Viral expansion after transfer is a primary driver of influenza A virus transmission bottlenecks.

For many viruses, narrow bottlenecks acting during transmission sharply reduce genetic diversity in a recipient host relative to the donor. Since genetic diversity represents adaptive potential, such losses of diversity are thought to limit the opportunity for viral populations to undergo antigenic change and other adaptive processes. Thus, a detailed picture of evolutionary dynamics during transmission is critical to understanding the forces driving viral evolution at an epidemiologic scale. To advance this understanding, we used a barcoded virus library and a guinea pig model of transmission to decipher where in the transmission process influenza A virus populations lose diversity. In inoculated guinea pigs, we show that a high level of viral barcode diversity is maintained. Within-host continuity in the barcodes detected across time furthermore indicates that stochastic effects are not pronounced within the inoculated hosts. Importantly, in both aerosol-exposed and direct contact animals, we observed many barcodes at the earliest time point(s) positive for infectious virus, indicating robust transfer of diversity through the environment. This high viral diversity is short-lived, however, with a sharp decline seen 1-2 days after initiation of infection. Although major losses of diversity at transmission are well described for influenza A virus, our data indicate that events that occur following viral transfer and during the earliest stages of natural infection have a central role in this process. This finding suggests that host factors, such as immune effectors, may have greater opportunity to impose selection during influenza A virus transmission than previously recognized.