Terrestrial connectivity, upstream aquatic history and seasonality shape bacterial community assembly within a large boreal aquatic network.

During transit from soils to the ocean, microbial communities are modified and re-assembled, generating complex patterns of ecological succession. The potential effect of upstream assembly on downstream microbial community composition is seldom considered within aquatic networks. Here, we reconstructed the microbial succession along a land-freshwater-estuary continuum within La Romaine river watershed in Northeastern Canada. We captured hydrological seasonality and differentiated the total and reactive community by sequencing both 16 S rRNA genes and transcripts. By examining how DNA- and RNA-based assemblages diverge and converge along the continuum, we inferred temporal shifts in the relative importance of assembly processes, with mass effects dominant in spring, and species selection becoming stronger in summer. The location of strongest selection within the network differed between seasons, suggesting that selection hotspots shift depending on hydrological conditions. The unreactive fraction (no/minor RNA contribution) was composed of taxa with diverse potential origins along the whole aquatic network, while the majority of the reactive pool (major RNA contribution) could be traced to soil/soilwater-derived taxa, which were distributed along the entire rank-abundance curve. Overall, our findings highlight the importance of considering upstream history, hydrological seasonality and the reactive microbial fraction to fully understand microbial community assembly on a network scale.