Additional file 4: Fig. S4. of G45R mutation in the nonstructural protein 1 of A/Puerto Rico/8/1934 (H1N1) enhances viral replication independent of dsRNA-binding activity and type I interferon biology

Plaque assay of the virus inoculums. The virus inoculums (WT and G45R/NS1) at MOI of 0.01 were serially diluted before they were infected onto MDCK cells to confirm that each inoculum had equal amount of the virus. Plaque assay of the viruses at 10−5 dilution is shown. The average virus titers of both WT and G45R/NS1 inoculums are 5× 106 pfu/ml. (TIFF 1781 kb

Additional file 3: Fig. S3. of G45R mutation in the nonstructural protein 1 of A/Puerto Rico/8/1934 (H1N1) enhances viral replication independent of dsRNA-binding activity and type I interferon biology

Western blot analysis of STAT1, pSTAT1 and NS1 protein. A549 cells were infected with rX31 encoding WT and G45R/NS1 viruses at an MOI of 2. At 8 h post infection, cell lysates were clarified by centrifugation at 13,000 rpm for 15 min. The lysates were mixed in Laemmli sample buffer (Bio-Rad) and denatured at 95 °C for 5 mins prior to perform SDS-PAGE gel electrophoresis. Immunoblots were probed for STAT1 (purified rabbit anti-Stat1 N-terminus, BD Transduction Laboratories), pSTAT1 (purified mouse anti-Stat1 pY701 BD Transduction Laboratories), NS1 (polyclonal rabbit anti-NS1; Thermoscientific) and actin (purified mouse anti-actin Ab-5; BD Transduction Laboratories). Antibodies were detected by incubation with goat anti-rabbit (GE healthcare) or goat anti-mouse HRP-linked antibody (Cell signaling). The immunoblots were visualized by using ChemiDoc XRS imager (Bio-Rad). Band intensity of proteins was quantified by using Image Lab version 5.0 (Bio-Rad). (TIFF 2319 kb

Viola brevistipulata W. Becker

原著和名: オホバキスミレ科名: スミレ科 = Violaceae採集地: 新潟県 柏崎市 東長鳥 峠 (越後 柏崎市 東長鳥 峠)採集日: 1988/4/26採集者: 萩庭丈壽整理番号: JH037661国立科学博物館整理番号: TNS-VS-98766

PCA analysis of normalized protein expression values of subset A.

<p>Principle component analysis (PCA) was performed with normalized log₂–transformed protein expression values from nasal washes for 18 samples from the subset A that was selected based viral load being higher than > 2⁸. The first two principal components are shown representing 42% and 16%, respectively, of the total variation. Healthy controls are labelled gray and IAV-positive samples are labelled red. In addition, sample identities (e.g., ID_4002) are shown. Horizontal and vertical axis represent principle component 1 and 2, respectively.</p

PCA analysis of normalized protein expression values.

<p>Principle component analysis (PCA) was performed with quantile normalized log₂–transformed protein expression values from nasal washes for all 24 samples. The first two principal components are shown representing 24% and 17%, respectively, of the total variation. Healthy controls are labeled gray and IAV-positive samples (in which influenza A or B was detected by PCR) are labeled red. In addition, sample identities (e.g. ID_4043) are shown. Horizontal and vertical axis represent principle component 1 and 2, respectively.</p

Scatter plot of RPS6KA5 expression levels and viral load.

<p>Scatter plot of normalized log₂–transformed relative protein concentrations of RPS6KA5 which was the DEP that most strongly correlated with viral titers and log₂-transformed virus titers are presented for patients from subset B. Red line: linear regression model.</p

Functional analysis of DEPs from subset B.

<p>Pathway enrichment analysis for normalized log₂–transformed expression values of proteins that were differentially expressed in IAV-positive versus IAV-negative patients from subset B is presented. Pathway terms that were enriched for the DEPs from subset B are indicated on the y-axis, the number of DEPs in the respective pathway category is indicated on the x-axis. The p-value for the probability that the observed distribution of expression occurred by chance is represented by colors of bars. The cut-off value for the pathway p-values was chosen at 0.05.</p

Age−specific proportions of children sero-protected (post-vaccination titre cut point > = 40) by N seasons vaccinated.

*<p>Age-group as of November 1, 2007.</p

Age−specific proportions of children sero-protected (post-vaccination titre cut point > = 320) by N seasons vaccinated.

*<p>Age-group as of November 1, 2007.</p

Vaccine antigens by year.

a<p>A/New York/55/04 is antigenically equivalent to the A/California/7/04 (H3N2) virus strain; B/Jiangsu/10/03 is antigenically equivalent to the B/Shanghai/361/02 virus strain <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051498#pone.0051498-National2" target="_blank">[10]</a> and is of the B Yamagata lineage <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051498#pone.0051498-National1" target="_blank">[1]</a>.</p>b<p>A/Hiroshima/52/05 is antigenically equivalent to the A/Wisconsin/67/05 virus strain <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051498#pone.0051498-National3" target="_blank">[11]</a>.</p>c<p>B/Malaysia/2506/04 belongs to the B/Victoria/02/87 lineage <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0051498#pone.0051498-National3" target="_blank">[11]</a>.</p