Cross-strain influenza A(H1N1) antibody responses measured by HI assay.^*

<p>HI  =  hemagglutination inhibition; TIV  =  trivalent inactivated influenza vaccine. Note that undetectable titres <10 were assigned a value of 5.</p>*<p>Unit of analysis pooled sera from 5 mice, thus 10 pools from 50 mice were available for this experiment.</p>a<p>Measured two weeks after specified TIV dose;</p>b<p>Measured two months after specified TIV dose.</p>†<p>Compared to pre-immunization for initiating antigens; compared to immediately preceding titre for boosting antigens.</p><p>Brisbane  =  A/Brisbane/59/2007(H1N1)-like, component of the northern hemisphere 2008–09 and 2009–10 TIV.</p><p>California  =  A/California/07/2009(H1N1)-like, component of the northern hemisphere 2010–11 TIV.</p

Cross-lineage influenza B antibody responses measured by HI and MN assays.^*

<p>HI  =  hemagglutination inhibition; MN  =  microneutralization; TIV  =  trivalent inactivated influenza vaccine. Note that undetectable titres <10 were assigned a value of 5.</p>*<p>Unit of analysis pooled sera from 5 mice, thus 10 pools from 50 mice were available for this experiment.</p>a<p>Measured two weeks after specified TIV dose;</p>b<p>Measured two months after specified TIV dose.</p>†<p>Compared to pre-immunization for initiating antigens; compared to immediately preceding titre for boosting antigens.</p><p>Yamagata  =  Florida/4/06(Yamagata)-like, the Yamagata lineage antigen included in the northern hemisphere 2008–09 TIV.</p><p>Victoria  =  Brisbane/60/08(Victoria)-like, the Victoria lineage antigen included in the northern hemisphere 2009–10 and 2010–11 TIV.</p

Cross-strain influenza A(H3N2) antibody responses measured by HI assay.^*

<p>HI = hemagglutination inhibition; TIV  =  trivalent inactivated influenza vaccine. Note that undetectable titres <10 were assigned a value of 5.</p>*<p>Unit of analysis pooled sera from 5 mice, thus 10 pools from 50 mice were available for this experiment.</p>a<p>Measured two weeks after specified TIV dose;</p>b<p>Measured two months after specified TIV dose.</p>†<p>Compared to pre-immunization for priming antigens; compared to immediately preceding titre for boosting antigens.</p><p>Brisbane-like  =  A/Uruguay/716/2007(NYMC 175C)(H3N2) considered antigenically equivalent to the WHO recommended A/Brisbane/10/2007(H3N2) component of the 2008–09 northern hemisphere TIV.</p><p>Perth  =  A/Perth/16/2009(H3N2)-like, component of the northern hemisphere 2010–11 TIV.</p

Clinical outcomes including weight loss, nasal wash and lung virus titers by study group and day.

<p>Clinical outcomes are displayed including: (A) Mean percentage weight relative to baseline by study group and day, with standard errors. (B) Nasal wash virus titers by study group and day. (C) Lung homogenate virus titers at day 5 post-challenge. Box plots (B and C) display mean (dot) and median (line) virus titres as log pfu/mL. Per usual, the box extends to the 25^th/75^th percentiles and whiskers extend to minimum/maximum values. Ch refers to challenge day and Ch+1, Ch+2 etc indicate day post-challenge (i.e. day one post-challenge, day two post-challenge etc). Ch+5 indicates day five post-challenge on which four animals per group were randomly selected for sacrifice. Statistically significant between-group differences are as specified.</p

HA1 microarray serological values by study antigens, group and day.

<p>Box plots display median (dash) and mean (dot) of log₁₀-transformed HA1 protein microarray signal values. The box extends to the 25^th/75^th percentiles and whiskers extend to minimum/maximum values. H1-07 indicates A/Brisbane/59/2007 (H1N1)-like; H3-07 indicates A/Brisbane/10/2007 (H3N2)-like; H1-09 indicates A/California/7/2009 (H1N1)pdm09-like (<b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086555#pone.0086555.s004" target="_blank">Table S3</a>;</b> grey-shaded). Sample size as follows: Pre-immunization Vaccine = 15, Placebo = 16 (3 ferrets each per group pre-shipment serum was substituted owing to insufficient day 0 available); Day 28 Vaccine = 14, Placebo = 15; Day 49 Vaccine = 12, Placebo = 11; Day 54 Vaccine = 2, Placebo = 4; Day 63 Vaccine = 9, Placebo = 8. **indicates statistical significance at p<0.01 and *indicates statistical significance at p<0.05 in comparing vaccine to placebo group at the designated time point. ΔΔ indicates statistical significance at p<0.01 and Δ indicates statistical significance at p<0.05 in comparing values within study groups at days 28, 49, 54 and 63 relative to pre-immunization, colour coded by vaccine (red) or placebo (blue). □□ indicates statistical significance at p<0.01 and □ indicates statistical significance at p<0.05 in comparing day 63 to day 49 within groups, colour coded per above by study group.</p

Study protocol.

<p>Randomized blinded placebo-controlled experiment of Canadian manufactured, commercially-available 2008–09 trivalent inactivated influenza vaccine (TIV: Fluviral) on A(H1N1)pdm09 disease risk in ferrets.</p

Influenza A antibody results by study day and group based on ELISA assay.

<p><b>Competitive Nucleoprotein-based IDEXX ELISA (Influenza A antibody). Ratios <0.60 classified as positive; ratios ≥0.60 classified as negative.</b></p><p>ND = Not done; IDEXX Inc. = Commercial ELISA assay (note the lower the ratio, the greater the antibody detected); Ch = challenge.</p>a<p>Where numbers tested differ from the number randomized per group in parentheses at the specified time point it is because insufficient sera remained for testing of all animals.</p>b<p>Number of sera tested by group shown in adjacent columns.</p>c<p>One day 0 serum in each group was insufficient for ELISA testing and substituted with pre-shipment values for these ferrets. Excluding these ferrets (leaving n = 15 per group) gives ELISA ratios of 1.04 (0.99–1.09) and 1.05 (1.01–1.09) for vaccine and placebo groups, respectively.</p>d<p>Two sera belonging to placebo group close to serologic threshold for positivity with ratios of 0.61.</p