Annual influenza A(H3N2) epidemics in the United States, 1997 - 2019.

A. Weekly incidence of influenza A(H1N1) (blue), A(H3N2) (red), and B (green) averaged across ten HHS regions (Region 1: Boston; Region 2: New York City; Region 3: Washington, DC; Region 4: Atlanta; Region 5: Chicago; Region 6: Dallas, Region 7: Kansas City; Region 8: Denver; Region 9: San Francisco; Region 10: Seattle). Incidences are the proportion of influenza-like illness (ILI) visits among all outpatient visits, multiplied by the proportion of respiratory samples testing positive for each influenza type/subtype. Time series are 95% confidence intervals of regional incidence estimates. Vertical dashed lines indicate January 1 of each year. B. Intensity of weekly influenza A(H3N2) incidence in ten HHS regions. White tiles indicate weeks when influenza-like-illness data or virological data were not reported. Data for Region 10 are not available in seasons prior to 2009.

Antigenic and genetic evolution of seasonal influenza A(H3N2) viruses, 1997 - 2019.

A-B. Temporal phylogenies of hemagglutinin (H3) and neuraminidase (N2) gene segments. Tip color denotes the Hamming distance from the root of the tree, based on the number of substitutions at epitope sites in H3 (N = 129 sites) and N2 (N = 223 sites). “X” marks indicate the phylogenetic positions of US recommended vaccine strains. C-D. Seasonal genetic and antigenic distances are the mean distance between A(H3N2) viruses circulating in the current season t versus the prior season (𝑡 – 1), measured by C. four sequence-based metrics (HA receptor binding site (RBS), HA stalk footprint, HA epitope, and NA epitope) and D. hemagglutination inhibition (HI) titer measurements. E. The Shannon diversity of H3 and N2 local branching index (LBI) values in each season. Vertical bars in C, D, and E and are 95% confidence intervals of seasonal estimates from five bootstrapped phylogenies.

Evolutionary indicators of seasonal viral fitness.

Evolutionary indicators are labeled by the influenza gene for which data are available (hemagglutinin, HA or neuraminidase, NA), the type of data they are based on, and the component of influenza fitness they represent. Table format is adapted from Huddleston et al., 2020.

Seasonal metrics of A(H3N2) epidemic dynamics.

Epidemic metrics are defined and labeled by which outcome category they represent.

Influenza A(H3N2) antigenic drift correlates with larger, more intense annual epidemics.

A(H3N2) epidemic size, peak incidence, transmissibility (effective reproduction number, 𝑅t), and epidemic intensity increase with antigenic drift, measured by A. hemagglutinin (H3) epitope distance, B. neuraminidase (N2) epitope distance, and C. hemagglutination inhibition (HI) log2 titer distance. Seasonal antigenic drift is the mean titer distance or epitope distance between viruses circulating in the current season t versus the prior season (𝑡 – 1) or two seasons ago (𝑡 – 2). Distances are scaled to aid in direct comparison of evolutionary indicators. Point color indicates the dominant influenza A virus (IAV) subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bands are 95% confidence intervals of regional estimates. Seasonal mean A(H3N2) epidemic metric values were fit as a function of antigenic or genetic distance using LMs (epidemic size, peak incidence), Gaussian GLMs (effective 𝑅t: inverse link), or Beta GLMs (epidemic intensity: logit link) with 1000 bootstrap resamples.

The proportion of influenza positive samples typed as A(H3N2) increases with antigenic drift.

A-B. Seasonal A(H3N2) subtype dominance increases with H3 and N2 epitope distance. Seasonal epitope distance is the mean epitope distance between viruses circulating in the current season t versus the prior season (𝑡 – 1) or two prior seasons ago (𝑡 – 2). Distances were scaled to aid in direct comparison of evolutionary indicators. Point color indicates the dominant influenza A virus (IAV) subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bands are 95% confidence intervals of regional estimates. Seasonal mean A(H3N2) dominance was fit as a function of H3 or N2 epitope distance using Beta GLMs with 1000 bootstrap resamples. C-D. Regional patterns of influenza type and subtype incidence during two seasons when A(H3N2) was nationally dominant. Pie charts represent the proportion of influenza positive samples typed as A(H3N2) (red), A(H1N1) (blue), or B (green) in each HHS region. The sizes of regional pie charts are proportional to the total number of influenza positive samples. Data for Region 10 (purple) are not available for seasons prior to 2009. C. Widespread A(H3N2) dominance during 2003-2004 after the emergence of a novel antigenic cluster, FU02 (A/Fujian/411/2002-like strains). D. Spatial heterogeneity in subtype circulation during 2007-2008, a season with low A(H3N2) antigenic novelty relative to the prior season.

Influenza A(H3N2) seasonal duration increases with the diversity of H3 and N2 clade growth rates in each season.

Seasonal diversity of clade growth rates is measured as the A. Shannon diversity or B. standard deviation (s.d.) of H3 and N2 local branching index (LBI) values of viruses circulating in each season. LBI values are scaled to aid in direct comparisons of H3 and N2 LBI diversity. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant). Mean seasonal duration was fit as a function of H3 or N2 LBI diversity using Gaussian GLMs (inverse link) with 1000 bootstrap resamples.

N2 epitope distance significantly correlates with the age distribution of outpatient influenza-like illness (ILI) cases.

Seasonal epitope distance is the mean distance between strains circulating in season t and those circulating in the prior season (𝑡 – 1) or two seasons ago (𝑡 – 2). Distances are scaled to aid in direct comparison of evolutionary indicators. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bars are 95% confidence intervals of regional age distribution estimates. The seasonal mean fraction of cases in each age group were fit as a function of H3 or N2 epitope distance using Beta GLMs (logit link) with 1000 bootstrap resamples.

The effects of influenza A(H1N1) and B epidemic size on A(H3N2) epidemic burden.

A. Influenza A(H1N1) epidemic size negatively correlates with A(H3N2) epidemic size, peak incidence, transmissibility (effective reproduction number, 𝑅t), and epidemic intensity. B. Influenza B epidemic size does not significantly correlate with A(H3N2) epidemic metrics. Point color indicates the dominant influenza A virus (IAV) subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical and horizontal bands are 95% confidence intervals of regional estimates. Seasonal mean A(H3N2) epidemic metrics were fit as a function of mean A(H1N1) or B epidemic size using Gaussian GLMs (epidemic size and peak incidence: inverse link; effective 𝑅t: log link) or Beta GLMs (epidemic intensity: logit link) with 1000 bootstrap resamples.

Variable importance rankings from conditional inference random forest models predicting seasonal region-specific influenza A(H3N2) epidemic dynamics.

Ranking of variables in predicting regional A(H3N2) A. epidemic size, B. peak incidence, C. transmissibility (maximum effective reproduction number, 𝑅t), D. epidemic intensity, and E. subtype dominance. Each forest was created by generating 3,000 regression trees from a repeated leave-one-season-out cross-validated sample of the data. Variables are ranked by their conditional permutation importance, with differences in prediction accuracy scaled by the total (null model) error. Black error bars are 95% confidence intervals of conditional permutation scores. Abbreviations: 𝑡 – 1 = one-season lag, 𝑡 – 2 = two-season lag, IAV = influenza A virus subtype, s.d. = standard deviation, HI = hemagglutination inhibition, LBI = local branching index, distance to vaccine = epitope distance between currently circulating strains and the recommended vaccine strain, VE = vaccine effectiveness.

Observed versus predicted values of seasonal region-specific influenza A(H3N2) A. epidemic size, B. peak incidence, C. maximum effective reproduction number, 𝑹𝒕, D. epidemic intensity, and E. subtype dominance from conditional random forest models.

Results are facetted by HHS region and epidemic metric. Point color and size corresponds to the degree of hemagglutinin (H3) epitope distance between viruses circulating in season t versus viruses circulating two seasons ago (𝑡 – 2). Large, yellow points indicate seasons with high antigenic novelty, and small blue points indicate seasons with low antigenic novelty. Regional Spearman’s rank correlation coefficients and associated P-values are in the top left section of each facet.

Predictors of seasonal A(H3N2) epidemic burden, transmissibility, intensity, and subtype dominance. Variables retained in the best fit model for each epidemic outcome were determined by BIC.

Intensity of weekly incidence of A. influenza A(H1N1) and B. influenza B in ten HHS regions, 1997 - 2019.

Incidences are the proportion of influenza-like illness (ILI) visits among all outpatient visits, multiplied by the proportion of respiratory samples testing positive for each influenza type/subtype. Seasonal and pandemic A(H1N1) are combined as A(H1N1), and the Victoria and Yamagata lineages of influenza B are combined as influenza B. White tiles indicate weeks when either influenza-like-illness cases or virological data were not reported. Data for Region 10 are not available in seasons prior to 2009.

Influenza test volume systematically increases in all HHS regions after the 2009 A(H1N1) pandemic.

Each point represents the total number of influenza tests in each HHS region in each season, as reported by the US CDC WHO Collaborating Center for Surveillance, Epidemiology and Control of Influenza. Approximately 100 public health laboratories and 300 clinical laboratories located throughout the US report influenza test results to the US CDC, through either the US WHO Collaborating Laboratories Systems or the National Respiratory and Enteric Virus Surveillance System (NREVSS).

Pairwise correlations between seasonal influenza A(H3N2), A(H1N1), and B epidemic metrics.

Spearman’s rank correlations among indicators of A(H3N2) epidemic timing, including onset week, peak week, regional variation (s.d.) in onset and peak timing, the number of days from epidemic onset to peak incidence, and seasonal duration, indicators of A(H3N2) epidemic magnitude, including epidemic intensity (i.e., the “sharpness” of the epidemic curve), transmissibility (maximum effective reproduction number, 𝑅t), subtype dominance, epidemic size, and peak incidence. We also considered relationships between the circulation of other influenza types/subtypes and A(H3N2) epidemic burden and timing. The Benjamini and Hochberg method was used to adjust P-values for multiple testing. The color of each circle indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation). Stars within circles indicate statistical significance (adjusted P < 0.05).

The number of A/H3 sequences in five subsampled datasets in each month and each influenza season.

In each figure, the five subsampled datasets are plotted individually but individual time series are difficult to discern due to minor differences in sequence counts across the datasets. A. The number of sequences in subsampled datasets in each month collected in North America (purple) versus nine other world regions combined (dark green). B. The total number of sequences in subsampled datasets collected in each month in all world regions combined. C. The number of sequences in subsampled datasets in each season collected in North America (purple) versus nine other world regions combined (dark green). D. The total number of sequences in subsampled datasets collected in each season in all world regions combined.

The number of A/N2 sequences in five subsampled datasets in each month and each influenza season.

In each figure, the five subsampled datasets are plotted individually but individual time series are difficult to discern due to minor differences in sequence counts across the datasets. A. The number of sequences in subsampled datasets in each month collected in North America (purple) versus nine other world regions combined (dark green). B. The total number of sequences in subsampled datasets collected in each month in all world regions combined. C. The number of sequences in subsampled datasets in each season collected in North America (purple) versus nine other world regions combined (dark green). D. The total number of sequences in subsampled datasets in each season in all world regions combined.

A/H3 sequence counts in five subsampled datasets.

We downloaded all H3 sequences and associated metadata from the GISAID EpiFlu database and focused our analysis on complete H3 sequences that were sampled between January 1, 1997, and October 1, 2019. To account for variation in sequence availability across global regions, we subsampled the selected sequences five times to representative sets of no more than 50 viruses per month, with preferential sampling for North America. Each month up to 25 viruses were selected from North America (when available) and up to 25 viruses were selected from nine other global regions (when available), with even sampling across the other global regions (China, Southeast Asia, West Asia, Japan and Korea, South Asia, Oceania, Europe, South America, and Africa).

A/N2 sequence counts in five subsampled datasets.

We downloaded all N2 sequences and associated metadata from the GISAID EpiFlu database and focused our analysis on complete N2 sequences that were sampled between January 1, 1997, and October 1, 2019. To account for variation in sequence availability across global regions, we subsampled the selected sequences five times to representative sets of no more than 50 viruses per month, with preferential sampling for North America. Each month up to 25 viruses were selected from North America (when available) and up to 25 viruses were selected from nine other global regions (when available), with even sampling across the other global regions (China, Southeast Asia, West Asia, Japan and Korea, South Asia, Oceania, Europe, South America, and Africa).

Comparison of seasonal antigenic drift measured by substitutions at hemagglutinin (H3) epitope sites and hemagglutination inhibition (HI) log2 titer measurements, from 1997-1998 to 2018-2019.

Spearman’s rank correlations between H3 epitope distance and HI log2 titer distance at A. one-season lags and B. two-season lags. Seasonal antigenic distance is the mean distance between strains circulating in season t and strains circulating in the prior season 𝑡 – 1 year (one season lags) or two seasons ago 𝑡 – 2 years (two season lags). Seasonal distances are scaled because H3 epitope distance and HI log2 titer distance use different units of measurement. Point labels indicate the current influenza season, and point color denotes the relative timing of influenza seasons, with earlier seasons shaded dark purple (e.g., 1997-1998) and later seasons shaded light yellow (e.g., 2018-2019). H3 epitope distance and HI log2 titer distance at two-season lags capture expected “jumps” in antigenic drift during key seasons previously associated with major antigenic transitions (Smith et al., 2004), such as the SY97 cluster seasons (1997-1998, 1998-1999, 1999-2000), the FU02 cluster season (2003-2004), and the CA04 cluster season (2004-2005).

Pairwise correlations between H3 and N2 evolutionary indicators (one season lags).

Spearman’s rank correlations between seasonal measures of H3 and N2 evolution, including H3 RBS distance, H3 epitope distance, H3 non-epitope distance, H3 stalk footprint distance, HI log2 titer distance, N2 epitope distance based on 223 or 53 epitope sites, N2 non-epitope distance, and the standard deviation (s.d.) and Shannon diversity of H3 and N2 local branching index (LBI) values in the current season t. Seasonal distances were estimated as the mean distance between strains circulating in the current season t and those circulating in the prior season (𝑡 – 1). The Benjamini and Hochberg method was used to adjust P-values for multiple testing. The color of each circle indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation). Stars within circles indicate statistical significance (adjusted P < 0.05).

Pairwise correlations between H3 and N2 evolutionary indicators (two season lags).

We measured Spearman’s rank correlations between seasonal measures of H3 and N2 evolution, including H3 RBS distance, H3 epitope distance, H3 non-epitope distance, H3 stalk footprint distance, HI log2 titer distance, N2 epitope distance based on 223 or 53 epitope sites, N2 non-epitope distance, and the standard deviation (s.d.) and Shannon diversity of H3 and N2 local branching index (LBI) values in the current season t. Seasonal distances were estimated as the mean distance between strains circulating in the current season t and those circulating two seasons ago (𝑡 – 2). The Benjamini and Hochberg method was used to adjust P-values for multiple testing. The color of each circle indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation). Stars within circles indicate statistical significance (adjusted P < 0.05).

Comparison of seasonal antigenic drift measured by substitutions at hemagglutinin (H3) and neuraminidase (N2) epitope sites, from 1997-1998 to 2018-2019.

Spearman’s rank correlations between H3 epitope distance and N2 epitope distance at A. one-season lags and B. two-season lags. Seasonal epitope distance is the mean distance between strains circulating in season t and strains circulating in the prior season 𝑡 – 1 (one season lag) or two seasons ago 𝑡 – 2 (two season lag). Point labels indicate the current influenza season, and point color denotes the relative timing of influenza seasons, with earlier seasons shaded dark purple (e.g., 1997-1998) and later seasons shaded light yellow (e.g., 2018-2019). H3 epitope distance at two-season lags and N2 epitope distance at one-season lags capture expected “jumps” in antigenic drift during key seasons previously associated with major antigenic transitions (Smith et al., 2004), such as the SY97 cluster seasons (1997-1998, 1998-1999, 1999-2000), the FU02 cluster season (2003-2004), and the CA04 cluster season (2004-2005).

Pairwise correlations between H3 and N2 evolutionary indicators (one- and two-season lags).

We measured Spearman’s rank correlations between seasonal measures of H3 and N2 evolution, including H3 RBS distance, H3 epitope distance, H3 non-epitope distance, H3 stalk footprint distance, HI log2 titer distance, N2 epitope distance based on 223 or 53 epitope sites, N2 non-epitope distance, and the standard deviation (s.d.) and Shannon diversity of H3 and N2 local branching index (LBI) values in the current season t. Seasonal distances were estimated as the mean distance between strains circulating in the current season t and those circulating in the prior season (𝑡 – 1) or two seasons ago (𝑡 – 2). The Benjamini and Hochberg method was used to adjust P-values for multiple testing. The color of each circle indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation). Stars within circles indicate statistical significance (adjusted P < 0.05).

Univariate correlations between influenza A(H3N2) evolutionary indictors and epidemic impact.

Mean Spearman’s rank correlation coefficients, 95% confidence intervals of correlation coefficients, and corresponding p-values of bootstrapped (N = 1000) evolutionary indicators (rows) and epidemic metrics (columns). Point color indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation), and stars indicate statistical significance (* P < 0.05, ** P < 0.01, *** P < 0.001). Abbreviations: 𝑡 – 1 = one-season lag, 𝑡 – 2 = two-season lag, RBS: receptor binding site, HI = hemagglutination inhibition, s.d. = standard deviation, LBI = local branching index.

Excess influenza A(H3N2) mortality increases with H3 and N2 antigenic drift, but correlations are not statistically significant.

The number of excess influenza deaths attributable to A(H3N2) (per 100,000 people) were estimated from a seasonal regression model fit to weekly pneumonia and influenza-coded deaths (Hansen et al., 2022). Seasonal epitope distance is the mean distance between strains circulating in season t and those circulating in the prior season (𝑡 – 1) or two seasons ago (𝑡 – 2). Distances are scaled to aid in direct comparison of evolutionary indicators. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bars are 95% confidence intervals of excess mortality estimates. Seasonal national excess mortality estimates were fit as a function of H3 or N2 epitope distance using Gaussian GLMs (log link) with 1000 bootstrap resamples.

Low seasonal diversity in the clade growth rates of circulating A(H3N2) viruses correlates with higher transmissibility and greater epidemic intensity.

A(H3N2) effective 𝑅t and epidemic intensity negatively correlate with the seasonal diversity of local branching index (LBI) values among circulating A(H3N2) lineages in the current season, measured by the standard deviation (s.d.) of A. H3 LBI values, and B. N2 LBI values. LBI values are scaled to aid in direct comparisons of H3 and N2 s.d. LBI values. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bands are 95% confidence intervals of regional estimates. Seasonal mean A(H3N2) epidemic metric values were fit as a function of H3 or N2 LBI diversity using Gaussian GLMs (effective 𝑅t: inverse link) or Beta GLMs (epidemic intensity: logit link) with 1000 bootstrap resamples.

Low seasonal diversity in the clade growth rates of circulating A(H3N2) viruses correlates with higher transmissibility and greater epidemic intensity.

A(H3N2) effective 𝑅t and epidemic intensity negatively correlate with the seasonal diversity of local branching index (LBI) values among circulating A(H3N2) lineages in the current season, measured by the Shannon diversity of A. H3 LBI values, and B. N2 LBI values. LBI values are scaled to aid in direct comparisons of H3 and N2 LBI diversity values. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bands are 95% confidence intervals of regional estimates. Seasonal mean A(H3N2) epidemic metric values were fit as a function of H3 or N2 LBI diversity using Gaussian GLMs (effective 𝑅t: inverse link) or Beta GLMs (epidemic intensity: logit link) with 1000 bootstrap resamples.

Regional patterns of influenza type and subtype circulation during seasons 1997-1998 to 2018-2019.

Pie charts represent the proportion of influenza positive samples that were typed as A(H3N2), A(H1N1) or A(H1N1)pdm09, and B in each HHS region. Data for Region 10 (purple) are not available for seasons prior to 2009.

Univariate correlations between influenza A(H3N2) evolutionary indicators and epidemic timing.

Mean Spearman’s rank correlation coefficients, 95% confidence intervals of correlation coefficients, and corresponding p-values of bootstrapped (N = 1000) evolutionary indicators (columns) and epidemic timing metrics (rows). Epidemic timing metrics are the week of epidemic onset, regional variation (s.d.) in onset timing, the week of epidemic peak, regional variation (s.d.) in peak timing, the number of days between epidemic onset and peak, and seasonal duration. Color indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation), and stars indicate statistical significance (* P < 0.05, ** P < 0.01, *** P < 0.001). Abbreviations: 𝑡 – 1 = one-season lag, 𝑡 – 2 = two-season lag, RBS: receptor binding site, HI = hemagglutination inhibition, s.d. = standard deviation, LBI = local branching index.

Epidemic speed increases with N2 antigenic drift.

N2 epitope distance significantly correlates with fewer days from epidemic onset to peak (A), while the relationship between H3 epitope distance and epidemic speed is weaker (B). Seasonal epitope distance is the mean distance between strains circulating in season t and those circulating in the prior season (𝑡 – 1) or two seasons ago (𝑡 – 2). Distances are scaled to aid in direct comparison of evolutionary indicators. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant). The seasonal mean number of days from onset to peak was fit as a function of H3 or N2 epitope distance using Gamma GLMs (inverse link) with 1000 bootstrap resamples.

Influenza A(H3N2) epidemic onsets and peaks are earlier in seasons with high antigenic novelty, but correlations are not statistically significant.

A. Epidemic onsets are earlier in seasons with increased H3 epitope distance (𝑡 – 2), but the correlation is not statistically significant. B. Epidemic peaks are earlier in seasons with increased H3 epitope distance (𝑡 – 2) and N2 epitope distance (𝑡 – 1), but correlations are not statistically significant. Seasonal epitope distance is the mean distance between strains circulating in season t and those circulating in the prior season (𝑡 – 1) or two seasons ago (𝑡 – 2). Distances are scaled to aid in direct comparison of evolutionary indicators. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant). Seasonal mean epidemic onsets and peaks were fit as a function of H3 or N2 epitope distance using Gaussian GLMs (inverse link) with 1000 bootstrap resamples.

Univariate correlations between A(H3N2) antigenic change and the age distribution of outpatient influenza-like illness (ILI) cases.

Mean Spearman’s rank correlation coefficients, 95% confidence intervals of correlation coefficients, and corresponding p-values of bootstrapped (N = 1000) evolutionary indicators (rows) and the proportion of ILI cases in individuals aged < 5 years, 5-24 years, 25-64 years, and ≥ 65 years (columns). Color indicates the strength and direction of the association, from dark red (strong positive correlation) to dark blue (strong negative correlation), and stars indicate statistical significance (* P < 0.05, ** P < 0.01, *** P < 0.001). Abbreviations: 𝑡 – 1 = one-season lag, 𝑡 – 2 = two-season lag, RBS: receptor binding site, and HI = hemagglutination inhibition.

National excess influenza A(H3N2) mortality decreases with A(H1N1) epidemic size but not B epidemic size.

Excess influenza deaths attributable to A(H3N2) (per 100,000 people) were estimated from a seasonal regression model fit to weekly pneumonia and influenza-coded deaths. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bands are 95% confidence intervals of model estimates. Seasonal national excess mortality estimates were fit as a function of A(H1N1) or B epidemic size using Gaussian GLMs (log link) with 1000 bootstrap resamples.

The effect of influenza A(H1N1) epidemic size on A(H3N2) epidemic burden during A. the entire study period (1997-2019), B. pre-2009 seasons, and C. post-2009 seasons.

Influenza A(H1N1) epidemic size negatively correlates with A(H3N2) epidemic size, peak incidence, transmissibility (maximum effective reproduction number, 𝑅t), and epidemic intensity. Point color indicates the dominant influenza A virus (IAV) subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical and horizontal bands are 95% confidence intervals of regional estimates. Seasonal mean A(H3N2) epidemic metrics were fit as a function of A(H1N1) epidemic size using Gaussian GLMs (epidemic size, peak incidence: inverse link; effective 𝑅t: log link) or Beta GLMs (epidemic intensity: logit link) with 1000 bootstrap resamples.

Wavelet analysis of influenza A(H3N2), A(H1N1), and B epidemic timing.

A. A(H3N2) incidence precedes A(H1N1) incidence in most seasons. Although A(H1N1) incidence sometimes leads or is in phase with A(H3N2) incidence (negative or zero phase lags), the direction of seasonal phase lags is not clearly associated with A(H1N1) epidemic size. B. A(H3N2) incidence leads B incidence (positive phase lag) during every season, irrespective of B epidemic size. Point color indicates the dominant influenza A subtype based on CDC influenza season summary reports (red: A(H3N2), blue: A(H1N1), purple: A(H1N1)pdm09, orange: A(H3N2)/A(H1N1)pdm09 co-dominant), and vertical bars are 95% confidence intervals of regional estimates. To estimate the relative timing of influenza subtype incidences, phase angle differences were calculated as phase in A(H3N2) minus phase in A(H1N1) (or B), with a positive value indicating that A(H1N1) (or B) incidence lags A(H3N2) incidence. To calculate seasonal phase lags, we averaged pairwise phase angle differences from epidemic week 40 to epidemic week 20. Seasonal phase lags were fit as a function of A(H1N1) or B epidemic size using LMs with 1000 bootstrap resamples.

Variable importance rankings from LASSO regression models predicting seasonal region-specific influenza A(H3N2) epidemic dynamics.

Ranking of variables in predicting regional A(H3N2) A. epidemic size, B. peak incidence, C. transmissibility (maximum effective reproduction number, 𝑅t), D. epidemic intensity, and E. subtype dominance. Models were tuned using a repeated leave-one-season-out cross-validated sample of the data. Variables are ranked by their coefficient estimates, with differences in prediction accuracy scaled by the total (null model) error. Abbreviations: 𝑡 – 1 = one-season lag, 𝑡 – 2 = two-season lag, IAV = influenza A virus subtype, s.d. = standard deviation, HI = hemagglutination inhibition, LBI = local branching index, distance to vaccine = epitope distance between currently circulating strains and the recommended vaccine strain, VE = vaccine effectiveness.

Relationships between the predictive accuracy of random forest models and seasonal H3 epitope distance.

Root mean squared errors between observed and model-predicted values were averaged across regions for each season, and results are facetted according to epidemic metric. Point color corresponds to the degree of H3 epitope distance in viruses circulating in season t relative to those circulating two seasons ago (𝑡 – 2), with bright yellow points indicating seasons with greater antigenic novelty. Spearman’s rank correlation coefficients and associated P-values are provided in the top left section of each facet.

Relationships between the predictive accuracy of random forest models and seasonal N2 epitope distance

Root mean squared errors between observed and model-predicted values were averaged across regions for each season, and results are facetted according to epidemic metric. Point color corresponds to the degree of N2 epitope distance in viruses circulating in season t relative to those circulating in the prior season (𝑡 – 1), with bright yellow points indicating seasons with greater antigenic novelty. Spearman’s rank correlation coefficients and associated P-values are provided in the top left section of each facet.

Comparison of influenza A(H3N2) epidemic timing between A(H3N2) and A(H1N1) dominant seasons. We defined influenza A virus (IAV) subtype dominance in each season based on the proportion of IAV positive samples typed as A(H3N2). We categorized seasons as A(H3N2) or A(H1N1) dominant when ≥ 70% of IAV positive samples were typed as one IAV subtype. We used two-sided Wilcoxon rank-sum tests to compare the distributions of epidemic timing metrics between A(H3N2) and A(H1N1) dominant seasons. Abbreviations: EW40 = epidemic week 40 (the start of the influenza season); s.d. = standard deviation.