Ecological niches and beta diversity are fundamental concepts providing insight into the structure and functioning of marine ecosystems. Both concepts help in understanding how communities are distributed in different habitats and how marine ecosystems respond to environmental change. Here, the study brings a functional approach to the relationship between phytoplankton ecological niches and beta diversity. Phytoplankton community (from pico- to microphytoplankton) was addressed during a spring bloom of Phaeocystis globosa and diatoms, from the eastern English Channel (EEC) toward the southern North Sea (SNS) in 8 distinct water bodies, from late-April 2017 to mid-May 2017. An automated flow cytometer was used to discriminate phytoplankton by their optical properties at the single-cell level from continuous subsurface pumping marine waters, allowing the characterization of 11 phytoplankton functional groups (PFGs) from pico- to microphytoplankton. The spatial segregation of PFGs was performed from total abundance and Local Contribution to Beta Diversity (LCBD) calculations, from the most abundant to the marginal PFGs, through niche's overlap. Nanoeukaryotes (RedNano) associated with Phaeocystis globosa and picophytoplankton associated with Synechococcus spp. (OraPicoProk) were the less marginal PFGs. However, the low niche overlap between these groups revealed they have contrasting habitat affinity. While nanoeukaryotes prefer estuarine habitats or at the vicinity of an estuary, Synechococcus spp. was more likely along the coasts without freshwater influence. Picoeukaryotes with high chlorophyll-a content (RedPico III), coccolithophores (HsNano), and Pseudo-nitzschia-like (RedMicro I) were highly marginal revealing a patchy distribution. Finally, the beta regression predicted changes in community composition (i.e., LCBD values) influenced positively by temperature and the distance to the coast and negatively by salinity. The overall contribution of the PFGs to these changes (i.e., Species Contribution to Beta Diversity) was positively linked to their niche position and negatively related to their environmental tolerance.