Quantifying within-host diversity of H5N1 influenza viruses in humans and poultry in Cambodia

Abstract

Avian influenza viruses (AIVs) periodically cross species barriers and infect humans. The likelihood that an AIV will evolve mammalian transmissibility depends on acquiring and selecting mutations during spillover, but data from natural infection is limited. We analyze deep sequencing data from infected humans and domestic ducks in Cambodia to examine how H5N1 viruses evolve during spillover. Overall, viral populations in both species are predominated by low-frequency (<10%) variation shaped by purifying selection and genetic drift, and half of the variants detected within-host are never detected on the H5N1 virus phylogeny. However, we do detect a subset of mutations linked to human receptor binding and replication (PB2 E627K, HA A150V, and HA Q238L) that arose in multiple, independent humans. PB2 E627K and HA A150V were also enriched along phylogenetic branches leading to human infections, suggesting that they are likely human-adaptive. Our data show that H5N1 viruses generate putative human-adapting mutations during natural spillover infection, many of which are detected at >5% frequency within-host. However, short infection times, genetic drift, and purifying selection likely restrict their ability to evolve extensively during a single infection. Applying evolutionary methods to sequence data, we reveal a detailed view of H5N1 virus adaptive potential, and develop a foundation for studying hostadaptation in other zoonotic viruses.

Phenotypic Effects of Substitutions within the Receptor Binding Site of Highly Pathogenic Avian Influenza H5N1 Virus Observed during Human Infection

“…Others also observed the 226R/K substitutions during human infection (35,40,50): 226R was also observed during contact transmission studies of chickens that were infected with the aerosol transmittable H5N1 virus containing Q226L/G228S (29). Richard et al (29) showed that besides substitutions PB2-E627K, HA substitutions Q226L and G228S caused substantial attenuation in chickens.…”

Transmission of SARS-CoV-2 in domestic cats imposes a narrow bottleneck

“…Purifying selection, which acts to purge deleterious mutations from a population, is known to result in an excess of low-frequency variants. In contrast, positive selection results in the accumulation of intermediate- and high-frequency variation [ 37 ]. Especially in the setting of an acute viral infection, exponential population growth is also expected to result in an excess of low-frequency variants [ 38 ].…”

“…Especially in the setting of an acute viral infection, exponential population growth is also expected to result in an excess of low-frequency variants [ 38 ]. To determine the type of evolutionary pressure acting on SARS-CoV-2 in cats, we plotted these distributions against a simple “neutral model” (light grey bars in Fig 2B ), which assumes a constant population size and the absence of selection [ 37 ]. This model predicted that ~43% of polymorphisms would fall in the 3–10% frequency bin, ~25% into the 10–20% bin, ~14% into the 20–30% bin, ~10% into the 30–40% bin, and ~8% into the 40–50% bin.…”

“…We plotted the counts of variants falling into each frequency bin using Matplotlib 3.3.2 ( https://matplotlib.org ). We used code written by Dr. Louise Moncla to generate the distribution of SNPs for a given population assuming no selection or change in population size, which is expected to follow a 1/x distribution [ 37 ]. The code to replicate this can be found in the GitHub accompanying this manuscript, specifically in the ‘code/SFS.ipynb’ Jupyter Notebook.…”

Identification and Characterization of Swine Influenza Virus H1N1 Variants Generated in Vaccinated and Nonvaccinated, Challenged Pigs

“…This finding may indicate a purifying selection event where the deleterious mutations are being eliminated, as previously reported. [ 51 , 52 ]. Thus, no positive selection pressure was found in nonvaccinated animals.…”