Spatiotemporal reconstruction of the North American A(H5N1) outbreak reveals successive lineage replacements by descendant reassortants

Abstract

The November 2021 introduction of highly pathogenic avian influenza A(H5N1) clade 2.3.4.4b into North America triggered a devastating outbreak, affecting more than 180 million domestic birds and spreading to more than 80 wildlife species across Canada and the US. From this outbreak, we have sequenced 2955 complete A(H5N1) viral genomes from samples collected in Canada and, in conjunction with previously published data, performed multifaceted phylodynamic analyses. These analyses reveal extensive diversification of A(H5N1) viruses via reassortment with low-pathogenic avian influenza viruses. We find evidence of repeated ancestral strain replacement by direct descendants, indicative of compounding viral fitness increases. Spatiotemporal modeling identified critical geographic areas facilitating transcontinental spread and demonstrated genotype-specific host dynamics, offering essential data for ongoing control and prevention strategies.

Fig. 1.. Emergence and surveillance of H5N1 clade 2.3.4.4b genotypes in Canada.

(A) Number of unique viral genotypes based on whole-genome sequence detected over time. (B) Relative sampling frequency of viral genotypes over time. (C) Reassortant network depicting the evolutionary origin of each genotype. The network is derived from a timescaled maximum likelihood tree based on concatenated nonreassorted genome segments HA, NA, and M (fig. S2) with discrete character reconstruction of viral genotypes at each node. Arrows represent reassortment events with the ancestral genotype at the base of the arrow and the derived genotype at the head. Only major reassortant genotypes (>150 samples) and their most recent Eurasian ancestor (A1) are given a unique color; all others are colored black.

Fig. 2.. Continuous phylogeographic inference of the three major North American genotypes with the largest contributions to the transcontinental spread of H5N1 clade 2.3.4.4b.

(A) Maps display the estimated spread for genotypes A1, B2.1, B3.2, and B3.6. Mapped points represent nodes of the MCC tree for each genotype, and lines represent transmission events, colored by the inferred median time of occurrence. Transmission lines are curved to represent the direction of dispersal, counterclockwise from origin to destination. Shaded areas represent the 80% HPD of nodes’ estimated location. (B) Number of viruses sequenced for genotypes A1, B2.1, B3.2, and B3.6 over time. Videos of the phylogeographic reconstruction presented in (A) are available in movies S1 to S4.

Fig. 3.. Continuous phylogeographic inference of two major North American H5N1 clade 2.3.4.4b genotypes.

Maps display the estimated spread for genotypes (A) B1.2 and (B) B1.3. Mapped points represent nodes of the MCC tree for each genotype, and lines represent transmission events, colored by the inferred median time of occurrence. Transmission lines are curved to represent the direction of dispersal, counterclockwise from origin to destination. Shaded areas represent the 80% HPD of nodes’ estimated location. (C) Number of viruses sequenced for genotypes B1.2 and B1.3 over time. Videos of the phylogeographic reconstruction presented in (A) and (B) are available in movies S5 and S6.

Fig. 4.. Estimates of relative fitness between A(H5N1) genotypes.

(A) Simplified maximum likelihood phylogenetic tree representing the evolutionary history of major A(H5N1) genotypes in the Americas. Estimation of relative fitness for each genotype was conducted by two methods, a multinomial logistic fitness model (B) and LBI (C). Colors of the tree branches in (A) and dotted lines between points in (B) and (C) represent genotype lineage ancestry. Error bars represent the 95% confidence interval of each data point.

Fig. 5.. Host dynamics for major H5N1 genotypes in North America.

The arrow width summarizes jump counts between host groups (Markov jumps with >0.70 posterior probability and a Bayes factor >3.0), where each arrow is colored according to the source host. The size of each node represents the median relative phylogenetic time spent in each host state (Markov rewards).