Divergent evolutionary trajectories of influenza B viruses underlie their contemporaneous epidemic activity

Abstract

Influenza B viruses have circulated in humans for over 80 y, causing a significant disease burden. Two antigenically distinct lineages ("B/Victoria/2/87-like" and "B/Yamagata/16/88-like," termed Victoria and Yamagata) emerged in the 1970s and have cocirculated since 2001. Since 2015 both lineages have shown unusually high levels of epidemic activity, the reasons for which are unclear. By analyzing over 12,000 influenza B virus genomes, we describe the processes enabling the long-term success and recent resurgence of epidemics due to influenza B virus. We show that following prolonged diversification, both lineages underwent selective sweeps across the genome and have subsequently taken alternate evolutionary trajectories to exhibit epidemic dominance, with no reassortment between lineages. Hemagglutinin deletion variants emerged concomitantly in multiple Victoria virus clades and persisted through epistatic mutations and interclade reassortment-a phenomenon previously only observed in the 1970s when Victoria and Yamagata lineages emerged. For Yamagata viruses, antigenic drift of neuraminidase was a major driver of epidemic activity, indicating that neuraminidase-based vaccines and cross-reactivity assays should be employed to monitor and develop robust protection against influenza B morbidity and mortality. Overall, we show that long-term diversification and infrequent selective sweeps, coupled with the reemergence of hemagglutinin deletion variants and antigenic drift of neuraminidase, are factors that contributed to successful circulation of diverse influenza B clades. Further divergence of hemagglutinin variants with poor cross-reactivity could potentially lead to circulation of 3 or more distinct influenza B viruses, further complicating influenza vaccine formulation and highlighting the urgent need for universal influenza vaccines.

Keywords: antigenic; genetic diversity; natural selection; phylogeny; vaccine.

Fig. 1.

Phylodynamics of global influenza B virus. Relative genetic diversities and temporal phylogenies of the HA and NA gene segments of the Victoria (A and B) and Yamagata lineages (C and D). Among the Victoria lineages, gray tip circles represent viruses in absence of HA deletions. Orange tip circles denote clade 1A.1 viruses with 2-aa deletions at positions 162–163 of the HA protein. Green, pink, and blue tip circles denote different clades (1A.2–1A.4) harboring a 3-aa deletion at positions 162–164 of the HA protein. For each gene phylogeny, trunk amino acid substitutions are mapped at the backbone of each tree. Amino acid substitutions in bold indicate potential N-linked glycosylation sites. (E) Three-dimensional structure of the trimeric HA (PDB ID code: 4FQM) and (F) tetrameric NA (PDB ID code: 3K36) proteins of B/Brisbane/60/2008 (Victoria lineage) and B/Perth/211/2001(Yamagata lineage) as reference, respectively. Trunk amino acid substitutions that occurred in the Victoria clade 1A.1–1A.3 and the Yamagata clade 3A viruses are mapped.

Fig. 2.

Schematic diagrams showing the putative cooccurring mutations within and between gene segments, and selection pressure of influenza B viruses. (A) HA phylogeny of the Victoria lineage. Orange branches denote clade 1A.1 viruses with 2-aa deletions (2-aa), and blue branches denote viruses with no deletion. (B) NA phylogeny of the Yamagata lineage. Purple branches denote clade 3A viruses. Colored circles represent trunk amino acid mutations on gene segments. Dotted arrows indicate putative cooccurring mutations. (C) Estimates of global d_N/d_S ratios for each gene segment inferred using SLAC method, with the significance level at P < 0.05. The means of d_N/d_S ratios are represented by circle-shaped symbols with error bars indicating 95% confidence intervals. Colored bars denote different gene segments.

Fig. 3.

Evolutionary relationships of each gene segment of the Victoria lineage, displaying interclade reassortment within the lineage, highlighting the deletion variants. Reassortment events of Yamagata viruses are shown in SI Appendix, Fig. S25.

Fig. 4.

Phylogeography and age distribution of influenza B viruses. (A) Proportion of ancestral geographical location states estimated on the phylogenetic trunk of Victoria (VIC) and Yamagata (YAM) viruses through time. Shaded areas represent estimated ancestral location state proportions in each location. (B) Patterns of migration rates are shown on a 2D matrix where each cell corresponds to mean migration rates between pairwise geographic locations. Blue and purple cells denote Victoria and Yamagata, respectively. Abbreviations for location states: Africa (AF), East Asia (EA), Europe (EU), Middle East/West Asia (ME/WA), North America (NAm), Oceania (OC), South America (SAm), South Asia (SA), and Southeast Asia (SEA). (C) Age density distribution of influenza B virus by lineage. The violin plot indicates the distribution, and the box-plot indicates the interquartile range (IQR). The median ages are represented by a notched horizontal line. The vertical line indicates the 1.5X IQR, and the outliers are represented by dots. The Mann–Whitney U test was used to compare the median ages, with statistical significance indicated by P < 0.0001. (D) Age density distribution of Victoria clade 1A viruses with multiple amino acid deletions on the HA protein. Orange plot represents viruses with 2-aa deletion (clade 1A.1), green plot indicates viruses with 3-aa deletion (clade 1A.2–1A.4), and gray plot indicates viruses with no deletion. Mann–Whitney U test P values indicated by P < 0.05.