of individual seed strains.