Within-host diversity in H5N1-infected humans and poultry in Cambodia

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

Louise H. Moncla1, Trevor Bedford1,2, Philippe Dussart3, Srey Viseth Horm3, Sareth Rith3, Philippe Buchy4, Erik A Karlsson3, Lifeng Li5,6, Yongmei Liu5,6, Huachen Zhu5,6, Yi Guan5,6, Thomas C. Friedrich7,8, Paul F. Horwood9,10

1Fred Hutchinson Cancer Research Center, Seattle, Washington, United States, 2University of Washington, Seattle, Washington, United States, 3Virology Unit, Institut Pasteur du Cambodge, Phnom Penh, Cambodia, 4GlaxoSmithKline, Vaccines R&D, Singapore, Singapore,5Joint Influenza Research Centre (SUMC/HKU), Shantou University Medical College, Shantou, People's Republic of China,6State Key Laboratory of Emerging Infectious Diseases/Centre of Influenza Research, School of Public Health, The University of Hong Kong, Hong Kong, SAR, People's Republic of China,7Department of Pathobiological Sciences, University of Wisconsin School of Veterinary Medicine, Madison, WI, United States,8Wisconsin National Primate Research Center, Madison, WI, United States,9Papua New Guinea Institute of Medical Research, Goroka, Paula New Guinea,10Australian Institute of Tropical Health and Medicine, James Cook University, Cairns, Australia.


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. We analyze deep sequencing data from infected humans and ducks in Cambodia to examine H5N1 evolution during spillover. Viral populations in both species are predominated by low-frequency (<10%) variation shaped by purifying selection and genetic drift. Viruses from humans contain some human-adapting mutations (PB2 E627K, HA A150V, and HA Q238L), but these mutations remain low-frequency. Within-host variants are not enriched along phylogenetic branches leading to human infections. Our data show that H5N1 viruses generate putative human-adapting mutations during natural spillover infection. However, short infections, randomness, and purifying selection limit the evolutionary capacity of H5N1 viruses within-host. Applying evolutionary methods to sequence data, we reveal a detailed view of H5N1 adaptive potential, and develop a foundation for studying host-adaptation in other zoonotic viruses.