総会抄録

<p>Antibody responses and protection of offspring when mothers were immunized via the IN route and their offspring via the IN route<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0157041#t004fn001" target="_blank">^a</a>.</p

Virus elution in <i>vitro</i>.

<p>50 µl two-fold dilutions of virus containing the HA titers of 1∶128 was incubated with 50 µl 0.5% chicken erythrocytes in microtiter plates at 4°C for 1 h. Then microtiter plates were incubated at 37°C, and the reduction of HA titers was measured periodically for 8 h.</p

Impact of different NA on initiation of influenza virus infection.

<p>MDCK cells were infected at an MOI of 0.001 in the presence of 1 µg/ml TPCK-trypsin. After adsorption for 1 h at 37°C, the inocula were removed and the cultures were washed 3 times. The cells were incubated at 37°C for 6 h. At the indicated time, the cells were processed for immunofluorescence, and the infected cells were detected with polyclonal antisera to whole viruses. (A) Fluorescence images of the infected cells at 6 h p.i. Fluorescent photomicrographs showing the intracellular expression of virus protein in cell culture. The FITC-fluorescence signal was expressed as the infected cells. (B) Volocity Demo software analysis of the ratios of infected cells according to the Fig. 5A. *, statistical significance (<i>p</i><0.05) (C) Flow-cytometric analysis of virus-infected cells at 6 h p.i. MDCK cells (2×10⁶) in suspension were incubated with PBS or anti-PR8 antibodies on ice. Then the FITC-conjugated IgG secondary antibodies were added. After washing, the cells were fixed and the number of infected cells was determined by flow cytometric analysis. *, statistical significance (<i>p</i><0.05).</p

Influenza virus induced cell-cell fusion. MDCK cells were infected with the viruses at MOI of 0.1 or 0.001 in the presence of 1 µg/ml TPCK-trypsin.

<p>After adsorption for 1 h at 37°C, the inocula were removed and the cultures were washed 3 times. The cells were incubated for the indicated times at 37°C in the maintenance media. At the indicated time, the cells were processed for indirect immunofluorescence assay, and the infected cells were detected with polyclonal antisera to whole viruses. (A) MOI at 0.1, 3 h p.i. (B) MOI at 0.1, 6 h p.i. (C) MOI at 0.001, 12 h p.i.</p

Pathogenicity of recombinant viruses in BALB/c mice.

<p>Survival rates (A) and bodyweight changes (B) after challenge with the viruses. BALB/c mice were intranasally inoculated with rPR8-H5N1NA, rPR8-H9N2NA, rPR8-H1N1NA or PR8-wt virus at 1×10^6.5 EID₅₀. The survival rates and bodyweights of five mice in each group were measured daily from the date of challenge to 14 days after challenge. Values represent mean ± SD of each group of mice.</p

Alignment of the deduced amino acid sequences of the NA genes from the influenza virus strains A/Chicken/Jiangsu/7/2002(H9N2), A/California/04/2009 (H1N1), A/chicken/Henan/12/2004 (H5N1) and PR8 (H1N1).

<p>Alignment of the deduced amino acid sequences of the NA genes from the influenza virus strains A/Chicken/Jiangsu/7/2002(H9N2), A/California/04/2009 (H1N1), A/chicken/Henan/12/2004 (H5N1) and PR8 (H1N1).</p

Replication of recombinant viruses in mice^a.

a<p>BALB/c mice were intranasally inoculated with recombinant or wild-type virus at 1×10^6.5 EID₅₀. On days 3 and 6 after infection, five mice from each group were killed for virus titration. Results are expressed as means ± SD.</p>b<p>ND, Not done.</p

Virus strains generated by reverse genetics and the amino acids comparison of NA^a.

a<p>The amino acids homology of wild-type PR8 virus and those of the H5N1, H9N2 or swine-H1N1.</p

Genotype Diversity of H9N2 Viruses Isolated from Wild Birds and Chickens in Hunan Province, China

<div><p>Three H9N2 avian influenza viruses were isolated from the Dongting Lake wetland, among which one was from fresh egret feces, the other two were from chicken cloacal swabs in poultry markets. Phylogenetic analyses suggested that eight genes of the egret-derived H9N2 virus might come from Korean-like or American-like lineages. The two poultry-derived H9N2 viruses were reassortants between the CK/BJ/94-like and G1-like viruses. Except the PB1 genes (90.6%), the nucleotide sequence of other internal genes of the two viruses exhibited high homology (>95%). In addition, they also exhibited high homology (96–98.3%) with some genes of the H7N9 virus that caused an epidemic in China in 2013. Nucleotide sequence of the poultry-derived and egret-derived H9N2 viruses shared low homology. Infection studies showed that the egret-derived H9N2 virus was non-pathogenic to both mice and chickens, and the virus was unable to infect chickens even through 8 passages continuously in the lung. On the other hand, the chickens infected by poultry-derived viruses showed obvious clinical symptoms and even died; the infected mice showed no noticeable clinical symptoms and weight loss, but viruses could be detected in their lungs. In conclusion, for the egret-derived H9N2 virus, it would take a long adaptation process to achieve cross-species transmission in poultry and mammals. H9N2 viruses isolated at different times from the same host species in the same geographical region presented different evolutionary status, and virus isolated from different hosts in the same geographical region exhibited genetic diversity. Therefore, it is important to continue the H9N2 virus surveillance for understanding their evolutionary trends so as to provide guidance for disease control and prevention.</p></div

Molecular characterizations of HA, NA, PB2, PB1, PA, NP, M and NS at representative sites.

a<p>E1: A/Egret/Hunan/1/2012; C1: A/Chicken/Hunan/1/2012; C12: A/Chicken/Hunan/12/2011.</p><p>-: There was no deletion. +: There was deletion.</p