Quantitative metagenomics reveals fine-scale population dynamics across bacteria, archaea, and microbial eukaryotes in an estuarine-coastal continuum

Microbial communities are foundational to marine ecosystem function, yet their diversity is often obscured by broad taxonomic groupings and relative-abundance surveys that mask the dynamics of individual populations. This limitation is especially important across estuarine-coastal gradients, where microbial standing stocks, environmental conditions, and community composition vary sharply over space and time. Here, we used quantitative, genome-resolved metagenomics to examine microbial population dynamics across a one-year estuary-to-ocean transect spanning the Neuse River Estuary, Pamlico Sound, and adjacent North Atlantic shelf waters. Internal standard normalization enabled absolute abundance estimates for single-copy genes and metagenome-assembled genomes (MAGs), allowing individual populations to be tracked as genome equivalents per liter. Bacterial standing stocks were higher in estuarine waters, and communities varied primarily with salinity and season. We recovered 415 MAGs, including 386 bacterial genomes that represented, on average, 52% of bacterial genome equivalents, along with archaeal and eukaryotic representatives. Many abundant MAGs lacked close reference genomes, demonstrating that numerically important coastal populations remain poorly characterized. Genome-resolved abundances revealed pronounced niche partitioning among closely related taxa, including seasonal and spatial turnover of Synechococcus, Cyanobium, and Vulcanococcus populations associated with distinct pigment-defined cytometric groups. Rhodobacteraceae MAGs also showed population-specific correlations with picoeukaryotic MAGs, including a winter offshore Planktomarina population that reached 18% of total bacterial genome equivalents during a Micromonas-associated bloom. By providing absolute population abundances, this study transformed coastal microbiome surveys into numerical frameworks for resolving microbial population structure, ecological interactions, and biogeochemical relevance across dynamic environmental gradients. IMPORTANCEEstuarine and coastal waters contain diverse microbial communities that help regulate food webs and the cycling of carbon and nutrients, but many of the individual microbial populations responsible for these processes remain poorly understood. In this study, we examined bacteria, archaea, and small algae across the Neuse River Estuary, Pamlico Sound, and nearby coastal ocean waters over one year. By measuring the absolute abundance of individual microbial genomes, rather than only their relative proportions, we showed that closely related populations can have very different seasonal and spatial patterns. This was especially clear for cyanobacteria related to Synechococcus and heterotrophic bacteria in the family Rhodobacteraceae, which showed distinct population dynamics and associations with small algae. These results demonstrate how quantitative genome-resolved measurements can reveal hidden microbial population structure and improve our understanding of how microorganisms shape coastal ecosystem function.