The Impact of Viral and Host Factors on the Influenza A Virus Transmission Bottleneck

Background. Transmission bottlenecks restrict the viral diversity that transmits from one host to another and have a major impact on the spread of mutations in a population. Previous work has identified a tight transmission bottleneck for influenza A virus (IAV) based on an analysis of 43 household pairs, largely from a single H3N2 predominate season. While many viral and host factors are known to influence IAV transmission in households, their impact on the transmission bottleneck is not clear.Methods. Nasal swabs were collected daily from IAV infected individuals in two case-ascertained household studies, FluTES (2017/2018 through 2019/2020 seasons) and RVTN (2021/2022 season). Viruses were sequenced in duplicate, and intrahost single nucleotide variants (iSNV) were identified at a 0.5% frequency threshold using a benchmarked pipeline with >99.99% specificity for mutations present in both replicates. Transmission pairs were defined based on co-residence, test date, and genetic distance. For each possible transmission pair, the bottleneck was estimated using a beta binomial and a clonal mutation model.Results. We sequenced 567 samples from 319 individuals and 102 households in duplicate. Based on epidemiologic linkage and a sequence-based cut-off, we defined 56 transmission pairs for the beta binomial model and 60 transmission pairs for the clonal mutation model. Across all pairs, we identified a bottleneck of 1 (CI: 1, 1) using the beta-binomial model and 1 (exact estimate: 0.91, CI: 0.91, 0.97) using the clonal mutation model. In our cohort, influenza season and subtype and host vaccination status, sex, and age did not alter the transmission bottleneck.Conclusions. IAV is subject to a tight genetic bottleneck during transmission, which limits the onward propagation of newly arising variants. Tight bottlenecks appear to be intrinsic to the transmission process, as host and viral factors do not affect its size.