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AbstractThe haemagglutinin (HA) gene of H5N1 Avian influenza virus(AIV) was amplified from the plasmid pGEM-HA and cloned into the baculovirus transfer plasmid pFastBacHT to construct the recombinant transfer vector pFastBacHT-HA. The pFastBacHT-HA was transformed into DH10Bac competent cells, transposed with a shuttle vector (Bacmid) and the transposition rBacmid-HA was constructed. The recombinant baculovirus was harvested from sf9 cells transfected with rBacmid-HA. The expressed HA protein was identified and analysed by SDS?PAGE, Western blot and haemadsorption assay. The 66 kDa protein could only react with chicken serum positive to H5 subtype AIV. The haemadsorption assay showed that the sf9 cells infected with rBacmid-HA baculovirus could absorb chicken red blood cells. These results indicated that the HA protein was successfully expressed in sf9 cells, with reaction specificity to H5 subtype antiserum. [ABSTRACT FROM AUTHOR]

Avian influenza virus is acquiring the ability to cross the species barrier between birds and mammals. However, the genetic mechanism of this ability is not well known. We successfully rescued an influenza A virus A/Chicken/Guangdong/03 (H5N1), which is highly pathogenic to both SPF chickens and BALB/c mice, by plasmid-based reverse genetics. The rescued virus, R-A/Chicken/Guangdong/03 (R-CG), and the wild-type A/Chicken/Guangdong/03 (W-CG) were found to share similar biological properties, such as in titres of 50% egg infectious dose (EID50), 50% tissue culture infectious dose (TCID50) and intravenous pathogenicity index (IPVI). R-CG, like W-CG, is highly pathogenic in mice following natural route infection. Both R-CG and W-CG viruses can be isolated from many organs of mice such as brain, lung, kidney and spleen. As a result, the constructed R-CG reverse genetic system can be used as a tool in the investigation of the molecular mechanism of infection of the mammalian host by avian influenza virus. [ABSTRACT FROM AUTHOR]

To evaluate the effects of H5 subtype avian influenza DNA vaccine with HA gene and expressive vector optimization, groups of 3-week-old specific pathogen free (SPF) chickens were inoculated intramuscularly with a single dose of 100 μg or 10 μg of pCIHA5, pCAGGHA5, pCIoptiHA5 or pCAGGoptiHA5 in 200 μl volume. Another group of chickens was injected with 200 μl phosphate-buffered saline (PBS) as controls. Sera were collected every week after vaccination to detect the HI and agar-gel precipitin (AGP) antibodies. Four weeks after the single vaccination, all chickens were challenged with 100LD50 of the highly pathogenic A/Goose/GuangDong/1/96(H5N1) [Gs/GD/1/96(H5N1)]. Oropharyngeal and cloacal swab specimens were collected from all chickens 3, 5 and 7 days after inoculation for titration of virus in eggs. Chickens were observed daily for 2 weeks for signs of disease and death. Results showed that in the groups vaccinated with 100 μg pCAGGoptiHA5 and pCAGGHA5 chickens were completely protected from virus challenge (no disease signs, no virus shedding and no deaths), while only partial protection occurred in chickens vaccinated with 100 μg pCIoptiHA5 (75%) and pCIHA5 (50%). In groups vaccinated with 10 μg pCAGGoptiHA5 and pCAGGHA5 chickens were protected from virus challenge (no disease signs and no deaths). In the group vaccinated with 10 μg pCIoptiHA5 the protection ratio was 75%, while chickens vaccinated with 10 μg pCIHA5 all died after challenge. Results indicated that codon usage optimization of the HA gene and β-actin promoter enhanced the protection efficiency of H5 subtype avian influenza DNA vaccine and the construction pCAGGoptiHA5 could protect chickens from lethal H5N1 virus challenge even at the low dose of 10 μg, implying the possibility for the commercialization of an avian influenza DNA vaccine in the near future. [ABSTRACT FROM AUTHOR]

The article reports on the findings of the literature review on the study of bird influenza. The study revealed that poultry trade is mainly responsible for the spread of bird flu instead of wild birds. Based on the study, wild birds serve as reservoirs of gene fragments of low pathogenic avian influenza, which can be transmitted to domestic birds. It was identified that high density rearing of domestic poultry was the ideal condition for the evolution of a virulent, infectious strain of the disease, which is disseminated through trade.