Preparation of a standardized, efficacious agricultural H5N3 vaccine by reverse genetics

Abstract

Options for the control of emerging and reemerging H5N1 influenza viruses include improvements in biosecurity and the use of inactivated vaccines. Commercially available H5N2 influenza vaccine prevents disease signs and reduces virus load but does not completely prevent virus shedding after challenge with H5N1 virus. By using reverse genetics, we prepared an H5N3 vaccine whose hemagglutinin is 99.6% homologous to that of A/CK/HK/86.3/02 (H5N1). We used the internal genes of A/PR/8/34 and the H5 of A/Goose/HK/437.4/99 (H5N1) after deletion of basic amino acids from its connecting peptide region. The resulting virus was not lethal to chicken embryos and grew to high HA titers in eggs, allowing preparation of HA protein-standardized vaccine in unconcentrated allantoic fluid. The N3 neuraminidase, derived from A/Duck/Germany/1215/73 (H2N3), permitted discrimination between vaccinated and naturally infected birds. The virus construct failed to replicate in quail and chickens. Similar to parental A/PR/8/34 (H1N1), it replicated in mice and ferrets and spread to the brains of mice; therefore, it should not be used as a live-attenuated vaccine. The H5N3 vaccine, at doses of 1.2 microg HA, induced HI antibodies in chickens and prevented death, signs of disease, and markedly reduced virus shedding after challenge with A/CK/HK/86.3/02 (H5N1) but did not provide sterilizing immunity. Thus, reverse genetics allows the inexpensive preparation of standardized, efficacious H5N3 poultry vaccines that may also reduce the reemergence of H5N1 genotypes.

Inactivated North American and European H5N2 avian influenza virus vaccines protect chickens from Asian H5N1 high pathogenicity avian influenza virus

“…Other vaccines, including traditional inactivated North American AI vaccine strains, H5N1 reverse genetic AI vaccine strain and fowlpox recombinant vaccines with H5 AI gene inserts, have been shown to protect against diverse Asian H5N1 HPAI strains in the laboratory (Swayne et al ., 2001;Liu et al ., 2003;Qiao et al ., 2003;Swayne, 2004;Swayne & Beck, 2005). This indicates that the need for frequent change in H5 AI vaccine strains for poultry, as occurs with human influenza A subtype H1 and H3 and influenza B vaccine strains, because of rapid drift, is not necessary.…”

Microbial adaptation and change: avian influenza

“…This technique provides a powerful new tool for the generation of vaccines for both poultry and humans. A recent study showed the efficacy of a reverse-genetic-derived H5N3 vaccine for agricultural use (25). The H5N3 vaccine was developed as follows: the HA gene came from A/goose/HK/437-4/99, which was altered by the deletion of basic amino acids from the connecting peptide region.…”

“…3) (25). The vaccine induced antibodies in the vaccinated birds, prevented death and clinical signs of disease and reduced virus shedding after challenge (25). There are several advantages in using reverse genetics to generate vaccines for poultry, as follows: -rapid and easy development of vaccines -uniform vaccination -low cost -safety and effectiveness.…”

Immunogenicity of the inactivated oil emulsion influenza A (H5N1) vaccine in chickens

“…According to Liu M et al (2003) the inactivated vaccine (Intervet) based on the heterogeneous strain A/CK/Mexico/232/94 (H5N2) formed 12-month-long immunity in twice vaccinated chickens. The geometric mean antibody titer was 1:100 on the 28 th day after the first vaccination and 1:333 after the second immunization.…”