The community composition of N2-fixing microorganisms (diazotrophs) was investigated in copepods (primarily Acartia spp.) in parallel to that of seawater in coastal waters off Denmark (Øresund) and New England, USA. The unicellular cyanobacterial diazotroph UCYN-A was detected from seawater and full-gut copepods, suggesting that the new N contributed by UCYN-A is directly transferred to higher trophic levels in these waters. Deltaproteobacterial and Cluster 3 nifH sequences were detected in > 1 μm seawater particles and full-gut copepods, suggesting that they associate with copepods primarily via feeding. The dominant communities in starved copepods were Vibrio spp. and related Gammaproteobacteria, suggesting they represent the most permanent diazotroph associations in the copepods. N2 fixation rates were up to 3.02 pmol N copepod−1 day−1. Although at a typical copepod density in estuarine waters, these volumetric rates are low; considering the small size of a copepod, these mesozooplanktonic crustaceans may serve as hotspots of N2 fixation, at 12.9–71.9 μmol N dm−3 copepod biomass day−1. Taken together, diazotroph associations range from more permanent attachments to copepod feeding on some groups. Similar diazotroph groups detected on the eastern and western Atlantic Ocean suggest that these associations are a general phenomenon and play a role in the coastal N cycles.