Fig. 2: Cross-reactive and protective potential of human N2 antibodies against subtype A(H9N2) influenza A viruses in Guangzhou cohort samples.

a Hemagglutination-inhibition (HI) titers against representative strains of subtype A(H9N2) AIV and antigenically-distinct human A(H3N2) influenza viruses in human circulation since 1968. Age-stratified b HI and c neuraminidase-inhibition (NI) antibody profiles against selected human A(H3N2) viruses. d Age-stratified NI-antibody profile against eight A(H9N2) AIV isolated between 1997 to 2015. e Age-stratified NI-titers against the 1968 pandemic strain, A/Aichi/2/1968 (H3N2) (AI68). f Weight loss and g survival curves of mice inoculated with pooled human sera from respective age-groups. All NI-antibody was detected using enzyme-linked lectin assay (ELLA) using recombinant viruses bearing the target NA with a HA gene from A/teal/Hong Kong/W312/1997 (H6N1) and the internal genes of A/Puerto Rico/8/1934 (H1N1) (PR8). Colored circles above the graph indicate those viruses circulating at the time of birth of the oldest participant in each age group. Dotted lines in (a–e) indicate limits of detection. The bar graphs indicate the geometric mean antibody titer with 95% confidence intervals, with n = 20 individuals per age group. Statistical significance in (d) was calculated using two-sided two-way ANOVA compared to ≥ 65-year-old age group using Dunnett’s multiple comparison test (****p < 0.0001 for all age groups). Statistical significance in (d) was calculated using two-sided two-way ANOVA compared to ≥ 65 yo group, adjusted with Dunnett’s multiple comparison test (****p < 0.0001 for all age groups). Statistical significance in (e) was calculated using two-sided one-way ANOVA compared ≥ 65 yo group, adjusted with Dunnett’s multiple comparison test (****p < 0.0001 for ≤ 5 yo, 6–10 yo, 11–20 yo and 21–39 yo, **p = 0.0057 for 40–64 yo). Weight loss was expressed as mean ± standard deviation. Survival rate in g was compared using two-sided Gehan-Breslow-Wilcoxon test with PBS as the reference group (**p = 0.0024 for 40–64 yo, ***p = 0.0002 for ≥ 65 yo). Each group had n = 10 mice, except for the 6–10 yo group and 21–39 yo group which had 8 mice, and the 40-64 yo group which had 9.