Characterization of Influenza A(H1N1)pdm09 Viruses Isolated in the 2018–2019 and 2019–2020 Influenza Seasons in Japan

Figure 1. Phylogenetic analysis of HA. Phylogenetic tree based on the HA amino acid sequences of A(H1N1)pdm09 viruses, generated by using the Maximum Likelihood method and the Jones–Taylor–Thornton (JTT) model. Several HA sequences of representative isolates that we sequenced were used. Gray and black circles indicate isolates that were isolated in this study during the 2018–2019 and 2019–2020 influenza seasons, respectively. Current and previous vaccine strains are indicated by black squares. The branch bootstrap score is listed next to the corresponding subtree node. Amino acid substitutions delineating major branches are shown. A/Tokyo/UT-WD039-0/2019 (WD039-0) and A/Tokyo/UT-BB251/2019 (BB251) indicated in green; A/Tokyo/UT-BB262/2019 (BB262) and A/Tokyo/UT-GR156/2019 (GR156) indicated in purple; A/Tokyo/UT-GR253/2020 (GR253), and A/Tokyo/UT-AC109/2020 (AC109) indicated in orange; and A/Singapore/GP1908/2015 (Singapore) shown in green were used for the antigenic analysis. A/California/04/2009 is defined as an outgroup.

Figure 2. The positions of the N156K/S, D187A, and Q189E substitutions in HA are shown in magenta. The positions (H1 numbering) are shown on the HA structure of A/California/04/2009 (PDB ID: 3ubn). The amino acids that are important for receptor binding are colored yellow.

Figure 3. HI and neutralization antigenic maps of A(H1N1)pdm09 viruses. HI (A) and neutralization (B) titers of ferret antisera were used to construct antigenic maps. Virus names are black, whereas amino acid substitutions are grey. Viruses are abbreviated as follows: A/Singapore/GP1908/2015 (Singapore); A/Tokyo/UT-WD039-0/2019 (WD039-0); A/Tokyo/UT-BB251/2019 (BB251); A/Tokyo/UT-BB262/2019 (BB262); A/Tokyo/UT-GR156/2019 (GR156); A/Tokyo/UT-GR253/2020 (GR253); and A/Tokyo/UT-AC109/2020 (AC109). Data were analyzed by using a nonparametric Friedman test followed by Dunn’s multiple comparisons.

Figure 4. Reactivity of human plasma samples against A(H1N1)pdm09 viruses. HI (A) and neutralization (B) titers of 63 human serum samples against the indicated seven viruses are plotted as black lines; a small amount of vertical jitter has been added to prevent lines from overlapping each other. The mean of all individuals is shown in red. Antigens are grouped on the x-axis according to the substitutions of interest. Viruses are abbreviated as follows: A/Singapore/GP1908/2015 (Singapore); A/Tokyo/UT-WD039-0/2019 (WD039-0); A/Tokyo/UT-BB251/2019 (BB251); A/Tokyo/UT-BB262/2019 (BB262); A/Tokyo/UT-GR156/2019 (GR156); A/Tokyo/UT-GR253/2020 (GR253); and A/Tokyo/UT-AC109/2020 (AC109). ** indicates p < 0.01 compared with Singapore.