We present here the first detailed description of the seasonal patterns in bacterial community composition (BCC) in shelf waters off the Ría de Vigo (Spain), based on monthly samplings during 2 years. Moreover, we studied the relationship between bacterial and small-sized eukaryotic community composition to identify potential biotic interactions among components of these two communities. Bacterial operational taxonomic unit (OTU) richness and diversity systematically peaked in autumn–winter, likely related to low resource availability during this period. BCC showed seasonal and vertical patterns, withRhodobacteraceaeandFlavobacteriaceaefamilies dominating in surface waters, and SAR11 clade dominating at the base of the photic zone (30 m depth). BCC variability was significantly explained by environmental variables (e.g., temperature of water, solar radiation, or dissolved organic matter). Interestingly, a strong and significant correlation was found between BCC and small-sized eukaryotic community composition (ECC), which suggests that biotic interactions may play a major role as structuring factors of the microbial plankton in this productive area. In addition, co-occurrence network analyses revealed strong and significant, mostly positive, associations between bacteria and small-sized phytoplankton. Positive associations likely result from mutualistic relationships (e.g., betweenDinophyceaeandRhodobacteraceae), while some negative correlations suggest antagonistic interactions (e.g., betweenPseudo-nitzchiasp. and SAR11). These results support the key role of biotic interactions as structuring factors of the small-sized eukaryotic community, mostly driven by positive associations between small-sized phytoplankton and bacteria.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00248-022-02125-2.